39 results on '"Monika Dieterle"'
Search Results
2. Enzymatic activity of glycosyltransferase GLT8D1 promotes human glioblastoma cell migration
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Elena I. Ilina, Camille Cialini, Dietlind L. Gerloff, Maitane Duarte Garcia-Escudero, Céline Jeanty, Marie-Laëtitia Thézénas, Antoine Lesur, Vincent Puard, François Bernardin, Alina Moter, Anne Schuster, Monika Dieterle, Anna Golebiewska, Jean-Jacques Gérardy, Gunnar Dittmar, Simone P. Niclou, Tanja Müller, and Michel Mittelbronn
- Subjects
Biochemistry ,Glycobiology ,Cell biology ,Cancer ,Science - Abstract
Summary: Glioblastoma (GBM) is the most aggressive primary brain tumor characterized by infiltrative growth of malignant glioma cells into the surrounding brain parenchyma. In this study, our analysis of GBM patient cohorts revealed a significantly higher expression of Glycosyltransferase 8 domain containing 1 (GLT8D1) compared to normal brain tissue and could be associated with impaired patient survival. Increased in vitro expression of GLT8D1 significantly enhanced migration of two different sphere-forming GBM cell lines. By in silico analysis we predicted the 3D-structure as well as the active site residues of GLT8D1. The introduction of point mutations in the predicted active site reduced its glycosyltransferase activity in vitro and consequently impaired GBM tumor cell migration. Examination of GLT8D1 interaction partners by LC-MS/MS implied proteins associated with cytoskeleton and intracellular transport as potential substrates. In conclusion, we demonstrated that the enzymatic activity of glycosyltransferase GLT8D1 promotes GBM cell migration.
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- 2022
- Full Text
- View/download PDF
3. Protocol for derivation of organoids and patient-derived orthotopic xenografts from glioma patient tumors
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Anaïs Oudin, Virginie Baus, Vanessa Barthelemy, Carina Fabian, Eliane Klein, Monika Dieterle, May Wantz, Ann-Christin Hau, Claire Dording, Amandine Bernard, Alessandro Michelucci, Yahaya A. Yabo, Georgia Kanli, Olivier Keunen, Rolf Bjerkvig, Simone P. Niclou, and Anna Golebiewska
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Cell Biology ,Cell culture ,Cell isolation ,Cancer ,Model Organisms ,Neuroscience ,Science (General) ,Q1-390 - Abstract
Summary: Tumor organoids and patient-derived orthotopic xenografts (PDOXs) are some of the most valuable pre-clinical tools in cancer research. In this protocol, we describe efficient derivation of organoids and PDOX models from glioma patient tumors. We provide detailed steps for organoid culture, intracranial implantation, and detection of tumors in the brain. We further present technical adjustments for standardized functional assays and drug testing.For complete details on the use and execution of this protocol, please refer to Golebiewska et al. (2020).
- Published
- 2021
- Full Text
- View/download PDF
4. Temozolomide-Induced RNA Interactome Uncovers Novel LncRNA Regulatory Loops in Glioblastoma
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Sabrina Fritah, Arnaud Muller, Wei Jiang, Ramkrishna Mitra, Mohamad Sarmini, Monika Dieterle, Anna Golebiewska, Tao Ye, Eric Van Dyck, Christel Herold-Mende, Zhongming Zhao, Francisco Azuaje, and Simone P. Niclou
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glioblastoma ,temozolomide ,lncRNA ,regulatory circuit ,chemoresistance ,transcriptome ,Neoplasms. Tumors. Oncology. Including cancer and carcinogens ,RC254-282 - Abstract
Resistance to chemotherapy by temozolomide (TMZ) is a major cause of glioblastoma (GBM) recurrence. So far, attempts to characterize factors that contribute to TMZ sensitivity have largely focused on protein-coding genes, and failed to provide effective therapeutic targets. Long noncoding RNAs (lncRNAs) are essential regulators of epigenetic-driven cell diversification, yet, their contribution to the transcriptional response to drugs is less understood. Here, we performed RNA-seq and small RNA-seq to provide a comprehensive map of transcriptome regulation upon TMZ in patient-derived GBM stem-like cells displaying different drug sensitivity. In a search for regulatory mechanisms, we integrated thousands of molecular associations stored in public databases to generate a background “RNA interactome”. Our systems-level analysis uncovered a coordinated program of TMZ response reflected by regulatory circuits that involve transcription factors, mRNAs, miRNAs, and lncRNAs. We discovered 22 lncRNAs involved in regulatory loops and/or with functional relevance in drug response and prognostic value in gliomas. Thus, the investigation of TMZ-induced gene networks highlights novel RNA-based predictors of chemosensitivity in GBM. The computational modeling used to identify regulatory circuits underlying drug response and prioritizing gene candidates for functional validation is applicable to other datasets.
