59 results on '"Lori Sussel"'
Search Results
2. Modeling Hypoxia-Induced Neuropathies Using a Fast and Scalable Human Motor Neuron Differentiation System
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Laura I. Hudish, Andrew N. Bubak, Lori Sussel, J. Matthew Taliaferro, Taylor M. Triolo, Maria A. Nagel, David S. Lorberbaum, Christy S. Niemeyer, and Holger A. Russ
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0301 basic medicine ,diabetic neuropathies ,Neurite ,Induced Pluripotent Stem Cells ,Neuronal Outgrowth ,Action Potentials ,Disease ,Mitochondrion ,Biology ,Biochemistry ,cell compartments ,Cell Line ,Mitochondrial Proteins ,neurites ,03 medical and health sciences ,0302 clinical medicine ,Downregulation and upregulation ,Report ,Genetics ,medicine ,Humans ,RNA, Messenger ,fractionation ,Cells, Cultured ,Motor Neurons ,Messenger RNA ,hypoxia ,Correction ,stem cell-derived human motor neurons ,Cell Biology ,Motor neuron ,Hypoxia (medical) ,Cell Hypoxia ,Mitochondria ,Oxygen ,030104 developmental biology ,medicine.anatomical_structure ,Soma ,medicine.symptom ,Neuroscience ,030217 neurology & neurosurgery ,soma ,Developmental Biology - Abstract
Summary Human motor neuron (MN) diseases encompass a spectrum of disorders. A critical barrier to dissecting disease mechanisms is the lack of appropriate human MN models. Here, we describe a scalable, suspension-based differentiation system to generate functional human MN diseases in 3 weeks. Using this model, we translated recent findings that mRNA mis-localization plays a role in disease development to the human context by establishing a membrane-based system that allows efficient fractionation of MN cell soma and neurites. In response to hypoxia, used to mimic diabetic neuropathies, MNs upregulated mitochondrial transcripts in neurites; however, mitochondria were decreased. These data suggest that hypoxia may disrupt translation of mitochondrial mRNA, potentially leading to neurite damage and development of neuropathies. We report the development of a novel human MN model system to investigate mechanisms of disease affecting soma and/or neurites that facilitates the rapid generation and testing of patient-specific MN diseases., Graphical Abstract, Highlights • Differentiation of human pluripotent stem cells into motor neurons in 3 weeks • Global transcriptomes of motor neuron soma and neurite fractionations • Modeling of diabetic neuropathies by hypoxia reveals neurite-specific defects • Study suggests mRNA mis-localization as novel disease mechanism, Russ and colleagues present here a rapid, scalable, suspension-culture system for the efficient generation of functional human motor neurons from pluripotent stem cells. Using a soma and neurite fractionation approach, they reveal that mitochondrial transcripts, but not proteins, are neurite enriched under conditions modeling diabetic neuropathies. These findings suggest mRNA mis-localizations as a potential novel disease mechanism in motor neurons.
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- 2020
3. Zinc transporter 8 haploinsufficiency protects against beta cell dysfunction in type 1 diabetes by increasing mitochondrial respiration
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Yong Kyung Kim, Jay A. Walters, Nicole D. Moss, Kristen L. Wells, Ryan Sheridan, Jose G. Miranda, Richard K.P. Benninger, Laura L. Pyle, Richard M. O'Brien, Lori Sussel, and Howard W. Davidson
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Cell Biology ,Molecular Biology - Abstract
Zinc transporter 8 (ZnT8) is a major humoral target in human type 1 diabetes (T1D). Polymorphic variants of Slc30A8, which encodes ZnT8, are also associated with protection from type 2 diabetes (T2D). The current study examined whether ZnT8 might play a role beyond simply being a target of autoimmunity in the pathophysiology of T1D.The phenotypes of NOD mice with complete or partial global loss of ZnT8 were determined using a combination of disease incidence, histological, transcriptomic, and metabolic analyses.Unexpectedly, while complete loss of ZnT8 accelerated spontaneous T1D, heterozygosity was partially protective. In vivo and in vitro studies of ZnT8 deficient NOD.SCID mice suggested that the accelerated disease was due to more rampant autoimmunity. Conversely, beta cells in heterozygous animals uniquely displayed increased mitochondrial fitness under mild proinflammatory conditions.In pancreatic beta cells and immune cell populations, Zn
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- 2022
4. An Ultradian Notch in Beta-Cell Development
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Utpal B. Pajvani and Lori Sussel
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business.industry ,Ultradian Cycles ,Cellular differentiation ,Cell ,Organogenesis ,General Medicine ,Protein expression ,Cell biology ,medicine.anatomical_structure ,Medicine ,Signal transduction ,Beta cell ,business ,Ultradian rhythm - Abstract
Rhythmic Protein Expression and the Beta Cell A study of genetically manipulated mouse models has shown that ultradian cycles of expression of a particular protein influence specification of cell t...
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- 2020
5. Murine Perinatal β-Cell Proliferation and the Differentiation of Human Stem Cell–Derived Insulin-Expressing Cells Require NEUROD1
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Lori Sussel, Lina Sui, Ruth A. Singer, Dieter Egli, and Anthony I. Romer
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0301 basic medicine ,Cell Survival ,Endocrinology, Diabetes and Metabolism ,Transgene ,Human Embryonic Stem Cells ,Mice, Transgenic ,Nerve Tissue Proteins ,030209 endocrinology & metabolism ,Biology ,Cell Line ,Mice ,03 medical and health sciences ,0302 clinical medicine ,Insulin-Secreting Cells ,Insulin Secretion ,Basic Helix-Loop-Helix Transcription Factors ,Internal Medicine ,Animals ,Humans ,Progenitor cell ,Transcription factor ,Cell Proliferation ,Cell growth ,Gene Expression Regulation, Developmental ,Cell Differentiation ,Embryonic stem cell ,Cell biology ,030104 developmental biology ,Islet Studies ,Cell culture ,NEUROD1 ,Stem cell - Abstract
Inactivation of the β-cell transcription factor NEUROD1 causes diabetes in mice and humans. In this study, we uncovered novel functions of NEUROD1 during murine islet cell development and during the differentiation of human embryonic stem cells (HESCs) into insulin-producing cells. In mice, we determined that Neurod1 is required for perinatal proliferation of α- and β-cells. Surprisingly, apoptosis only makes a minor contribution to β-cell loss when Neurod1 is deleted. Inactivation of NEUROD1 in HESCs severely impaired their differentiation from pancreatic progenitors into insulin-expressing (HESC-β) cells; however, survival or proliferation was not affected at the time points analyzed. NEUROD1 was also required in HESC-β cells for the full activation of an essential β-cell transcription factor network. These data reveal conserved and distinct functions of NEUROD1 during mouse and human β-cell development and maturation, with important implications about the function of NEUROD1 in diabetes.
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- 2019
6. Groucho co-repressor proteins regulate β cell development and proliferation by repressing Foxa1 in the developing mouse pancreas
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Ruth A. Singer, Alexander Paul, Anila Narayana, Diana C. Garofalo, Alexandra Theis, and Lori Sussel
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0303 health sciences ,Co-Repressor Proteins ,Cell growth ,Cell ,Biology ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,medicine.anatomical_structure ,Downregulation and upregulation ,NEUROD1 ,medicine ,Ectopic expression ,Molecular Biology ,Psychological repression ,Transcription factor ,030217 neurology & neurosurgery ,030304 developmental biology ,Developmental Biology - Abstract
Groucho-related genes (GRGs) are transcriptional co-repressors that are crucial for many developmental processes. Several essential pancreatic transcription factors are capable of interacting with GRGs; however, the in vivo role of GRG-mediated transcriptional repression in pancreas development is still not well understood. In this study, we used complex mouse genetics and transcriptomic analyses to determine that GRG3 is essential for β cell development, and in the absence of Grg3 there is compensatory upregulation of Grg4. Grg3/4 double mutant mice have severe dysregulation of the pancreas gene program with ectopic expression of canonical liver genes and Foxa1, a master regulator of the liver program. Neurod1, an essential β cell transcription factor and predicted target of Foxa1, becomes downregulated in Grg3/4 mutants, resulting in reduced β cell proliferation, hyperglycemia, and early lethality. These findings uncover novel functions of GRG-mediated repression during pancreas development.
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- 2021
7. Single Molecule-based FliFISH Validates Radial and Heterogeneous Gene Expression Patterns in Pancreatic Islet β Cells
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Lori Sussel, Galya Orr, Cailin Dieter, Dehong Hu, Charles Ansong, and Fangjia Li
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0301 basic medicine ,endocrine system ,Maf Transcription Factors, Large ,Endocrinology, Diabetes and Metabolism ,medicine.medical_treatment ,Cell ,030209 endocrinology & metabolism ,RGS4 ,Transcriptome ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Insulin-Secreting Cells ,Gene expression ,Internal Medicine ,medicine ,Animals ,RNA-Seq ,Gene ,In Situ Hybridization, Fluorescence ,Urocortins ,geography ,geography.geographical_feature_category ,medicine.diagnostic_test ,biology ,Sequence Analysis, RNA ,Insulin ,Islet ,Cell biology ,030104 developmental biology ,medicine.anatomical_structure ,Islet Studies ,biology.protein ,Single-Cell Analysis ,RGS Proteins ,Fluorescence in situ hybridization - Abstract
Single cell RNA sequencing (scRNA-Seq) technologies have greatly enhanced our understanding of islet cell transcriptomes and have revealed the existence of β cell heterogeneity. However, comparison of scRNA-Seq datasets from different groups have highlighted inconsistencies in gene expression patterns, primarily due to variable detection of lower abundance transcripts. Furthermore, such analyses are unable to uncover the spatial organization of heterogeneous gene expression. Here we used fluctuation localization imaging-based fluorescence in situ hybridization (fliFISH) to quantify transcripts in single cells in mouse pancreatic islet sections. We compared the expression patterns of Insulin 2 (Ins2) with Mafa and Ucn3 – two genes expressed in β cells as they mature, as well as Rgs4 – a factor with variably reported expression in the islet. This approach accurately quantified transcripts across a wide range of expression levels - from single copies to over hundred copies per cell in one islet. Importantly, fliFISH allowed evaluation of transcript heterogeneity in the spatial context of an intact islet. These studies confirm the existence of a high degree of heterogeneous gene expression levels within the islet and highlight relative and radial expression patterns that likely reflect distinct β cell maturation states along the radial axis of the islet.
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- 2021
8. Islet Regeneration: Endogenous and Exogenous Approaches
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Fiona M. Docherty and Lori Sussel
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0301 basic medicine ,030209 endocrinology & metabolism ,Review ,Biology ,Catalysis ,Inorganic Chemistry ,lcsh:Chemistry ,03 medical and health sciences ,0302 clinical medicine ,Insulin-Secreting Cells ,Diabetes Mellitus ,Glucose homeostasis ,Animals ,Humans ,pancreas ,Physical and Theoretical Chemistry ,Induced pluripotent stem cell ,Beta (finance) ,Molecular Biology ,lcsh:QH301-705.5 ,Spectroscopy ,Cell Proliferation ,diabetes ,islet ,Regeneration (biology) ,Organic Chemistry ,Endogenous regeneration ,Transdifferentiation ,Cell Differentiation ,General Medicine ,beta cell ,Computer Science Applications ,Cell biology ,Transplantation ,030104 developmental biology ,lcsh:Biology (General) ,lcsh:QD1-999 ,regeneration ,Beta cell ,Stem Cell Transplantation - Abstract
Both type 1 and type 2 diabetes are characterized by a progressive loss of beta cell mass that contributes to impaired glucose homeostasis. Although an optimal treatment option would be to simply replace the lost cells, it is now well established that unlike many other organs, the adult pancreas has limited regenerative potential. For this reason, significant research efforts are focusing on methods to induce beta cell proliferation (replication of existing beta cells), promote beta cell formation from alternative endogenous cell sources (neogenesis), and/or generate beta cells from pluripotent stem cells. In this article, we will review (i) endogenous mechanisms of beta cell regeneration during steady state, stress and disease; (ii) efforts to stimulate endogenous regeneration and transdifferentiation; and (iii) exogenous methods of beta cell generation and transplantation.
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- 2021
9. mRNA Processing: An Emerging Frontier in the Regulation of Pancreatic β Cell Function
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Nicole D Moss and Lori Sussel
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0301 basic medicine ,lcsh:QH426-470 ,RNA methylation ,RNA-binding protein ,Review ,Biology ,Transcriptome ,03 medical and health sciences ,0302 clinical medicine ,Genetics ,Glucose homeostasis ,Genetics (clinical) ,Messenger RNA ,diabetes ,Alternative splicing ,RNA ,pancreatic islet ,Cell biology ,lcsh:Genetics ,beta cells ,030104 developmental biology ,RNA processing ,030220 oncology & carcinogenesis ,Molecular Medicine ,RNA binding proteins ,Function (biology) - Abstract
Robust endocrine cell function, particularly β cell function, is required to maintain blood glucose homeostasis. Diabetes can result from the loss or dysfunction of β cells. Despite decades of clinical and basic research, the precise regulation of β cell function and pathogenesis in diabetes remains incompletely understood. In this review, we highlight RNA processing of mRNAs as a rapidly emerging mechanism regulating β cell function and survival. RNA-binding proteins (RBPs) and RNA modifications are primed to be the next frontier to explain many of the poorly understood molecular processes that regulate β cell formation and function, and provide an exciting potential for the development of novel therapeutics. Here we outline the current understanding of β cell specific functions of several characterized RBPs, alternative splicing events, and transcriptome wide changes in RNA methylation. We also highlight several RBPs that are dysregulated in both Type 1 and Type 2 diabetes, and discuss remaining knowledge gaps in the field.