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- 2020
- Full Text
- View/download PDF
5. Supplementary Movie S4. from Actin Cytoskeleton Remodeling Drives Breast Cancer Cell Escape from Natural Killer–Mediated Cytotoxicity
- Author
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Clément Thomas, Bassam Janji, Guy Berchem, Salem Chouaib, Jean-Paul Thiery, Monika Dieterle, Carla Pou Casellas, Rémi Petrolli, Joshua Brown-Clay, Xianqing Mao, Flora Moreau, Celine Hoffmann, Coralie Guerin, Hannah Wurzer, and Antoun Al Absi
- Abstract
Time lapse imaging movie showing typical actin cytoskeleton configurations and dynamics in MCF-7 cells upon NK cell attack. Individual Z stacks were taken every minute over a period of 77 minutes. Time lapse movies were assembled from the Z stacks in NIH ImageJ and played at a fast frame rate (5 frames per second). NK cells are stained in red (PKH26) whereas the LifeAct-mEGFP-expressing tumour cell appears in green.
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- 2023
6. Supplementary Materials and Methods from Actin Cytoskeleton Remodeling Drives Breast Cancer Cell Escape from Natural Killer–Mediated Cytotoxicity
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Clément Thomas, Bassam Janji, Guy Berchem, Salem Chouaib, Jean-Paul Thiery, Monika Dieterle, Carla Pou Casellas, Rémi Petrolli, Joshua Brown-Clay, Xianqing Mao, Flora Moreau, Celine Hoffmann, Coralie Guerin, Hannah Wurzer, and Antoun Al Absi
- Abstract
Supplementary Materials and Methods
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- 2023
7. Data from Actin Cytoskeleton Remodeling Drives Breast Cancer Cell Escape from Natural Killer–Mediated Cytotoxicity
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Clément Thomas, Bassam Janji, Guy Berchem, Salem Chouaib, Jean-Paul Thiery, Monika Dieterle, Carla Pou Casellas, Rémi Petrolli, Joshua Brown-Clay, Xianqing Mao, Flora Moreau, Celine Hoffmann, Coralie Guerin, Hannah Wurzer, and Antoun Al Absi
- Abstract
Elucidation of the underlying molecular mechanisms of immune evasion in cancer is critical for the development of immunotherapies aimed to restore and stimulate effective antitumor immunity. Here, we evaluate the role of the actin cytoskeleton in breast cancer cell resistance to cytotoxic natural killer (NK) cells. A significant fraction of breast cancer cells responded to NK-cell attack via a surprisingly rapid and massive accumulation of F-actin near the immunologic synapse, a process we termed “actin response.” Live-cell imaging provided direct evidence that the actin response is associated with tumor cell resistance to NK-cell–mediated cell death. High-throughput imaging flow cytometry analyses showed that breast cancer cell lines highly resistant to NK cells were significantly enriched in actin response-competent cells as compared with susceptible cell lines. The actin response was not associated with a defect in NK-cell activation but correlated with reduced intracellular levels of the cytotoxic protease granzyme B and a lower rate of apoptosis in target cells. Inhibition of the actin response by knocking down CDC42 or N-WASP led to a significant increase in granzyme B levels in target cells and was sufficient to convert resistant breast cancer cell lines into a highly susceptible phenotype. The actin response and its protective effects were fully recapitulated using donor-derived primary NK cells as effector cells. Together, these findings establish the pivotal role of actin remodeling in breast cancer cell resistance to NK-cell–mediated killing.Significance: These findings establish the pivotal role of the actin cytoskeleton in driving breast cancer cell resistance to natural killer cells, a subset of cytotoxic lymphocytes with important roles in innate antitumor immunity. Cancer Res; 78(19); 5631–43. ©2018 AACR.