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- 2020
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10. A Notch in Time: Spatiotemporal Analysis of Notch Signaling during Pancreas Development
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Lori Sussel and David S. Lorberbaum
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JAG1 ,Cellular differentiation ,Regulator ,Notch signaling pathway ,Context (language use) ,Biology ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,0302 clinical medicine ,Spatio-Temporal Analysis ,medicine ,HES1 ,Molecular Biology ,Pancreas ,030304 developmental biology ,0303 health sciences ,Receptors, Notch ,Cell Differentiation ,Cell Biology ,Cell biology ,medicine.anatomical_structure ,Pancreatic progenitor cell ,030217 neurology & neurosurgery ,Developmental Biology ,Signal Transduction - Abstract
Notch signaling is a major regulator of pancreas development, yet how it precisely controls pancreatic cell fates has remained obscure. Seymour et al. (2020) use sophisticated Notch- based genetic tools to uncover highly context- and temporally-specific roles for DLL1, JAG1, and HES1 in regulating pancreatic progenitor cell growth and specification.
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- 2020
11. The role of islet lipid composition remodeling in regulation of beta-cell death via ADP-ribosyl-acceptor glycohydrolase ARH3 signaling in insulitis
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Jonàs Juan-Mateu, Ruichuan Yin, Farooq Syed, Yi Cui, Bobbie-Jo M. Webb-Robertson, Raghavendra G. Mirmira, Cailin Deiter, Jennifer E. Kyle, Julia Laskin, Lori Sussel, Ernesto S. Nakayasu, Carmella Evans-Molina, Kristin E. Burnum-Johnson, Michelle A. Guney, Charles Ansong, Decio L. Eizirik, Carrie D. Nicora, Thomas O. Metz, Galya Orr, and Dylan Sarbaugh
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0303 health sciences ,geography ,Chemokine ,geography.geographical_feature_category ,biology ,Antigen processing ,030209 endocrinology & metabolism ,Lipid metabolism ,medicine.disease ,Islet ,Cell biology ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Lysophosphatidylcholine ,chemistry ,Apoptosis ,medicine ,biology.protein ,Beta cell ,Insulitis ,030304 developmental biology - Abstract
SummaryLipids have been implicated as mediators of insulitis and β-cell death in type 1 diabetes development, but the mechanisms underlying this association are poorly understood. Here, we investigated the changes in islet/β-cell lipid composition using three models of insulitis: human islets and EndoC-βH1 β-cells treated with the cytokines IL-1β and IFN-γ, and islets from non-obese diabetic mice. Across all three models, lipidomic analyses showed a consistent change in abundance of the lysophosphatidylcholine, phosphatidylcholine and triacylglycerol species. We also showed that lysophosphatidylcholine and its biosynthetic enzyme PLA2G6 are enriched in murine islets. We determined that the ADP-ribosyl-acceptor glycohydrolase ARH3 is regulated by cytokines downstream of PLA2G6, which in turn regulates proteins involved in apoptosis, lipid metabolism, antigen processing and presentation and chemokines. ARH3 reduced cytokine-induced apoptosis, which may represent a negative feedback mechanism. Overall, these data show the importance of lipid metabolism in regulating β-cell death in type 1 diabetes.HighlightsLipidomics of 3 insulitis models revealed commonly regulated lipid classes.Identification of 35 proteins regulated by cytokines via PLA2G6 signaling.ARH3 reduces cytokine-induced apoptosis via PLA2G6 regulation.ARH3 regulates the levels of proteins related to insulitis and type 1 diabetes.
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- 2020
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12. Animal Models of Pancreas Development, Developmental Disorders, and Disease
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Fiona M. Docherty, Lori Sussel, and David S. Lorberbaum
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Pluripotent Stem Cells ,Organogenesis ,Blood sugar ,Acinar Cells ,Disease ,Biology ,Pancreas, Exocrine ,Article ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,medicine.anatomical_structure ,Models, Animal ,medicine ,Animals ,Humans ,Endocrine system ,030212 general & internal medicine ,Pancreas ,Induced pluripotent stem cell ,Enzyme secretion ,Homeostasis ,Hormone - Abstract
The pancreas is a glandular organ responsible for diverse homeostatic functions, including hormone production from the endocrine islet cells to regulate blood sugar levels and enzyme secretion from the exocrine acinar cells to facilitate food digestion. These pancreatic functions are essential for life; therefore, preserving pancreatic function is of utmost importance. Pancreas dysfunction can arise either from developmental disorders or adult onset disease, both of which are caused by defects in shared molecular pathways. In this chapter, we discuss what is known about the molecular mechanisms controlling pancreas development, how disruption of these mechanisms can lead to developmental defects and disease, and how essential pancreas functions can be modeled using human pluripotent stem cells. At the core of understanding of these molecular processes are animal model studies that continue to be essential for elucidating the mechanisms underlying human pancreatic functions and diseases.
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- 2020
13. Pancreatic β cell identity requires continual repression of non–β cell programs
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Lori Sussel, Stephen Kelly, Vincenzo Cirulli, Aaron S. Bender, Teresa L. Mastracci, Klaus H. Kaestner, Aristotelis Tsirigos, and Giselle Dominguez Gutierrez
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0301 basic medicine ,Regulation of gene expression ,Cell ,030209 endocrinology & metabolism ,Enteroendocrine cell ,General Medicine ,Biology ,Cell biology ,03 medical and health sciences ,030104 developmental biology ,0302 clinical medicine ,medicine.anatomical_structure ,Downregulation and upregulation ,medicine ,Gene ,Psychological repression ,Transcription factor ,Reprogramming - Abstract
Loss of β cell identity, the presence of polyhormonal cells, and reprogramming are emerging as important features of β cell dysfunction in patients with type 1 and type 2 diabetes. In this study, we have demonstrated that the transcription factor NKX2.2 is essential for the active maintenance of adult β cell identity as well as function. Deletion of Nkx2.2 in β cells caused rapid onset of a diabetic phenotype in mice that was attributed to loss of insulin and downregulation of many β cell functional genes. Concomitantly, NKX2.2-deficient murine β cells acquired non-β cell endocrine features, resulting in populations of completely reprogrammed cells and bihormonal cells that displayed hybrid endocrine cell morphological characteristics. Molecular analysis in mouse and human islets revealed that NKX2.2 is a conserved master regulatory protein that controls the acquisition and maintenance of a functional, monohormonal β cell identity by directly activating critical β cell genes and actively repressing genes that specify the alternative islet endocrine cell lineages. This study demonstrates the highly volatile nature of the β cell, indicating that acquiring and sustaining β cell identity and function requires not only active maintaining of the expression of genes involved in β cell function, but also continual repression of closely related endocrine gene programs.
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- 2016
14. Notch signaling dynamically regulates adult β cell proliferation and maturity
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Utpal B. Pajvani, Lori Sussel, Changyu Zhu, and Alberto Bartolomé
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0301 basic medicine ,Aging ,Cell ,Notch signaling pathway ,Mice, Transgenic ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Insulin-Secreting Cells ,Glucose Intolerance ,Insulin Secretion ,medicine ,Endocrine system ,Animals ,Obesity ,Cell Proliferation ,geography ,geography.geographical_feature_category ,Receptors, Notch ,Cell growth ,Chemistry ,Proliferative capacity ,Embryogenesis ,General Medicine ,Metabolism ,Islet ,Cell biology ,030104 developmental biology ,medicine.anatomical_structure ,Glucose ,030220 oncology & carcinogenesis ,Research Article ,Signal Transduction - Abstract
Notch signaling regulates differentiation of the pancreatic endocrine lineage during embryogenesis, but the role of Notch in mature β cells is unclear. We found that islets derived from lean mice show modest β cell Notch activity, which increases in obesity and in response to high glucose. This response appeared maladaptive, as mice with β cell-specific-deficient Notch transcriptional activity showed improved glucose tolerance when subjected to high-fat diet feeding. Conversely, mice with β cell-specific Notch gain of function (β-NICD) had a progressive loss of β cell maturity, due to proteasomal degradation of MafA, leading to impaired glucose-stimulated insulin secretion and glucose intolerance with aging or obesity. Surprisingly, Notch-active β cells had increased proliferative capacity, leading to increased but dysfunctional β cell mass. These studies demonstrate a dynamic role for Notch in developed β cells for simultaneously regulating β cell function and proliferation.
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- 2018
15. Inherent Beta Cell Dysfunction Contributes to Autoimmune Susceptibility
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Howard W. Davidson, Yong Kyung Kim, and Lori Sussel
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0301 basic medicine ,Cell type ,type 1 diabetes ,medicine.medical_treatment ,lcsh:QR1-502 ,Review ,Mitochondrion ,Biochemistry ,lcsh:Microbiology ,Epitopes ,03 medical and health sciences ,0302 clinical medicine ,Insulin-Secreting Cells ,Insulin Secretion ,Autophagy ,medicine ,Animals ,Humans ,Glucose homeostasis ,Secretion ,Beta (finance) ,Molecular Biology ,Cellular Senescence ,Type 1 diabetes ,Chemistry ,Insulin ,autoimmunity ,Mitophagy ,pancreatic islet ,medicine.disease ,beta cell ,Mitochondria ,Cell biology ,Diabetes Mellitus, Type 1 ,030104 developmental biology ,Beta cell ,030217 neurology & neurosurgery - Abstract
The pancreatic beta cell is a highly specialized cell type whose primary function is to secrete insulin in response to nutrients to maintain glucose homeostasis in the body. As such, the beta cell has developed unique metabolic characteristics to achieve functionality; in healthy beta cells, the majority of glucose-derived carbons are oxidized and enter the mitochondria in the form of pyruvate. The pyruvate is subsequently metabolized to induce mitochondrial ATP and trigger the downstream insulin secretion response. Thus, in beta cells, mitochondria play a pivotal role in regulating glucose stimulated insulin secretion (GSIS). In type 2 diabetes (T2D), mitochondrial impairment has been shown to play an important role in beta cell dysfunction and loss. In type 1 diabetes (T1D), autoimmunity is the primary trigger of beta cell loss; however, there is accumulating evidence that intrinsic mitochondrial defects could contribute to beta cell susceptibility during proinflammatory conditions. Furthermore, there is speculation that dysfunctional mitochondrial responses could contribute to the formation of autoantigens. In this review, we provide an overview of mitochondrial function in the beta cells, and discuss potential mechanisms by which mitochondrial dysfunction may contribute to T1D pathogenesis.
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- 2021
16. βlinc1 encodes a long noncoding RNA that regulates islet β-cell formation and function
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Dina A. Balderes, Luis Arnes, Ildem Akerman, Jorge Ferrer, and Lori Sussel
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0301 basic medicine ,β cell ,Transcriptome ,Gene Knockout Techniques ,Mice ,Insulin-Secreting Cells ,Glucose homeostasis ,Genetics & Heredity ,Genetics ,geography.geographical_feature_category ,Gene Expression Regulation, Developmental ,11 Medical And Health Sciences ,Non-coding RNA ,Islet ,Long non-coding RNA ,Cell biology ,GENOME ,RNA, Long Noncoding ,Beta cell ,Life Sciences & Biomedicine ,EXPRESSION ,GENES ,Endocrine System ,Biology ,Cell Line ,17 Psychology And Cognitive Sciences ,Research Communication ,03 medical and health sciences ,REVEALS ,Glucose Intolerance ,Animals ,Humans ,long noncoding RNA ,TRANSCRIPTOME ,Gene ,Transcription factor ,NKX2.2 ,geography ,Science & Technology ,SEQUENCES ,Cell Biology ,RESEARCH RESOURCE ,06 Biological Sciences ,PANCREAS DEVELOPMENT ,beta cell ,Mice, Inbred C57BL ,FACTORIES ,030104 developmental biology ,Transcription Factors ,Developmental Biology - Abstract
Pancreatic β cells are responsible for maintaining glucose homeostasis; their absence or malfunction results in diabetes mellitus. Although there is evidence that long noncoding RNAs (lncRNAs) play important roles in development and disease, none have been investigated in vivo in the context of pancreas development. In this study, we demonstrate that βlinc1 (β-cell long intergenic noncoding RNA 1), a conserved lncRNA, is necessary for the specification and function of insulin-producing β cells through the coordinated regulation of a number of islet-specific transcription factors located in the genomic vicinity of βlinc1. Furthermore, deletion of βlinc1 results in defective islet development and disruption of glucose homeostasis in adult mice.
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- 2016
17. Unique functions of Gata4 in mouse liver induction and heart development
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Lori Sussel, Matthew J. Borok, and Virginia E. Papaioannou
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0301 basic medicine ,endocrine system ,DNA, Complementary ,Mesenchyme ,Septum transversum ,Morphogenesis ,Biology ,Article ,03 medical and health sciences ,Mice ,0302 clinical medicine ,medicine ,Animals ,Humans ,Transcription factor ,Molecular Biology ,reproductive and urinary physiology ,Genetics ,GATA6 ,Heart development ,GATA4 ,Heart ,Cell Biology ,respiratory system ,Embryonic stem cell ,Cell biology ,GATA4 Transcription Factor ,030104 developmental biology ,medicine.anatomical_structure ,HEK293 Cells ,Liver ,030220 oncology & carcinogenesis ,embryonic structures ,cardiovascular system ,Developmental Biology - Abstract
Gata4 and Gata6 are closely related transcription factors that are essential for the development of a number of embryonic tissues. While they have nearly identical DNA-binding domains and similar patterns of expression, Gata4 and Gata6 null embryos have strikingly different embryonic lethal phenotypes. To determine whether the lack of redundancy is due to differences in protein function or Gata4 and Gata6 expression domains, we generated mice that contained the Gata6 cDNA in place of the Gata4 genomic locus. Gata4(Gata6/Gata6) embryos survived through embryonic day (E)12.5 and successfully underwent ventral folding morphogenesis, demonstrating that Gata6 is able to replace Gata4 function in extraembryonic tissues. Surprisingly, Gata6 is unable to replace Gata4 function in the septum transversum mesenchyme or the epicardium, leading to liver agenesis and lethal heart defects in Gata4(Gata6/Gata6) embryos. These studies suggest that Gata4 has evolved distinct functions in the development of these tissues that cannot be performed by Gata6, even when it is provided in the identical expression domain. Our work has important implications for the respective mechanisms of Gata function during development, as well as the functional evolution of these essential transcription factors.
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- 2016
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18. The Long Noncoding RNA Paupar Modulates PAX6 Regulatory Activities to Promote Alpha Cell Development and Function
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Ruth A. Singer, Yuqian Gao, Kristin E. Burnum-Johnson, Michelle A. Guney, Jiguang Wang, Charles Ansong, Yi Cui, Galya Orr, Luis Arnes, Raul Rabadan, and Lori Sussel
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0301 basic medicine ,endocrine system ,PAX6 Transcription Factor ,Physiology ,Biology ,Article ,Alpha cell ,Transcriptome ,Mice ,03 medical and health sciences ,0302 clinical medicine ,Diabetes Mellitus ,medicine ,Animals ,Humans ,Glucose homeostasis ,Molecular Biology ,Transcription factor ,Cells, Cultured ,Mice, Knockout ,Cell growth ,Gene Expression Profiling ,Pancreatic islets ,Glucagon secretion ,Cell Biology ,Embryo, Mammalian ,Glucagon ,Cell biology ,Mice, Inbred C57BL ,Glucose ,030104 developmental biology ,medicine.anatomical_structure ,Glucagon-Secreting Cells ,RNA, Long Noncoding ,PAX6 ,030217 neurology & neurosurgery - Abstract
Many studies have highlighted the role of dysregulated glucagon secretion in the etiology of hyperglycemia and diabetes. Accordingly, understanding the mechanisms underlying pancreatic islet α cell development and function has important implications for the discovery of new therapies for diabetes. In this study, comparative transcriptome analyses between embryonic mouse pancreas and adult mouse islets identified several pancreatic lncRNAs that lie in close proximity to essential pancreatic transcription factors, including the Pax6-associated lncRNA Paupar. We demonstrate that Paupar is enriched in glucagon-producing α cells where it promotes the alternative splicing of Pax6 to an isoform required for activation of essential α cell genes. Consistently, deletion of Paupar in mice resulted in dysregulation of PAX6 α cell target genes and corresponding α cell dysfunction, including blunted glucagon secretion. These findings illustrate a distinct mechanism by which a pancreatic lncRNA can coordinate glucose homeostasis by cell-specific regulation of a broadly expressed transcription factor.
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- 2019
19. FISHing for β Cells
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David S. Lorberbaum, Laura I. Hudish, and Lori Sussel
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endocrine system ,endocrine system diseases ,Population ,Cell ,Mice, Transgenic ,Enteroendocrine cell ,Biology ,Article ,General Biochemistry, Genetics and Molecular Biology ,Diabetes Mellitus, Experimental ,Genetic Heterogeneity ,Islets of Langerhans ,03 medical and health sciences ,0302 clinical medicine ,Insulin-Secreting Cells ,Insulin Secretion ,medicine ,Animals ,Insulin ,education ,Pancreas ,Molecular Biology ,030304 developmental biology ,0303 health sciences ,geography ,education.field_of_study ,geography.geographical_feature_category ,Cell Biology ,Islet ,Cell biology ,Glucose ,medicine.anatomical_structure ,030217 neurology & neurosurgery ,Function (biology) ,Proinsulin ,Developmental Biology - Abstract
Pancreatic beta cells have been shown to be heterogeneous at multiple levels. However, spatially interrogating transcriptional heterogeneity in the intact tissue has been challenging. Here, we developed an optimized protocol for single-molecule transcript imaging in the intact pancreas and used it to identify a sub-population of "extreme" beta cells with elevated mRNA levels of insulin and other secretory genes. Extreme beta cells contain higher ribosomal and proinsulin content but lower levels of insulin protein in fasted states, suggesting they may be tuned for basal insulin secretion. They exhibit a distinctive intra-cellular polarization pattern, with elevated mRNA concentrations in an apical ER-enriched compartment, distinct from the localization of nascent and mature proteins. The proportion of extreme cells increases in db/db diabetic mice, potentially facilitating the required increase in basal insulin. Our results thus highlight a sub-population of beta cells that may carry distinct functional roles along physiological and pathological timescales.
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- 2019
20. Gotta Have GATA for Human Pancreas Development
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Lori Sussel and David S. Lorberbaum
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0301 basic medicine ,Pluripotent Stem Cells ,medicine.medical_specialty ,endocrine system ,Cellular differentiation ,Cell ,pancreatic agenesis ,Mutation, Missense ,β cell ,030209 endocrinology & metabolism ,Biology ,medicine.disease_cause ,Article ,03 medical and health sciences ,Mice ,0302 clinical medicine ,GATA6 ,stem cells ,Internal medicine ,GATA6 Transcription Factor ,Insulin-Secreting Cells ,Genetics ,medicine ,retinoic acid ,Animals ,Humans ,Pancreas ,Mutation ,Gene Expression Regulation, Developmental ,Cell Differentiation ,Cell Biology ,030104 developmental biology ,medicine.anatomical_structure ,Endocrinology ,Cancer research ,Molecular Medicine ,Stem cell ,definitive endoderm ,Haploinsufficiency ,Definitive endoderm - Abstract
Summary Induced pluripotent stem cells were created from a pancreas agenesis patient with a mutation in GATA6. Using genome-editing technology, additional stem cell lines with mutations in both GATA6 alleles were generated and demonstrated a severe block in definitive endoderm induction, which could be rescued by re-expression of several different GATA family members. Using the endodermal progenitor stem cell culture system to bypass the developmental block at the endoderm stage, cell lines with mutations in one or both GATA6 alleles could be differentiated into β-like cells but with reduced efficiency. Use of suboptimal doses of retinoic acid during pancreas specification revealed a more severe phenotype, more closely mimicking the patient’s disease. GATA6 mutant β-like cells fail to secrete insulin upon glucose stimulation and demonstrate defective insulin processing. These data show that GATA6 plays a critical role in endoderm and pancreas specification and β-like cell functionality in humans., Highlights • GATA6 is required for definitive endoderm specification in human ES/iPS cells • Bypassing the endoderm defect allows GATA6 mutants to generate β-like cells • Suboptimal retinoic acid signaling blocks pancreas specification in GATA6 mutants • GATA6 is critical for human β cell function in vitro, In this study, Gadue and colleagues studied GATA6 mutant pluripotent stem cells and demonstrate that GATA6 is necessary for human definitive endoderm specification. GATA6 also plays an important role in pancreas specification which could be partially bypassed by retinoic acid signaling. Furthermore, derivative β cells lacked glucose responsiveness in vitro, showing that GATA6 is vital for appropriate human pancreas development.
- Published
- 2017
21. Genetic evidence that Nkx2.2 and Pdgfra are major determinants of the timing of oligodendrocyte differentiation in the developing CNS
- Author
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Qiang Zhu, Kang Zheng, Hong Li, YiPing Chen, Teresa L. Mastracci, Michael Wegner, Lori Sussel, Xiaofeng Zhao, Hao Huang, Mengsheng Qiu, and Zunyi Zhang
- Subjects
Central Nervous System ,Receptor, Platelet-Derived Growth Factor alpha ,Time Factors ,Cellular differentiation ,Molecular Sequence Data ,PDGFRA ,Biology ,Mice ,Growth factor receptor ,Cell Movement ,medicine ,Animals ,Cell Lineage ,Promoter Regions, Genetic ,Molecular Biology ,Transcription factor ,Myelin Sheath ,Cell Proliferation ,Homeodomain Proteins ,Genetics ,Base Sequence ,Stem Cells ,Oligodendrocyte differentiation ,Gene Expression Regulation, Developmental ,Cell Differentiation ,Zebrafish Proteins ,Stem Cells and Regeneration ,Oligodendrocyte ,Cell biology ,Oligodendroglia ,Homeobox Protein Nkx-2.2 ,Phenotype ,medicine.anatomical_structure ,Mutation ,embryonic structures ,Homeobox ,Stem cell ,Protein Binding ,Transcription Factors ,Developmental Biology - Abstract
In the central nervous system (CNS), oligodendrocyte maturation and axonal myelination occur on a predictable schedule, but the underlying timing mechanisms are largely unknown. In the present study, we demonstrate that Nkx2.2 homeodomain transcription factor is a key regulator for the timing of oligodendrocyte differentiation during development. Whereas induced expression of Nkx2.2 in early oligodendrocyte precursor cells (OPCs) causes precocious differentiation of oligodendrocytes, conditional ablation of Nkx2.2 temporally delays oligodendrocyte maturation. Moreover, Nkx2.2 can directly bind to the promoter of platelet-derived growth factor receptor alpha (Pdgfra) and repress its gene expression. Genetic ablation of Pdgfra mimics the effect of Nkx2.2 overexpression in accelerating OPC differentiation in the developing spinal cord. Together, our findings strongly suggest that Nkx2.2 functions as a major ‘switch’ to turn off Pdgfra signaling in OPCs and initiate the intrinsic program for oligodendrocyte differentiation.
- Published
- 2014
22. Author response: Genetic evidence that Nkx2.2 acts primarily downstream of Neurog3 in pancreatic endocrine lineage development
- Author
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Giselle Dominguez Gutierrez, David S. Lorberbaum, Lori Sussel, Ruth A. Singer, Kevin A Fischer, and Angela J. Churchill
- Subjects
Lineage (genetic) ,Downstream (manufacturing) ,Endocrine system ,Biology ,Cell biology - Published
- 2016
23. Nkx2.2:Cre knock-in mouse line: A novel tool for pancreas- and CNS-specific gene deletion
- Author
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Mark A. Magnuson, Lori Sussel, and Dina A. Balderes
- Subjects
Cell type ,Hindbrain ,Enteroendocrine cell ,Cell Biology ,respiratory system ,Cell fate determination ,Biology ,Embryonic stem cell ,Molecular biology ,Endocrinology ,medicine.anatomical_structure ,stomatognathic system ,embryonic structures ,cardiovascular system ,Genetics ,medicine ,Transcriptional regulation ,Progenitor cell ,Pancreas - Abstract
Nkx2.2 is a homeodomain-containing transcriptional regulator necessary for the appropriate differentiation of ventral neuronal populations in the spinal cord and hindbrain, and endocrine cell populations in the pancreas and intestine. In each tissue, Nkx2.2 inactivation leads to reciprocal cell fate alterations. To confirm the cell fate changes are due to respecification of Nkx2.2-expressing progenitors and to provide a novel tool for lineage tracing in the pancreas and CNS, we generated an Nkx2.2:Cre mouse line by knocking in a Cre-EGFP cassette into the Nkx2.2 genomic locus and inactivating endogenous Nkx2.2. The R26R-CAG-LSL-tdTomato reporter was used to monitor the specificity and efficiency of Nkx2.2:Cre activity; the tomato reporter faithfully recapitulated endogenous Nkx2.2 expression and could be detected as early as embryonic day (e) 9.25 in the developing CNS and was initiated shortly thereafter at e9.5 in the pancreas. Lineage analyses in the CNS confirmed the cell populations thought to be derived from Nkx2.2-expressing progenitor domains. Furthermore, lineage studies verified Nkx2.2 expression in the earliest pancreatic progenitors that give rise to all cell types of the pancreas; however they also revealed more robust Cre activity in the dorsal versus ventral pancreas. Thus, the Nkx2.2:Cre line provides a novel tool for gene manipulations in the CNS and pancreas.