- Published
- 2023
8. Supplementary Figures S1-S6 from Actin Cytoskeleton Remodeling Drives Breast Cancer Cell Escape from Natural Killer–Mediated Cytotoxicity
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Clément Thomas, Bassam Janji, Guy Berchem, Salem Chouaib, Jean-Paul Thiery, Monika Dieterle, Carla Pou Casellas, Rémi Petrolli, Joshua Brown-Clay, Xianqing Mao, Flora Moreau, Celine Hoffmann, Coralie Guerin, Hannah Wurzer, and Antoun Al Absi
- Abstract
Supplementary Figures S1-S6 and their legends
- Published
- 2023
9. Protocol for derivation of organoids and patient-derived orthotopic xenografts from glioma patient tumors
- Author
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Georgia Kanli, Anna Golebiewska, Yahaya A Yabo, Rolf Bjerkvig, Carina Fabian, Alessandro Michelucci, Claire Dording, Virginie Baus, Amandine Bernard, Olivier Keunen, Ann-Christin Hau, Simone P. Niclou, Monika Dieterle, May Wantz, Vanessa Barthelemy, Anais Oudin, and Eliane Klein
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Male ,Science (General) ,ved/biology.organism_classification_rank.species ,Cell Culture Techniques ,Antineoplastic Agents ,General Biochemistry, Genetics and Molecular Biology ,Mice ,03 medical and health sciences ,Q1-390 ,0302 clinical medicine ,Model Organisms ,Glioma ,Tumor Cells, Cultured ,Protocol ,medicine ,Organoid ,Animals ,Humans ,Cell isolation ,Model organism ,030304 developmental biology ,Cancer ,0303 health sciences ,General Immunology and Microbiology ,Brain Neoplasms ,ved/biology ,business.industry ,General Neuroscience ,Cell Biology ,medicine.disease ,3. Good health ,Organoids ,Cell culture ,Cancer research ,Heterografts ,Female ,Drug Screening Assays, Antitumor ,business ,030217 neurology & neurosurgery ,Neuroscience - Abstract
Summary Tumor organoids and patient-derived orthotopic xenografts (PDOXs) are some of the most valuable pre-clinical tools in cancer research. In this protocol, we describe efficient derivation of organoids and PDOX models from glioma patient tumors. We provide detailed steps for organoid culture, intracranial implantation, and detection of tumors in the brain. We further present technical adjustments for standardized functional assays and drug testing. For complete details on the use and execution of this protocol, please refer to Golebiewska et al. (2020)., Graphical abstract, Highlights • Organoids can be generated from diverse glioma patient tumors • High-grade glioma organoids give rise to patient-derived orthotopic xenografts • Serial transplantation in vivo allows for consistent expansion of human tumor cells • The adapted protocol for reconstitution of uniform organoids for functional assays, Tumor organoids and patient-derived orthotopic xenografts (PDOXs) are some of the most valuable pre-clinical tools in cancer research. In this protocol, we describe efficient derivation of organoids and PDOX models from glioma patient tumors. We provide detailed steps for organoid culture, intracranial implantation, and detection of tumors in the brain. We further present technical adjustments for standardized functional assays and drug testing.
- Published
- 2021
- Full Text
- View/download PDF
10. Enzymatic Activity of Glycosyltransferase Glt8d1 Promotes Human Glioblastoma Cells Migration
- Author
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Tanja R. Müller, Monika Dieterle, Jean-Jacques Gérardy, Anne Schuster, Michel Mittelbronn, Elena I. Ilina, Céline Jeanty, Maitane Duarte Garcia-Escudero, Camille Cialini, Simone P. Niclou, Alina Moter, Gunnar Dittmar, Dietlind L. Gerloff, Vincent Puard, and Anna Golebiewska
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biology ,Chemistry ,Brain tumor ,Cell migration ,medicine.disease ,In vitro ,Cell culture ,Glioma ,Glycosyltransferase ,Parenchyma ,biology.protein ,medicine ,Cancer research ,Cytoskeleton - Abstract
Glioblastoma (GBM) is the most aggressive primary brain tumor characterized by infiltrative migration of malignant glioma cells into the surrounding brain parenchyma. In this study, our analysis of GBM patient cohorts displayed significant higher expression of Glycosyltransferase 8 domain containing 1 ( GLT8D1 ) compared to normal brain tissue and could be associated with impaired patient survival. In vitro an increased GLT8D1 expression significantly enhanced migration of two different sphere-forming GBM cell lines. By in silico analysis we predicted the 3D-structure as well as the active site residues of GLT8D1. The introduction of point mutations in predicted active site reduced its glycosyltransferase activity in vitro and consequently impaired GBM tumor cell migration. Examination of GLT8D1 interaction partners by LC-MS/MS implied proteins associated with cytoskeleton and intracellular transport as potential substrates. In conclusion, we demonstrated that the enzymatic activity of glycosyltransferase GLT8D1 promotes GBM cell migration.