- Published
- 2013
24. Generation of mice encoding a conditional allele of Nkx2.2
- Author
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Teresa L. Mastracci, Chyuan-Sheng Lin, and Lori Sussel
- Subjects
Cell type ,Fluorescent Antibody Technique ,Cre recombinase ,Mice, Transgenic ,In situ hybridization ,Biology ,Real-Time Polymerase Chain Reaction ,Article ,Mice ,stomatognathic system ,Genetics ,medicine ,Animals ,Allele ,Pancreas ,Transcription factor ,In Situ Hybridization ,DNA Primers ,Homeodomain Proteins ,Integrases ,Zebrafish Proteins ,respiratory system ,Null allele ,Molecular biology ,Cell biology ,Homeobox Protein Nkx-2.2 ,medicine.anatomical_structure ,embryonic structures ,cardiovascular system ,Homeobox ,Animal Science and Zoology ,Agronomy and Crop Science ,Gene Deletion ,Transcription Factors ,Biotechnology - Abstract
Nkx2.2 is a homeobox transcription factor that is expressed in the pancreas, intestine and central nervous system (CNS) during embryogenesis and in the adult. In mice, global deletion of Nkx2.2 results in cell mis-specification in each of the tissues where it is expressed, and the null mice die as neonates with severe hyperglycemia. Although a whole body knockout demonstrates the importance of Nkx2.2 in cell specification and postnatal viability, it precludes assessment of the cell-autonomous and postnatal functions of Nkx2.2. In this study we report the generation and functional characterization of mice encoding a conditional allele of Nkx2.2. We demonstrate the functional integrity of the conditional Nkx2.2 allele and report successful in vivo deletion using a pancreas-specific Cre recombinase. The pancreas-specific deletion of Nkx2.2 results in similar defects found in the Nkx2.2 null pancreas and the mice die shortly after birth, demonstrating that the neonatal lethality associated with the null allele is caused by pancreatic islet dysfunction. The generation of a conditional Nkx2.2 allele provides an important tool for identifying the cell-autonomous and postnatal activities of Nkx2.2 in establishing and maintaining cell type identities and functions in the pancreas, intestine and CNS.
- Published
- 2013
25. Genetic evidence that Nkx2.2 acts primarily downstream of Neurog3 in pancreatic endocrine lineage development
- Author
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Giselle Dominguez Gutierrez, Kevin A Fischer, Ruth A. Singer, Angela J. Churchill, David S. Lorberbaum, and Lori Sussel
- Subjects
0301 basic medicine ,Mouse ,Cellular differentiation ,Mice ,0302 clinical medicine ,Insulin-Secreting Cells ,Basic Helix-Loop-Helix Transcription Factors ,Biology (General) ,Genetics ,Mice, Knockout ,Transcriptional networks ,education.field_of_study ,geography.geographical_feature_category ,General Neuroscience ,Cell Differentiation ,General Medicine ,respiratory system ,Islet ,Cell biology ,Homeobox Protein Nkx-2.2 ,embryonic structures ,cardiovascular system ,Medicine ,Stem cell ,Research Article ,endocrine system ,QH301-705.5 ,Science ,Population ,Nerve Tissue Proteins ,Biology ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,Islets of Langerhans ,stomatognathic system ,Animals ,Progenitor cell ,education ,Transcription factor ,Progenitor ,Homeodomain Proteins ,geography ,General Immunology and Microbiology ,Zebrafish Proteins ,pancreatic islet ,beta cells ,030104 developmental biology ,Developmental Biology and Stem Cells ,Developmental biology ,030217 neurology & neurosurgery ,Transcription Factors - Abstract
Many pancreatic transcription factors that are essential for islet cell differentiation have been well characterized; however, because they are often expressed in several different cell populations, their functional hierarchy remains unclear. To parse out the spatiotemporal regulation of islet cell differentiation, we used a Neurog3-Cre allele to ablate Nkx2.2, one of the earliest and most broadly expressed islet transcription factors, specifically in the Neurog3+ endocrine progenitor lineage (Nkx2.2△endo). Remarkably, many essential components of the β cell transcriptional network that were down-regulated in the Nkx2.2KO mice, were maintained in the Nkx2.2△endo mice - yet the Nkx2.2△endo mice displayed defective β cell differentiation and recapitulated the Nkx2.2KO phenotype. This suggests that Nkx2.2 is not only required in the early pancreatic progenitors, but has additional essential activities within the endocrine progenitor population. Consistently, we demonstrate Nkx2.2 functions as an integral component of a modular regulatory program to correctly specify pancreatic islet cell fates. DOI: http://dx.doi.org/10.7554/eLife.20010.001
- Published
- 2016
26. Lmx1a functions in intestinal serotonin-producing enterochromaffin cells downstream of Nkx2.2
- Author
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Dina A. Balderes, Teresa L. Mastracci, José M. Dias, Diana C. Garofalo, Lori Sussel, Thomas Perlmann, Johan Ericson, and Stefanie Gross
- Subjects
0301 basic medicine ,medicine.medical_specialty ,TPH1 ,Enteroendocrine cell ,Biology ,digestive system ,Secretin ,Cell biology ,03 medical and health sciences ,030104 developmental biology ,Endocrinology ,Internal medicine ,Enterochromaffin cell ,medicine ,Progenitor cell ,Stem cell ,Molecular Biology ,Developmental Biology ,Gastrin ,Cholecystokinin - Abstract
The intestinal hormone-producing cells represent the largest endocrine system in the body; however, there is still remarkably little known about enteroendocrine cell type specification in the embryo and adult. We analyzed stage and cell-type specific deletions of Nkx2.2 and its functional domains to characterize its precise role in the development and maintenance of enteroendocrine cell lineages in the duodenum and colon. Although Nkx2.2 regulates enteroendocrine cell specification in the duodenum at all stages examined, Nkx2.2 controls the differentiation of progressively fewer enteroendocrine cell populations when deleted from Neurogenin 3 (Ngn3)+ progenitor cells or in the adult duodenum. During embryonic development Nkx2.2 regulates all enteroendocrine cell types, except gastrin and preproglucagon. In the developing Ngn3-expressing enteroendocrine progenitor cells, Nkx2.2 is also not required for the specification of neuropeptide Y and vasoactive intestinal polypeptide, indicating that a subset of these cell populations derive from an Nkx2.2-independent lineage. In the adult duodenum, Nkx2.2 also becomes dispensable for cholecystokinin and secretin production. In all stages and Nkx2.2 mutant conditions, serotonin-producing enterochromaffin cells were the most severely reduced enteroendocrine lineage in the duodenum and the colon. We determined that the transcription factor Lmx1a is expressed in enterochromaffin cells and functions downstream of Nkx2.2. Consistently, Lmx1a-deficient mice have reduced expression of Tph1, the rate-limiting enzyme for serotonin biosynthesis. These data clarify the function of Nkx2.2 in the specification and homeostatic maintenance of enteroendocrine populations, and identify Lmx1a as a novel enterochromaffin cell marker that is also essential for the production of the serotonin biosynthetic enzyme Tph1.
- Published
- 2016
27. The endocrine pancreas: insights into development, differentiation, and diabetes
- Author
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Lori Sussel and Teresa L. Mastracci
- Subjects
Ductal cells ,Cellular differentiation ,Cell Biology ,Biology ,Cell fate determination ,Embryonic stem cell ,Cell biology ,Diabetes mellitus genetics ,medicine.anatomical_structure ,Immunology ,medicine ,Stem cell ,Progenitor cell ,Pancreas ,Molecular Biology ,Developmental Biology - Abstract
In the developing embryo, appropriate patterning of the endoderm fated to become pancreas requires the spatial and temporal coordination of soluble factors secreted by the surrounding tissues. Once pancreatic progenitor cells are specified in the developing gut tube epithelium, epithelial-mesenchymal interactions, as well as a cascade of transcription factors, subsequently delineate three distinct lineages, including endocrine, exocrine, and ductal cells. Simultaneous morphological changes, including branching, vascularization, and proximal organ development, also influence the process of specification and differentiation. Decades of research using mouse genetics have uncovered many of the key factors involved in pancreatic cell fate decisions. When pancreas development or islet cell functions go awry, due to mutations in genes important for proper organogenesis and development, the result can lead to a common pancreatic affliction, diabetes mellitus. Current treatments for diabetes are adequate but not curative. Therefore, researchers are utilizing the current understanding of normal embryonic pancreas development in vivo, to direct embryonic stem cells toward a pancreatic fate with the goal of transplanting these in vitro generated 'islets' into patients. Mimicking development in vitro has proven difficult; however, significant progress has been made and the current differentiation protocols are becoming more efficient. The continued partnership between developmental biologists and stem cell researchers will guarantee that the in vitro generation of insulin-producing β cells is a possible therapeutic option for the treatment of diabetes.
- Published
- 2012
28. The L6 domain tetraspanin Tm4sf4 regulates endocrine pancreas differentiation and directed cell migration
- Author
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Christopher W. Johnson, Ruth A. Singer, Dina A. Balderes, Keith R. Anderson, Lori Sussel, Kristin Bruk Artinger, and Laura Hernandez-Lagunas
- Subjects
rho GTP-Binding Proteins ,Cell type ,Cellular differentiation ,Molecular Sequence Data ,Nerve Tissue Proteins ,Cell fate determination ,Cell Line ,Islets of Langerhans ,Mice ,Tetraspanin ,Cell Movement ,Basic Helix-Loop-Helix Transcription Factors ,Animals ,Humans ,Amino Acid Sequence ,Progenitor cell ,Cell adhesion ,Molecular Biology ,Zebrafish ,Research Articles ,Homeodomain Proteins ,Mice, Knockout ,Membrane Glycoproteins ,biology ,Stem Cells ,Gene Expression Regulation, Developmental ,Membrane Proteins ,Nuclear Proteins ,Cell Differentiation ,Cell migration ,Zebrafish Proteins ,biology.organism_classification ,Ghrelin ,Cell biology ,Homeobox Protein Nkx-2.2 ,embryonic structures ,Sequence Alignment ,Transcription Factors ,Developmental Biology - Abstract
The homeodomain transcription factor Nkx2.2 is essential for pancreatic development and islet cell type differentiation. We have identified Tm4sf4, an L6 domain tetraspanin family member, as a transcriptional target of Nkx2.2 that is greatly upregulated during pancreas development in Nkx2.2–/– mice. Tetraspanins and L6 domain proteins recruit other membrane receptors to form active signaling centers that coordinate processes such as cell adhesion, migration and differentiation. In this study, we determined that Tm4sf4 is localized to the ductal epithelial compartment and is prominent in the Ngn3+ islet progenitor cells. We also established that pancreatic tm4sf4 expression and regulation by Nkx2.2 is conserved during zebrafish development. Loss-of-function studies in zebrafish revealed that tm4sf4 inhibits α and β cell specification, but is necessary for ε cell fates. Thus, Tm4sf4 functional output opposes that of Nkx2.2. Further investigation of how Tm4sf4 functions at the cellular level in vitro showed that Tm4sf4 inhibits Rho-activated cell migration and actin organization in a ROCK-independent fashion. We propose that the primary role of Nkx2.2 is to inhibit Tm4sf4 in endocrine progenitor cells, allowing for delamination, migration and/or appropriate cell fate decisions. Identification of a role for Tm4sf4 during endocrine differentiation provides insight into islet progenitor cell behaviors and potential targetable regenerative mechanisms.
- Published
- 2011
29. A homeodomain feedback circuit underlies step-function interpretation of a Shh morphogen gradient during ventral neural patterning
- Author
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Sanja Kurdija, Madelen Lek, José M. Dias, Johan Ericson, Christopher W. Uhde, Ulrika Marklund, Hans-Henning Arnold, John L.R. Rubenstein, Michael P. Matise, Lori Sussel, Thomas M. Jessell, and Qiubo Lei
- Subjects
Time Factors ,animal structures ,PAX6 Transcription Factor ,Biology ,Models, Biological ,Mice ,Mice, Neurologic Mutants ,Animals ,Paired Box Transcription Factors ,Hedgehog Proteins ,Eye Proteins ,Molecular Biology ,Transcription factor ,Body Patterning ,Floor plate ,Progenitor ,Feedback, Physiological ,Homeodomain Proteins ,Motor Neurons ,Neurons ,Genetics ,Regulation of gene expression ,Gene Expression Profiling ,Stem Cells ,Gene Expression Regulation, Developmental ,Cell Differentiation ,Cell biology ,Repressor Proteins ,embryonic structures ,Homeobox ,PAX6 ,Signal transduction ,Signal Transduction ,Developmental Biology ,Morphogen - Abstract
The deployment of morphogen gradients is a core strategy to establish cell diversity in developing tissues, but little is known about how small differences in the concentration of extracellular signals are translated into robust patterning output in responding cells. We have examined the activity of homeodomain proteins, which are presumed to operate downstream of graded Shh signaling in neural patterning, and describe a feedback circuit between the Shh pathway and homeodomain transcription factors that establishes non-graded regulation of Shh signaling activity. Nkx2 proteins intrinsically strengthen Shh responses in a feed-forward amplification and are required for ventral floor plate and p3 progenitor fates. Conversely, Pax6 has an opposing function to antagonize Shh signaling, which provides intrinsic resistance to Shh responses and is important to constrain the inductive capacity of the Shh gradient over time. Our data further suggest that patterning of floor plate cells and p3 progenitors is gated by a temporal switch in neuronal potential, rather than by different Shh concentrations. These data establish that dynamic, non-graded changes in responding cells are essential for Shh morphogen interpretation, and provide a rationale to explain mechanistically the phenomenon of cellular memory of morphogen exposure.
- Published
- 2010
30. Islet β-Cell-Specific MafA Transcription Requires the 5′-Flanking Conserved Region 3 Control Domain
- Author
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Chad S. Hunter, Eva Henderson, Roland Stein, Min Guo, Takeshi Ogihara, Isabella Artner, Lori Sussel, Raghavendra G. Mirmira, Beatriz Sosa-Pineda, Jeffrey C. Raum, and Lynda Elghazi
- Subjects
Maf Transcription Factors, Large ,PAX6 Transcription Factor ,Transcription, Genetic ,5' Flanking Region ,Molecular Sequence Data ,5' flanking region ,Repressor ,Mice, Transgenic ,Nerve Tissue Proteins ,Regulatory Sequences, Nucleic Acid ,Biology ,Mice ,Transcription (biology) ,Insulin-Secreting Cells ,Basic Helix-Loop-Helix Transcription Factors ,Animals ,Humans ,Paired Box Transcription Factors ,Eye Proteins ,Molecular Biology ,Cells, Cultured ,Homeodomain Proteins ,Base Sequence ,Articles ,Cell Biology ,Transfection ,Molecular biology ,Repressor Proteins ,Glucose ,Regulatory sequence ,Mutation ,NEUROD1 ,PAX6 ,Chromatin immunoprecipitation ,Protein Binding - Abstract
MafA is a key transcriptional activator of islet beta cells, and its exclusive expression within beta cells of the developing and adult pancreas is distinct among pancreatic regulators. Region 3 (base pairs -8118 to -7750 relative to the transcription start site), one of six conserved 5' cis domains of the MafA promoter, is capable of directing beta-cell-line-selective expression. Transgenic reporters of region 3 alone (R3), sequences spanning regions 1 to 6 (R1-6; base pairs -10428 to +230), and R1-6 lacking R3 (R1-6(DeltaR3)) were generated. Only the R1-6 transgene was active in MafA(+) insulin(+) cells during development and in adult cells. R1-6 also mediated glucose-induced MafA expression. Conversely, pancreatic expression was not observed with the R3 or R1-6(DeltaR3) line, although much of the nonpancreatic expression pattern was shared between the R1-6 and R1-6(DeltaR3) lines. Further support for the importance of R3 was also shown, as the islet regulators Nkx6.1 and Pax6, but not NeuroD1, activated MafA in gel shift, chromatin immunoprecipitation (ChIP), and transfection assays and in vivo mouse knockout models. Lastly, ChIP demonstrated that Pax6 and Pdx-1 also bound to R1 and R6, potentially functioning in pancreatic and nonpancreatic expression. These data highlight the nature of the cis- and trans-acting factors controlling the beta-cell-specific expression of MafA.
- Published
- 2010
31. Ets Transcription Factors Control Epithelial Maturation and Transit and Crypt-Villus Morphogenesis in the Mammalian Intestine
- Author
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Paul Jedlicka, Lori Sussel, Xiaomei Sui, and Arthur Gutierrez-Hartmann
- Subjects
Cellular differentiation ,Crypt ,Repressor ,Mice, Transgenic ,Biology ,Transfection ,digestive system ,Pathology and Forensic Medicine ,Mice ,Intestinal mucosa ,Animals ,Humans ,Intestinal Mucosa ,CDX2 ,Transcription factor ,Genetics ,Proto-Oncogene Proteins c-ets ,ETS transcription factor family ,digestive, oral, and skin physiology ,Cell Differentiation ,Immunohistochemistry ,engrailed ,Cell biology ,Enterocytes ,HeLa Cells ,Regular Articles - Abstract
Members of the Ets transcription factor family are widely expressed in both the developing and mature mammalian intestine, but their biological functions remain primarily uncharacterized. We used a dominant repressor transgene approach to probe the function of epithelial Ets factors in the homeostasis of the crypt-villus unit, the functional unit of the small intestine. We show that targeted expression in small intestinal epithelium of a fusion protein composed of the Engrailed repressor domain and the Erm DNA-binding domain (En/Erm) results in marked disruption of normal crypt-villus homeostasis, including a cell-autonomous disturbance of epithelial maturation, increased epithelial transit, severe villus dysmorphogenesis, and crypt dysmorphogenesis. The epithelial maturation disturbance is independent of the regulation of TGFbetaRII levels, in contrast to Ets-mediated epithelial differentiation during development; rather, regulation of Cdx2 expression may play a role. The villus dysmorphogenesis is independent of alterations in the crypt-villus boundary and inappropriate beta-catenin activation, and thus appears to represent a new mechanism controlling villus architectural organization. An Analysis of animals mosaic for En/Erm expression suggests that crypt nonautonomous mechanisms underlie the crypt dysmorphogenesis phenotype. Our studies thus uncover novel Ets-regulated pathways of intestinal homeostasis in vivo. Interestingly, the overall En/Erm phenotype of disturbed crypt-villus homeostasis is consistent with recently identified Ets function(s) in the restriction of intestinal epithelial tumorigenesis.
- Published
- 2009
32. Genetic identification of a novel NeuroD1 function in the early differentiation of islet α, PP and ε cells
- Author
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Zoe Loomis, Christina S. Chao, Jacqueline E. Lee, and Lori Sussel
- Subjects
Cellular differentiation ,Mice ,0302 clinical medicine ,Basic Helix-Loop-Helix Transcription Factors ,Cell differentiation ,Mice, Knockout ,0303 health sciences ,geography.geographical_feature_category ,Genetic analysis ,Islet development ,Gene Expression Regulation, Developmental ,Nkx2.2 ,Pancreatic Polypeptide-Secreting Cells ,Islet ,Phenotype ,Ghrelin ,Cell biology ,Homeobox Protein Nkx-2.2 ,α Cells ,medicine.medical_specialty ,NeuroD1 ,Mice, Transgenic ,Nerve Tissue Proteins ,Biology ,Glucagon ,Article ,Islets of Langerhans ,03 medical and health sciences ,Internal medicine ,medicine ,Animals ,RNA, Messenger ,Molecular Biology ,Transcription factor ,030304 developmental biology ,Homeodomain Proteins ,geography ,Cell Biology ,Zebrafish Proteins ,Embryo, Mammalian ,Embryonic stem cell ,Endocrinology ,Glucagon-Secreting Cells ,POU Domain Factors ,NEUROD1 ,030217 neurology & neurosurgery ,Transcription Factors ,Developmental Biology - Abstract
Nkx2.2 and NeuroD1 are vital for proper differentiation of pancreatic islet cell types. Nkx2.2-null mice fail to form beta cells, have reduced numbers of alpha and PP cells and display an increase in ghrelin-producing epsilon cells. NeuroD1-null mice display a reduction of alpha and beta cells after embryonic day (e) 17.5. To begin to determine the relative contributions of Nkx2.2 and NeuroD1 in islet development, we generated Nkx2.2-/-;NeuroD1-/- double knockout (DKO) mice. As expected, the DKO mice fail to form beta cells, similar to the Nkx2.2-null mice, suggesting that the Nkx2.2 phenotype may be dominant over the NeuroD1 phenotype in the beta cells. Surprisingly, however, the alpha, PP and epsilon phenotypes of the Nkx2.2-null mice are partially rescued by the simultaneous elimination of NeuroD1, even at early developmental time points when NeuroD1 null mice alone do not display a phenotype. Our results indicate that Nkx2.2 and NeuroD1 interact to regulate pancreatic islet cell fates, and this epistatic relationship is cell-type dependent. Furthermore, this study reveals a previously unappreciated early function of NeuroD1 in regulating the specification of alpha, PP and epsilon cells.
- Published
- 2007
33. Epigenetic modifications and long noncoding RNAs influence pancreas development and function
- Author
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Luis Arnes and Lori Sussel
- Subjects
Cellular differentiation ,Biology ,Article ,Epigenesis, Genetic ,Islets of Langerhans ,Insulin-Secreting Cells ,Genetics ,medicine ,Glucose homeostasis ,Animals ,Humans ,Epigenetics ,Regulation of gene expression ,geography ,geography.geographical_feature_category ,Models, Genetic ,Endoderm ,Gene Expression Regulation, Developmental ,Cell Differentiation ,Islet ,Cell biology ,medicine.anatomical_structure ,RNA, Long Noncoding ,Pancreas ,Function (biology) - Abstract
Insulin-producing β cells within the pancreatic islet of Langerhans are responsible for maintaining glucose homeostasis; the loss or malfunction of β cells results in diabetes mellitus. Recent advances in cell purification strategies and sequencing technologies as well as novel molecular tools have revealed that epigenetic modifications and long noncoding RNAs (lncRNAs) represent an integral part of the transcriptional mechanisms regulating pancreas development and β cell function. Importantly, these findings have uncovered a new layer of gene regulation in the pancreas that can be exploited to enhance the restoration and/or repair of β cells to treat diabetes.
- Published
- 2015
34. Nkx2.2 is expressed in a subset of enteroendocrine cells with expanded lineage potential
- Author
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Stefanie Gross, Samuel Asfaha, Jing Liu, Dina A. Balderes, Lori Sussel, Guoqiang Gu, and Timothy C. Wang
- Subjects
Pluripotent Stem Cells ,Genotype ,Physiology ,Enteroendocrine Cells ,Cellular differentiation ,Crypt ,Mice, Transgenic ,Nerve Tissue Proteins ,Enteroendocrine cell ,Stem cells ,Biology ,Stem cell marker ,Receptors, G-Protein-Coupled ,Lgr5 ,Intestinal mucosa ,Proto-Oncogene Proteins ,Physiology (medical) ,Basic Helix-Loop-Helix Transcription Factors ,Animals ,Cell Lineage ,Intestinal Mucosa ,Induced pluripotent stem cell ,Cells, Cultured ,Cell Proliferation ,Homeodomain Proteins ,Polycomb Repressive Complex 1 ,Enteroendocrine cells ,Hepatology ,Gastroenterology ,Nkx2.2 ,Cell Differentiation ,Zebrafish Proteins ,Intestinal epithelium ,Bmi1 ,Cell biology ,Mice, Inbred C57BL ,Luminescent Proteins ,Homeobox Protein Nkx-2.2 ,Phenotype ,Immunology ,embryonic structures ,Call for Papers ,Stem cell ,Biomarkers ,Whole-Body Irradiation ,Transcription Factors - Abstract
There are two major stem cell populations in the intestinal crypt region that express either Bmi1 or Lgr5; however, it has been shown that other populations in the crypt can regain stemness. In this study, we demonstrate that the transcription factor NK2 homeobox 2 (Nkx2.2) is expressed in enteroendocrine cells located in the villus and crypt of the intestinal epithelium and is coexpressed with the stem cell markers Bmi1 and Lgr5 in a subset of crypt cells. To determine whether Nkx2.2-expressing enteroendocrine cells display cellular plasticity and stem cell potential, we performed genetic lineage tracing of the Nkx2.2-expressing population using Nkx2.2Cre/+; R26RTomato mice. These studies demonstrated that Nkx2.2+ cells are able to give rise to all intestinal epithelial cell types in basal conditions. The proliferative capacity of Nkx2.2-expressing cells was also demonstrated in vitro using crypt organoid cultures. Injuring the intestine with irradiation, systemic inflammation, and colitis did not enhance the lineage potential of Nkx2.2-expressing cells. These findings demonstrate that a rare mature enteroendocrine cell subpopulation that is demarcated by Nkx2.2 expression display stem cell properties during normal intestinal epithelial homeostasis, but is not easily activated upon injury.