- Published
- 2021
11. AN1-type zinc finger protein 3 (ZFAND3) is a transcriptional regulator that drives Glioblastoma invasion
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Daniel Pérez-Hernández, Arnon Møldrup Knudsen, Rolf Bjerkvig, Arnaud Muller, Carina Fabian, Anna Golebiewska, Sophie Rodius, Anne Schuster, Eliane Klein, Ann Christin Hau, Petr V. Nazarov, Monika Dieterle, Barbara Klink, Gunnar Dittmar, Virginie Neirinckx, Bjarne Winther Kristensen, Simone P. Niclou, Christel Herold-Mende, and Anais Oudin
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0301 basic medicine ,General Physics and Astronomy ,Transcriptome ,Mice ,0302 clinical medicine ,Cell Movement ,Transcriptional regulation ,Nuclear protein ,Cancer ,Regulation of gene expression ,Zinc finger ,Multidisciplinary ,Brain Neoplasms ,DNA-Binding Proteins ,Gene Expression Regulation, Neoplastic ,030220 oncology & carcinogenesis ,Gene Knockdown Techniques ,Heterografts ,Fibronectins/genetics ,Cell Adhesion Molecules/genetics ,Science ,Protein domain ,Brain tumor ,Transcription Factors/genetics ,Collagen Type VI ,Biology ,General Biochemistry, Genetics and Molecular Biology ,Article ,03 medical and health sciences ,Collagen Type VI/genetics ,Protein Domains ,Cell Line, Tumor ,medicine ,Animals ,Humans ,Neoplasm Invasiveness ,Brain Neoplasms/genetics ,General Chemistry ,medicine.disease ,Glioblastoma/genetics ,nervous system diseases ,Fibronectins ,CNS cancer ,030104 developmental biology ,Cancer research ,Neoplasm Invasiveness/genetics ,Glioblastoma ,Cell Adhesion Molecules ,Nuclear localization sequence ,DNA-Binding Proteins/genetics ,Transcription Factors - Abstract
The infiltrative nature of Glioblastoma (GBM), the most aggressive primary brain tumor, critically prevents complete surgical resection and masks tumor cells behind the blood brain barrier reducing the efficacy of systemic treatment. Here, we use a genome-wide interference screen to determine invasion-essential genes and identify the AN1/A20 zinc finger domain containing protein 3 (ZFAND3) as a crucial driver of GBM invasion. Using patient-derived cellular models, we show that loss of ZFAND3 hampers the invasive capacity of GBM, whereas ZFAND3 overexpression increases motility in cells that were initially not invasive. At the mechanistic level, we find that ZFAND3 activity requires nuclear localization and integral zinc-finger domains. Our findings indicate that ZFAND3 acts within a nuclear protein complex to activate gene transcription and regulates the promoter of invasion-related genes such as COL6A2, FN1, and NRCAM. Further investigation in ZFAND3 function in GBM and other invasive cancers is warranted., Glioblastomas (GBMs) are highly invasive brain tumours, but the underlying mechanisms of GBM invasion are unclear. Here, the authors perform an RNA interference screen and identify AN1-Type Zinc Finger protein 3 (ZFAND3) as a regulator of GBM invasion, and find that it acts through the transcriptional regulation of invasion-related genes.