- Published
- 2015
35. Gata6 is an important regulator of mouse pancreas development
- Author
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Kimberly J. Decker, Devorah C Goldman, Catherine L Grasch, and Lori Sussel
- Subjects
Gata4 ,Gata6 ,Organogenesis ,Cellular differentiation ,Regulator ,Enteroendocrine cell ,Mice ,0302 clinical medicine ,GATA6 Transcription Factor ,Pancreas development ,Drosophila Proteins ,Promoter Regions, Genetic ,reproductive and urinary physiology ,0303 health sciences ,geography.geographical_feature_category ,GATA6 ,Cell Differentiation ,respiratory system ,Islet ,Homeobox Protein Nkx-2.2 ,medicine.anatomical_structure ,embryonic structures ,cardiovascular system ,Pancreas ,endocrine system ,Cell type ,Mice, Transgenic ,Biology ,Article ,Cell Line ,03 medical and health sciences ,medicine ,Animals ,Cell Lineage ,Molecular Biology ,030304 developmental biology ,Homeodomain Proteins ,geography ,Cell Biology ,Zebrafish Proteins ,Epithelium ,GATA4 Transcription Factor ,Rats ,Trans-Activators ,Cancer research ,030217 neurology & neurosurgery ,Transcription Factors ,Developmental Biology - Abstract
Gata4, Gata5, and Gata6 represent a subfamily of zinc-finger transcriptional regulators that are important in the development and differentiation of numerous tissues, including many endodermally-derived organs. We demonstrate that Gata4 and Gata6 have overlapping expression patterns in the early pancreatic epithelium. Later, Gata4 becomes restricted to exocrine tissue and Gata6 becomes restricted to a subset of endocrine cells. In addition, we show Gata6, but not Gata4, physically interacts with Nkx2.2, an essential islet transcription factor. To begin determining the roles that Gata4 and Gata6 play during pancreatic development, we expressed Gata4-Engrailed and Gata6-Engrailed dominant repressor fusion proteins in the pancreatic epithelium and in the islet. At e17.5, transgenic Gata6-Engrailed embryos exhibit two distinct phenotypes: a complete absence of pancreas or a reduction in pancreatic tissue. In the embryos that do form pancreas, there is a significant reduction of all pancreatic cell types, with the few differentiated endocrine cells clustered within, or in close proximity to, enlarged ductal structures. Conversely, the majority of transgenic Gata4-Engrailed embryos do not have a pancreatic phenotype. This study suggests that Gata6 is an important regulator of pancreas specification.
- Published
- 2006
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36. The concerted activities of Pax4 and Nkx2.2 are essential to initiate pancreatic β-cell differentiation
- Author
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Hasan Kizilocak, Lynda Elghazi, Lori Sussel, Susan E Parker, Masahide Asano, Junfeng Wang, and Beatriz Sosa-Pineda
- Subjects
Mouse ,Cellular differentiation ,Homeobox A1 ,030209 endocrinology & metabolism ,Enteroendocrine cell ,Biology ,Islets of Langerhans ,Mice ,03 medical and health sciences ,0302 clinical medicine ,medicine ,Animals ,Paired Box Transcription Factors ,Glucose homeostasis ,CDX2 ,Pancreas ,Molecular Biology ,In Situ Hybridization ,030304 developmental biology ,Homeodomain Proteins ,Mice, Knockout ,Pax4 ,β-cell differentiation ,0303 health sciences ,Nkx2.2 ,Cell Differentiation ,Cell Biology ,Zebrafish Proteins ,Immunohistochemistry ,Molecular biology ,Pax6 ,Cell biology ,Homeobox Protein Nkx-2.2 ,medicine.anatomical_structure ,PDX1 ,PAX4 ,Transcription Factors ,Developmental Biology - Abstract
Pancreatic beta cells play a central role in maintaining glucose homeostasis because they secrete insulin in response to increased level of blood glucose; failure of this capacity constitutes a major component of the pathogenesis of diabetes. The identification of key regulators of pancreatic beta-cell differentiation is relevant for the overall understanding of this process and for future experiments aimed at regenerating insulin-producing beta cells from pancreatic or embryonic stem cells. Several studies using transgenic or knockout mice have established that the development and function of pancreatic beta cells are controlled by several genes encoding specific transcription factors. By inactivating the homeobox gene Pax4, we previously demonstrated that its function is required for the formation of mature insulin-producing cells. Here, we show that during pancreas ontogeny, Pax4 is expressed in differentiating endocrine cells, including beta cells. Pax4 activity appears essential for appropriate initiation of beta-cell differentiation because loss of Pax4 prevents the expression of Pdx1, HB9 and insulin in beta-cell precursors. This role of Pax4 appears to be accomplished via its genetic interaction with another homeobox gene, Nkx2.2.
- Published
- 2004
37. Transcription Factor Occupancy of the Insulin Gene in Vivo
- Author
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Eva Henderson, Roland Stein, Michelle A. Cissell, Li Zhao, and Lori Sussel
- Subjects
Sp1 transcription factor ,General transcription factor ,Sp3 transcription factor ,Pioneer factor ,Response element ,Promoter ,E-box ,Cell Biology ,Biology ,Enhancer ,Molecular Biology ,Biochemistry ,Molecular biology - Abstract
Consensus-binding sites for many transcription factors are relatively non-selective and found at high frequency within the genome. This raises the possibility that factors that are capable of binding to a cis-acting element in vitro and regulating transcription from a transiently transfected plasmid, which would not have higher order chromatin structure, may not occupy this site within the endogenous gene. Closed chromatin structure and competition from another DNA-binding protein with similar nucleotide specificity are two possible mechanisms by which a transcription factor may be excluded from a potential binding sitein vivo. Multiple transcription factors, including Pdx-1, BETA-2, and Pax6, have been implicated in expression of the insulin gene in pancreatic β cells. In this study, the chromatin immunoprecipitation assay has been used to show that these factors do, in fact, bind to insulin control region sequences in intact β cells. In addition, another key islet-enriched transcription factor, Nkx2.2, was found to occupy this region using the chromatin immunoprecipitation assay. In vitro DNA-binding and transient transfection assays defined how Nkx2.2 affected insulin gene expression. Pdx-1 was also shown to bind within a region of the endogenous islet amyloid polypeptide, pax-4, and glucokinase genes that were associated with control in vitro. Because Pdx-1 does not regulate gene transcription in isolation, these sequences were examined for occupancy by the other insulin transcriptional regulators. BETA-2, Pax6, and Nkx2.2 were also found to bind to amyloid polypeptide, glucokinase, andpax-4 control sequences in vivo. These studies reveal the broad application of the Pdx-1, BETA-2, Pax6, and Nkx2.2 transcription factors in regulating expression of genes selectively expressed in islet β cells.
- Published
- 2003
38. Homeobox gene Nkx6.1 lies downstream of Nkx2.2 in the major pathway of β-cell formation in the pancreas
- Author
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Lori Sussel, A. Hayes-Jordan, David W. Scheel, Maike Sander, F. Dela Cruz, Valerie M. Schwitzgebel, Jennifer R. Conners, Michael S. German, and J. Kalamaras
- Subjects
Cellular differentiation ,Mutant ,Pancreas/cytology/embryology/metabolism ,Pancreas morphogenesis ,Transcription Factors/genetics/metabolism ,Neogenesis ,Islets of Langerhans ,Mice ,03 medical and health sciences ,0302 clinical medicine ,Animals ,Pancreas ,Molecular Biology ,030304 developmental biology ,Homeodomain Proteins ,Mice, Knockout ,Regulation of gene expression ,Genetics ,0303 health sciences ,geography ,ddc:618 ,geography.geographical_feature_category ,biology ,Genes, Homeobox ,Gene Expression Regulation, Developmental ,Cell Differentiation ,Islets of Langerhans/cytology/embryology/metabolism ,Zebrafish Proteins ,biology.organism_classification ,Islet ,Cell biology ,Homeobox Protein Nkx-2.2 ,Mutation ,embryonic structures ,Homeobox ,Homeodomain Proteins/genetics ,Beta cell ,030217 neurology & neurosurgery ,Transcription Factors ,Developmental Biology - Abstract
Most insulin-producing β-cells in the fetal mouse pancreas arise during the secondary transition, a wave of differentiation starting at embryonic day 13. Here, we show that disruption of homeobox gene Nkx6.1 in mice leads to loss of β-cell precursors and blocks β-cell neogenesis specifically during the secondary transition. In contrast, islet development in Nkx6.1/Nkx2.2 double mutant embryos is identical to Nkx2.2 single mutant islet development: β-cell precursors survive but fail to differentiate into β-cells throughout development. Together, these experiments reveal two independently controlled pathways for β-cell differentiation, and place Nkx6.1 downstream of Nkx2.2 in the major pathway of β-cell differentiation.
- Published
- 2000
39. Loss of Nkx2.1 homeobox gene function results in a ventral to dorsal molecular respecification within the basal telencephalon: evidence for a transformation of the pallidum into the striatum
- Author
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S. Kimura, Oscar Marín, Lori Sussel, and John L.R. Rubenstein
- Subjects
Male ,Telencephalon ,Ganglionic eminence ,Thyroid Nuclear Factor 1 ,Striatum ,Biology ,Globus Pallidus ,Mice ,Cell Movement ,Interneurons ,Pregnancy ,Basal ganglia ,medicine ,Animals ,Hedgehog Proteins ,Cholinergic neuron ,Molecular Biology ,Body Patterning ,DNA Primers ,Homeodomain Proteins ,Mice, Knockout ,Basal forebrain ,Base Sequence ,Cerebrum ,DLX2 ,Genes, Homeobox ,Gene Expression Regulation, Developmental ,Nuclear Proteins ,Proteins ,Anatomy ,Corpus Striatum ,Mice, Mutant Strains ,Cell biology ,Phenotype ,medicine.anatomical_structure ,nervous system ,Cerebral cortex ,embryonic structures ,Trans-Activators ,Female ,Transcription Factors ,Developmental Biology - Abstract
The telencephalon is organized into distinct longitudinal domains: the cerebral cortex and the basal ganglia. The basal ganglia primarily consists of a dorsal region (striatum) and a ventral region (pallidum). Within the telencephalon, the anlage of the pallidum expresses the Nkx2.1 homeobox gene. A mouse deficient in Nkx2.1 function does not form pallidal structures, lacks basal forebrain TrkA-positive neurons (probable cholinergic neurons) and has reduced numbers of cortical cells expressing GABA, DLX2 and calbindin that migrate from the pallidum through the striatum and into the cortex. We present evidence that these phenotypes result from a ventral-to-dorsal transformation of the pallidal primordium into a striatal-like anlage.
- Published
- 1999
40. Homeobox gene Nkx2.2 and specification of neuronal identity by graded Sonic hedgehog signalling
- Author
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Palle Serup, Thomas M. Jessell, James Briscoe, Dennis J. Hartigan-O'Connor, Johan Ericson, Lori Sussel, and John L.R. Rubenstein
- Subjects
Cell type ,PAX6 Transcription Factor ,Interneuron ,Neural tube patterning ,Mice ,Interneurons ,Culture Techniques ,medicine ,Animals ,Paired Box Transcription Factors ,Cell Lineage ,Hedgehog Proteins ,Sonic hedgehog ,Progenitor cell ,Eye Proteins ,Body Patterning ,Embryonic Induction ,Homeodomain Proteins ,Motor Neurons ,Neurons ,Genetics ,Multidisciplinary ,biology ,Genes, Homeobox ,Proteins ,Zebrafish Proteins ,Motor neuron ,Cell biology ,DNA-Binding Proteins ,Repressor Proteins ,Rhombencephalon ,Homeobox Protein Nkx-2.2 ,medicine.anatomical_structure ,Spinal Cord ,Mutation ,embryonic structures ,Trans-Activators ,biology.protein ,Neuron ,PAX6 ,Signal Transduction ,Transcription Factors - Abstract
During vertebrate development, the specification of distinct cell types is thought to be controlled by inductive signals acting at different concentration thresholds1. The degree of receptor activation in response to these signals is a known determinant of cell fate2, but the later steps at which graded signals are converted into all-or-none distinctions in cell identity remain poorly resolved. In the ventral neural tube, motor neuron and interneuron generation depends on the graded activity of the signalling protein Sonic hedgehog (Shh)3,4,5. These neuronal subtypes derive from distinct progenitor cell populations that express the homeodomain proteins Nkx2.2 or Pax6 in response to graded Shh signalling6,7. In mice lacking Pax6, progenitor cells generate neurons characteristic of exposure to greater Shh activity6,7. However, Nkx2.2 expression expands dosally in Pax6 mutants6, raising the possibility that Pax6 controls neuronal pattern indirectly. Here we provide evidence that Nkx2.2 has a primary role in ventral neuronal patterning. In Nkx2.2 mutants, Pax6 expression is unchanged but cells undergo a ventral-to-dorsal transformation in fate and generate motor neurons rather than interneurons. Thus, Nkx2.2 has an essential role in interpreting graded Shh signals and selecting neuronal identity.
- Published
- 1999
41. Regulation of Neurod1 contributes to the lineage potential of Neurogenin3+ endocrine precursor cells in the pancreas
- Author
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Keith R. Anderson, Lori Sussel, James B. Papizan, and Teresa L. Mastracci
- Subjects
Cancer Research ,endocrine system ,Histology ,lcsh:QH426-470 ,Cellular differentiation ,Enteroendocrine cell ,Biology ,Alpha cell ,Islets of Langerhans ,Mice ,03 medical and health sciences ,0302 clinical medicine ,Insulin-Secreting Cells ,Basic Helix-Loop-Helix Transcription Factors ,Genetics ,medicine ,Animals ,Humans ,Cell Lineage ,Progenitor cell ,Pancreas ,Molecular Biology ,Genetics (clinical) ,Ecology, Evolution, Behavior and Systematics ,030304 developmental biology ,Homeodomain Proteins ,0303 health sciences ,Stem Cells ,Gene Expression Regulation, Developmental ,Nuclear Proteins ,Cell Differentiation ,Zebrafish Proteins ,Cell biology ,lcsh:Genetics ,Homeobox Protein Nkx-2.2 ,medicine.anatomical_structure ,Glucagon-Secreting Cells ,Epsilon cell ,PDX1 ,Stem cell ,030217 neurology & neurosurgery ,Signal Transduction ,Transcription Factors ,Research Article ,Developmental Biology - Abstract
During pancreatic development, transcription factor cascades gradually commit precursor populations to the different endocrine cell fate pathways. Although mutational analyses have defined the functions of many individual pancreatic transcription factors, the integrative transcription factor networks required to regulate lineage specification, as well as their sites of action, are poorly understood. In this study, we investigated where and how the transcription factors Nkx2.2 and Neurod1 genetically interact to differentially regulate endocrine cell specification. In an Nkx2.2 null background, we conditionally deleted Neurod1 in the Pdx1+ pancreatic progenitor cells, the Neurog3+ endocrine progenitor cells, or the glucagon+ alpha cells. These studies determined that, in the absence of Nkx2.2 activity, removal of Neurod1 from the Pdx1+ or Neurog3+ progenitor populations is sufficient to reestablish the specification of the PP and epsilon cell lineages. Alternatively, in the absence of Nkx2.2, removal of Neurod1 from the Pdx1+ pancreatic progenitor population, but not the Neurog3+ endocrine progenitor cells, restores alpha cell specification. Subsequent in vitro reporter assays demonstrated that Nkx2.2 represses Neurod1 in alpha cells. Based on these findings, we conclude that, although Nkx2.2 and Neurod1 are both necessary to promote beta cell differentiation, Nkx2.2 must repress Neurod1 in a Pdx1+ pancreatic progenitor population to appropriately commit a subset of Neurog3+ endocrine progenitor cells to the alpha cell lineage. These results are consistent with the proposed idea that Neurog3+ endocrine progenitor cells represent a heterogeneous population of unipotent cells, each restricted to a particular endocrine lineage., Author Summary Diabetes mellitus is a family of metabolic diseases that can result from either destruction or dysfunction of the insulin-producing beta cells of the pancreas. Recent studies have provided hope that generating insulin-producing cells from alternative cell sources may be a possible treatment for diabetes; this includes the observation that pancreatic glucagon-expressing alpha cells can be converted into beta cells under certain physiological or genetic conditions. Our study focuses on two essential beta cell regulatory factors, Nkx2.2 and Neurod1, and demonstrates how their genetic interactions can promote the development of other hormone-expressing cell types, including alpha cells. We determined that, while Nkx2.2 is required to activate Neurod1 to promote beta cell formation, Nkx2.2 must prevent expression of Neurod1 to allow alpha cell formation. Furthermore, the inactivation of Neurod1 must occur in the earliest pancreatic progenitors, at a stage in the differentiation process earlier than previously believed. These studies contribute to our understanding of the overlapping gene regulatory networks that specify islet cell types and identify the importance of timing and cellular context for these regulatory interactions. Furthermore, our data have broad implications regarding the manipulation of alpha cells or human pluripotent stem cells to generate insulin-producing beta cells for therapeutic purposes.
- Published
- 2013
42. ALDH1B1 is a potential stem / progenitor marker for multiple pancreas progenitor pools
- Author
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Surendra Singh, Luis Arnes, Ioannis Serafimidis, Marilia Ioannou, Anthony Gavalas, Vasilis Vasiliou, and Lori Sussel
- Subjects
Time Factors ,Genotype ,Pancreas injury ,Aldehyde dehydrogenase ,Article ,Aldehyde Dehydrogenase 1 Family ,Catalysis ,Centroacinar cells ,03 medical and health sciences ,NGN3 ,Mice ,0302 clinical medicine ,medicine ,Animals ,Progenitor cell ,Molecular Biology ,Pancreas ,In Situ Hybridization ,030304 developmental biology ,Progenitor ,0303 health sciences ,PDX1 ,biology ,Aldehyde Dehydrogenase, Mitochondrial ,Stem Cells ,Gene Expression Regulation, Developmental ,Cell Biology ,Aldehyde Dehydrogenase ,Oligonucleotides, Antisense ,Streptozotocin ,Embryonic stem cell ,Molecular biology ,Up-Regulation ,Pancreas stem and progenitor cells ,medicine.anatomical_structure ,Microscopy, Fluorescence ,030220 oncology & carcinogenesis ,Mutation ,biology.protein ,Stem cell ,Developmental Biology ,medicine.drug - Abstract
Aldehyde dehydrogenase (ALDH) genes are increasingly associated with stem/progenitor cell status but their role in the maintenance of pluripotency remains uncertain. In a screen conducted for downstream Ngn3 target genes using ES derived pancreas progenitors we identified Aldh1b1, encoding a mitochondrial enzyme, as one of the genes strongly up regulated in response to Ngn3 expression. We found both by in situ hybridization and immunofluorescence using a specific antibody that ALDH1B1 is exclusively expressed in the emerging pancreatic buds of the early embryo (9.5 dpc) in a Pdx1 dependent manner. Around the time of secondary transition, ALDH1B1 expression was restricted in the tip tripotent progenitors of the branching epithelium and in a subset of the trunk epithelium. Expression in the latter was Ngn3 dependent. Subsequently, ALDH1B1 expression persisted only in the tip cells that become restricted to the exocrine lineage and declined rapidly as these cells mature. In the adult pancreas we identified rare ALDH1B1+ cells that become abundant following pancreas injury in either the caerulein or streptozotocin paradigms. Blocking ALDH catalytic activity in pancreas embryonic explants resulted in reduced size of the explants and accelerated differentiation suggesting for the first time that ALDH activity may be necessary in the developing pancreas for the maintenance and expansion of progenitor pools.
- Published
- 2012
43. Pancreatic β-Cell Dedifferentiation As Mechanism Of Diabetic β-Cell Failure
- Author
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Shouhong Xuan, Domenico Accili, Lori Sussel, Chutima Talchai, and Hua V. Lin
- Subjects
Male ,Homeobox protein NANOG ,medicine.medical_specialty ,Cell ,FOXO1 ,Enteroendocrine cell ,Cell fate determination ,Biology ,General Biochemistry, Genetics and Molecular Biology ,Article ,Mice ,Insulin-Secreting Cells ,Internal medicine ,medicine ,Animals ,Insulin ,Pancreas ,Forkhead Box Protein O1 ,Cell growth ,Biochemistry, Genetics and Molecular Biology(all) ,Forkhead Transcription Factors ,Cell Dedifferentiation ,Cell cycle ,Cell biology ,medicine.anatomical_structure ,Endocrinology ,Diabetes Mellitus, Type 2 - Abstract
SummaryDiabetes is associated with β cell failure. But it remains unclear whether the latter results from reduced β cell number or function. FoxO1 integrates β cell proliferation with adaptive β cell function. We interrogated the contribution of these two processes to β cell dysfunction, using mice lacking FoxO1 in β cells. FoxO1 ablation caused hyperglycemia with reduced β cell mass following physiologic stress, such as multiparity and aging. Surprisingly, lineage-tracing experiments demonstrated that loss of β cell mass was due to β cell dedifferentiation, not death. Dedifferentiated β cells reverted to progenitor-like cells expressing Neurogenin3, Oct4, Nanog, and L-Myc. A subset of FoxO1-deficient β cells adopted the α cell fate, resulting in hyperglucagonemia. Strikingly, we identify the same sequence of events as a feature of different models of murine diabetes. We propose that dedifferentiation trumps endocrine cell death in the natural history of β cell failure and suggest that treatment of β cell dysfunction should restore differentiation, rather than promoting β cell replication.PaperFlick
- Published
- 2012
44. Ghrelin expression in the mouse pancreas defines a unique multipotent progenitor population
- Author
-
Luis Arnes, Jonathon T. Hill, Stefanie Gross, Mark A. Magnuson, and Lori Sussel
- Subjects
Cellular differentiation ,lcsh:Medicine ,Enteroendocrine cell ,Mice ,0302 clinical medicine ,Endocrinology ,Molecular Cell Biology ,Basic Helix-Loop-Helix Transcription Factors ,Signaling in Cellular Processes ,lcsh:Science ,0303 health sciences ,Multidisciplinary ,geography.geographical_feature_category ,Stem Cells ,Cell Differentiation ,Islet ,Ghrelin ,Cell biology ,medicine.anatomical_structure ,Homeobox Protein Nkx-2.2 ,Medicine ,Cellular Types ,Pancreas ,Research Article ,Signal Transduction ,medicine.medical_specialty ,Cell type ,endocrine system ,Nerve Tissue Proteins ,Gastroenterology and Hepatology ,Biology ,Glucose Signaling ,03 medical and health sciences ,Islets of Langerhans ,Internal medicine ,medicine ,Animals ,Progenitor cell ,030304 developmental biology ,Homeodomain Proteins ,Diabetic Endocrinology ,Delta cell ,geography ,Multipotent Stem Cells ,lcsh:R ,Zebrafish Proteins ,Mice, Inbred C57BL ,Multipotent Stem Cell ,lcsh:Q ,030217 neurology & neurosurgery ,Transcription Factors ,Developmental Biology - Abstract
Pancreatic islet cells provide the major source of counteractive endocrine hormones required for maintaining glucose homeostasis; severe health problems result when these cell types are insufficiently active or reduced in number. Therefore, the process of islet endocrine cell lineage allocation is critical to ensure there is a correct balance of islet cell types. There are four endocrine cell types within the adult islet, including the glucagon-producing alpha cells, insulin-producing beta cells, somatostatin-producing delta cells and pancreatic polypeptide-producing PP cells. A fifth islet cell type, the ghrelin-producing epsilon cells, is primarily found during gestational development. Although hormone expression is generally assumed to mark the final entry to a determined cell state, we demonstrate in this study that ghrelin-expressing epsilon cells within the mouse pancreas do not represent a terminally differentiated endocrine population. Ghrelin cells give rise to significant numbers of alpha and PP cells and rare beta cells in the adult islet. Furthermore, pancreatic ghrelin-producing cells are maintained in pancreata lacking the essential endocrine lineage regulator Neurogenin3, and retain the ability to contribute to cells within the pancreatic ductal and exocrine lineages. These results demonstrate that the islet ghrelin-expressing epsilon cells represent a multi-potent progenitor cell population that delineates a major subgrouping of the islet endocrine cell populations. These studies also provide evidence that many of hormone-producing cells within the adult islet represent heterogeneous populations based on their ontogeny, which could have broader implications on the regulation of islet cell ratios and their ability to effectively respond to fluctuations in the metabolic environment during development.
- Published
- 2012
45. Generation of Nkx2.2:lacZ mice using recombination-mediated cassette exchange technology
- Author
-
Kevin Leclerc, Mark A. Magnuson, Lori Sussel, Jessica M. Friel, Luis Arnes, and Susan B. Hipkens
- Subjects
Central Nervous System ,Genotype ,Mice, Transgenic ,Biology ,Article ,Mice ,Endocrinology ,stomatognathic system ,Neural Stem Cells ,Gene Order ,Genetics ,Animals ,Progenitor cell ,Homologous Recombination ,Transcription factor ,Pancreas ,Progenitor ,Homeodomain Proteins ,Recombinase-mediated cassette exchange ,Gene Expression Regulation, Developmental ,Embryo ,Heterozygote advantage ,Cell Biology ,respiratory system ,Zebrafish Proteins ,beta-Galactosidase ,Phenotype ,Molecular biology ,Homeobox Protein Nkx-2.2 ,embryonic structures ,Gene Targeting ,cardiovascular system ,Homeobox ,Transcription Factors - Abstract
Nkx2.2 encodes a homeodomain transcription factor required for the correct specification and/or differentiation of cells in the pancreas, intestine, and central nervous system (CNS). To follow the fate of cells deleted for Nkx2.2 within these tissues, we generated Nkx2.2:lacZ knockin mice using a recombination-mediated cassette exchange (RMCE) approach. Expression analysis of lacZ and/or β-galactosidase in Nkx2.2lacZ/+ heterozygote embryos and adults demonstrates that lacZ faithfully recapitulates endogenous Nkx2.2 expression. Furthermore, the Nkx2.2lacZ/lacZ homozygous embryos display phenotypes indistinguishable from the previously characterized Nkx2.2−/− strain. LacZ expression analyses in the Nkx2.2lacZ/lacZ homozygous embryos indicate that Nkx2.2-expressing progenitor cells within the pancreas are generated in their normal numbers and are not mislocalized within the pancreatic ductal epithelium or developing islets. In the CNS of Nkx2.2lacZ/lacZ embryos, LacZ-expressing cells within the ventral P3 progenitor domain display different migration properties depending on the developmental stage and their respective differentiation potential. genesis 50:612–624, 2012. © 2012 Wiley Periodicals, Inc.
- Published
- 2011
46. Nkx2.2 and Arx genetically interact to regulate pancreatic endocrine cell development and endocrine hormone expression
- Author
-
Lori Sussel, Crystal L. Wilcox, Teresa L. Mastracci, Catherine Lee May, Jeffrey A. Golden, Casandra Panea, and Luis Arnes
- Subjects
medicine.medical_specialty ,Endocrine cell fate ,Population ,Enteroendocrine cell ,Biology ,PP ,Article ,Mice ,Transcriptional regulation ,stomatognathic system ,Internal medicine ,medicine ,Animals ,Cell Lineage ,Progenitor cell ,education ,Molecular Biology ,Pancreas ,Homeodomain Proteins ,education.field_of_study ,Nkx2.2 ,Cell Biology ,respiratory system ,Zebrafish Proteins ,Pancreatic Hormones ,Ghrelin ,Cell biology ,medicine.anatomical_structure ,Endocrinology ,Somatostatin ,Homeobox Protein Nkx-2.2 ,Epsilon cell ,embryonic structures ,cardiovascular system ,Arx ,PP cell ,Developmental Biology ,Transcription Factors - Abstract
Nkx2.2 and Arx are essential pancreatic transcription factors. Nkx2.2 is necessary for the appropriate specification of the islet alpha, beta, PP and epsilon cell lineages, whereas Arx is required to form the correct ratio of alpha, beta, delta and PP cells. To begin to understand the cooperative functions of Nkx2.2 and Arx in the development of endocrine cell lineages, we generated progenitor cell-specific deletions of Arx on the Nkx2.2 null background. The analysis of these mutants demonstrates that expansion of the ghrelin cell population in the Nkx2.2 null pancreas is not dependent on Arx; however, Arx is necessary for the upregulation of ghrelin mRNA levels in Nkx2.2 mutant epsilon cells. Alternatively, in the absence of Arx, delta cell numbers are increased and Nkx2.2 becomes essential for the repression of somatostatin gene expression. Interestingly, the dysregulation of ghrelin and somatostatin expression in the Nkx2.2/Arx compound mutant (Nkx2.2null;ArxΔpanc) results in the appearance of ghrelin+/somatostatin+ co-expressing cells. These compound mutants also revealed a genetic interaction between Nkx2.2 and Arx in the regulation of the PP cell lineage; the PP cell population is reduced when Nkx2.2 is deleted but is restored back to wildtype numbers in the Nkx2.2null;ArxΔpanc mutant. Moreover, conditional deletion of Arx in specific pancreatic cell populations established that the functions of Arx are necessary in the Neurog3+ endocrine progenitors. Together, these experiments identify novel genetic interactions between Nkx2.2 and Arx within the endocrine progenitor cells that ensure the correct specification and regulation of endocrine hormone-producing cells.
- Published
- 2011
47. Cooperative Transcriptional Regulation of the Essential Pancreatic Islet Gene NeuroD1 (Beta2) by Nkx2.2 and Neurogenin 3*
- Author
-
Christopher B. Newgard, James Hagman, Keely Solomon, Keith R. Anderson, Lori Sussel, Thomas C. Becker, Ciara A. Torres, and Christopher V.E. Wright
- Subjects
Cell type ,endocrine system ,Transcription, Genetic ,Mice, Transgenic ,Nerve Tissue Proteins ,Biology ,Cell fate determination ,Biochemistry ,Islets of Langerhans ,Mice ,Molecular Basis of Cell and Developmental Biology ,Insulin-Secreting Cells ,Transcriptional regulation ,Basic Helix-Loop-Helix Transcription Factors ,Animals ,Progenitor cell ,Molecular Biology ,Transcription factor ,Zebrafish ,Regulation of gene expression ,Homeodomain Proteins ,Mice, Knockout ,geography ,geography.geographical_feature_category ,Cell growth ,Gene Expression Regulation, Developmental ,Cell Biology ,Zebrafish Proteins ,Islet ,Molecular biology ,Cell biology ,Homeobox Protein Nkx-2.2 ,embryonic structures ,Transcription Factors - Abstract
Nkx2.2 and NeuroD1 are two critical regulators of pancreatic beta cell development. Nkx2.2 is a homeodomain transcription factor that is essential for islet cell type specification and mature beta cell function. NeuroD1 is a basic helix-loop-helix transcription factor that is critical for islet beta cell maturation and maintenance. Although both proteins influence beta cell development directly downstream of the endocrine progenitor factor, neurogenin3 (Ngn3), a connection between the two proteins in the regulation of beta cell fate and function has yet to be established. In this study, we demonstrate that Nkx2.2 transcriptional activity is required to facilitate the activation of NeuroD1 by Ngn3. Furthermore, Nkx2.2 is necessary to maintain high levels of NeuroD1 expression in developing mouse and zebrafish islets and in mature beta cells. Interestingly, Nkx2.2 regulates NeuroD1 through two independent promoter elements, one that is bound and activated directly by Nkx2.2 and one that appears to be regulated by Nkx2.2 through an indirect mechanism. Together, these findings suggest that Nkx2.2 coordinately activates NeuroD1 with Ngn3 within the endocrine progenitor cell and also plays a role in the maintenance of NeuroD1 expression to regulate beta cell function in the mature islet. Collectively, these findings further define the conserved regulatory networks involved in islet beta cell formation and function.
- Published
- 2009
48. Pancreatic beta cells require NeuroD to achieve and maintain functional maturity
- Author
-
Klaus H. Kaestner, Chunyan Gu, Jacqueline E. Lee, Lori Sussel, Gretchen H. Stein, Klaus-Armin Nave, Sandra Goebbels, Pedro Luis Herrera, Hanna Hörnberg, Ning Pan, and Peter White
- Subjects
medicine.medical_specialty ,animal structures ,Physiology ,medicine.medical_treatment ,HUMDISEASE ,Protein Array Analysis ,Nerve Tissue Proteins ,Biology ,Carbohydrate metabolism ,Article ,Mice ,Oxygen Consumption ,KATP Channels ,Internal medicine ,Insulin-Secreting Cells ,Insulin Secretion ,medicine ,Basic Helix-Loop-Helix Transcription Factors ,Animals ,Insulin ,Glycolysis ,Secretion ,Neuropeptide Y ,Beta (finance) ,Molecular Biology ,ddc:616 ,NeuroD ,Cell Biology ,Endocrinology ,medicine.anatomical_structure ,Glucose ,embryonic structures ,sense organs ,Beta cell ,Pancreas - Abstract
NeuroD, a transactivator of the insulin gene, is critical for development of the endocrine pancreas, and NeuroD mutations cause MODY6 in humans. To investigate the role of NeuroD in differentiated beta cells, we generated mice in which neuroD is deleted in insulin-expressing cells. These mice exhibit severe glucose intolerance. Islets lacking NeuroD respond poorly to glucose and display a glucose metabolic profile similar to immature beta cells, featuring increased expression of glycolytic genes and LDHA, elevated basal insulin secretion and O2 consumption, and overexpression of NPY. Moreover, the mutant islets appear to have defective K(ATP) channel-mediated insulin secretion. Unexpectedly, virtually all insulin in the mutant mice is derived from ins2, whereas ins1 expression is almost extinguished. Overall, these results indicate that NeuroD is required for beta cell maturation and demonstrate the importance of NeuroD in the acquisition and maintenance of fully functional glucose-responsive beta cells.
- Published
- 2009
49. Nkx2.2 regulates cell fate choice in the enteroendocrine cell lineages of the intestine
- Author
-
Zoe Loomis, Michelle J. Doyle, Shailey Desai, Erica L McCoy, Jessica M. Schrunk, Aimee E. Pugh-Bernard, Angela Minic, and Lori Sussel
- Subjects
medicine.medical_specialty ,Cell type ,Cellular differentiation ,Population ,Cell ,Enteroendocrine cell ,Biology ,Cell fate determination ,Article ,03 medical and health sciences ,Mice ,0302 clinical medicine ,stomatognathic system ,Internal medicine ,Endocrine Glands ,Intestine, Small ,medicine ,Intestinal epithelial cell differentiation ,Animals ,Cell Lineage ,education ,Molecular Biology ,030304 developmental biology ,Enteroendocrine cells ,Homeodomain Proteins ,0303 health sciences ,education.field_of_study ,geography ,geography.geographical_feature_category ,Nkx2.2 ,Gene Expression Regulation, Developmental ,Cell Differentiation ,Cell Biology ,respiratory system ,Zebrafish Proteins ,Islet ,Ghrelin ,Hormones ,Cell biology ,Intestine ,Endocrinology ,medicine.anatomical_structure ,Homeobox Protein Nkx-2.2 ,030220 oncology & carcinogenesis ,Cell type specification ,embryonic structures ,cardiovascular system ,Developmental Biology ,Transcription Factors - Abstract
Nkx2.2 is a homeodomain-containing transcription factor essential for pancreatic islet cell specification. In this study we investigate the role of Nkx2.2 within the small intestine. We have determined that Nkx2.2 is expressed at the onset of intestinal epithelial cell differentiation in specific intestinal cell populations, including a subset of enteroendocrine cells. Similar to its role in the pancreatic islet, Nkx2.2 regulates cell fate choices within the intestinal enteroendocrine population; in the Nkx2.2 null mice, several hormone-producing enteroendocrine cell populations are absent or reduced and the ghrelin-producing cell population is upregulated. The remaining intestinal cell populations, including the paneth cells, goblet cells, and enterocytes appear to be unaffected by the loss of Nkx2.2. Furthermore, similar to the pancreatic islet, Nkx2.2 appears to function upstream of Pax6 in regulating intestinal cell fates; Pax6 mRNA and protein expression is decreased in the Nkx2.2 null mice. These studies identify a novel role for Nkx2.2 in intestinal endocrine cell development and reveal the regulatory similarities between cell type specification in the pancreatic islet and in the enteroendocrine population of the intestine.
- Published
- 2006
50. FoxA2, Nkx2.2, and PDX-1 Regulate Islet β-Cell-Specific mafA Expression through Conserved Sequences Located between Base Pairs −8118 and −7750 Upstream from the Transcription Start Site
- Author
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Isabella Artner, Kevin Gerrish, Jonathan C. Schisler, Lori Sussel, Min Guo, Christopher B. Newgard, Eva Henderson, Jeffrey C. Raum, and Roland Stein
- Subjects
Maf Transcription Factors, Large ,Transcription, Genetic ,Molecular Sequence Data ,Biology ,Regulatory Sequences, Nucleic Acid ,Conserved sequence ,Cell Line ,Mice ,Transcription (biology) ,Insulin-Secreting Cells ,Animals ,Humans ,RNA, Small Interfering ,Molecular Biology ,Gene ,Transcription factor ,Regulation of gene expression ,Homeodomain Proteins ,Messenger RNA ,Base Sequence ,Nuclear Proteins ,Cell Biology ,Articles ,Zebrafish Proteins ,Molecular biology ,Rats ,Homeobox Protein Nkx-2.2 ,Gene Expression Regulation ,Hepatocyte Nuclear Factor 3-beta ,Trans-Activators ,FOXA2 ,Chromatin immunoprecipitation ,Chickens ,Sequence Alignment ,Transcription Factors - Abstract
The MafA transcription factor is both critical to islet beta-cell function and has a unique pancreatic cell-type-specific expression pattern. To localize the potential transcriptional regulatory region(s) involved in directing expression to the beta cell, areas of identity within the 5' flanking region of the mouse, human, and rat mafA genes were found between nucleotides -9389 and -9194, -8426 and -8293, -8118 and -7750, -6622 and -6441, -6217 and -6031, and -250 and +56 relative to the transcription start site. The identity between species was greater than 75%, with the highest found between bp -8118 and -7750 ( approximately 94%, termed region 3). Region 3 was the only upstream mammalian conserved region found in chicken mafA (88% identity). In addition, region 3 uniquely displayed beta-cell-specific activity in cell-line-based reporter assays. Important regulators of beta-cell formation and function, PDX-1, FoxA2, and Nkx2.2, were shown to specifically bind to region 3 in vivo using the chromatin immunoprecipitation assay. Mutational and functional analyses demonstrated that FoxA2 (bp -7943 to -7910), Nkx2.2 (bp -7771 to -7746), and PDX-1 (bp -8087 to -8063) mediated region 3 activation. Consistent with a role in transcription, small interfering RNA-mediated knockdown of PDX-1 led to decreased mafA mRNA production in INS-1-derived beta-cell lines (832/13 and 832/3), while MafA expression was undetected in the pancreatic epithelium of Nkx2.2 null animals. These results suggest that beta-cell-type-specific mafA transcription is principally controlled by region 3-acting transcription factors that are essential in the formation of functional beta cells.
- Published
- 2006
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