- Published
- 2020
12. Temozolomide-Induced RNA Interactome Uncovers Novel LncRNA Regulatory Loops in Glioblastoma
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Francisco Azuaje, Tao Ye, Zhongming Zhao, Arnaud Muller, Wei Jiang, Ramkrishna Mitra, Monika Dieterle, Anna Golebiewska, Mohamad Sarmini, Eric Van Dyck, Christel Herold-Mende, Simone P. Niclou, Sabrina Fritah, univOAK, Archive ouverte, Luxembourg Institute of Health (LIH), Vanderbilt University Medical Center [Nashville], Vanderbilt University [Nashville], Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Universität Heidelberg, The University of Texas Health Science Center at Houston (UTHealth), and University of Bergen (UiB)
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0301 basic medicine ,regulatory circuit ,Cancer Research ,[SDV.NEU.NB]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]/Neurobiology ,Gene regulatory network ,[SDV.CAN]Life Sciences [q-bio]/Cancer ,Computational biology ,temozolomide ,Biology ,Interactome ,lcsh:RC254-282 ,Article ,Transcriptome ,03 medical and health sciences ,0302 clinical medicine ,lncRNA ,[SDV.CAN] Life Sciences [q-bio]/Cancer ,microRNA ,medicine ,Gene ,Transcription factor ,Temozolomide ,glioblastoma ,[SDV.NEU.NB] Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]/Neurobiology ,RNA ,chemoresistance ,lcsh:Neoplasms. Tumors. Oncology. Including cancer and carcinogens ,030104 developmental biology ,Oncology ,030220 oncology & carcinogenesis ,[SDV.SP.PHARMA] Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology ,[SDV.SP.PHARMA]Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology ,transcriptome ,medicine.drug - Abstract
Simple Summary Glioblastoma (GBM) is the most aggressive brain tumor and most resistant to therapy. The identification of novel predictive biomarkers or targets to counteract chemoresistance, requires a better understanding of the GBM primary response to therapy. The aim of our study was to assess the molecular response of GBM to the standard of care chemotherapy by temozolomide (TMZ). We established a comprehensive map of gene expression changes after treatment and discovered that GBM cells elicit a coordinated gene expression program after chemotherapy that differs between sensitive and resistant cells. We found that a novel class of genes expressed as long non-coding RNAs (lncRNAs) is involved in gene regulatory circuits in GBM and could represent novel markers of GBM patient prognosis. By shedding light on the involvement of the non-coding genome in GBM, our results may provide new mechanistic insight on lncRNAs and their importance in chemoresistance. Abstract Resistance to chemotherapy by temozolomide (TMZ) is a major cause of glioblastoma (GBM) recurrence. So far, attempts to characterize factors that contribute to TMZ sensitivity have largely focused on protein-coding genes, and failed to provide effective therapeutic targets. Long noncoding RNAs (lncRNAs) are essential regulators of epigenetic-driven cell diversification, yet, their contribution to the transcriptional response to drugs is less understood. Here, we performed RNA-seq and small RNA-seq to provide a comprehensive map of transcriptome regulation upon TMZ in patient-derived GBM stem-like cells displaying different drug sensitivity. In a search for regulatory mechanisms, we integrated thousands of molecular associations stored in public databases to generate a background “RNA interactome”. Our systems-level analysis uncovered a coordinated program of TMZ response reflected by regulatory circuits that involve transcription factors, mRNAs, miRNAs, and lncRNAs. We discovered 22 lncRNAs involved in regulatory loops and/or with functional relevance in drug response and prognostic value in gliomas. Thus, the investigation of TMZ-induced gene networks highlights novel RNA-based predictors of chemosensitivity in GBM. The computational modeling used to identify regulatory circuits underlying drug response and prioritizing gene candidates for functional validation is applicable to other datasets.
- Published
- 2020
13. Actin Cytoskeleton Remodeling Drives Breast Cancer Cell Escape from Natural Killer–Mediated Cytotoxicity
- Author
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Céline Hoffmann, Xianqing Mao, Rémi Petrolli, Carla Pou Casellas, Antoun Al Absi, Clément Thomas, Flora Moreau, Monika Dieterle, Jean Paul Thiery, Guy Berchem, Hannah Wurzer, Salem Chouaib, Joshua D. Brown-Clay, Coralie L. Guerin, and Bassam Janji
- Subjects
Cytotoxicity, Immunologic ,0301 basic medicine ,Cancer Research ,Wiskott-Aldrich Syndrome Protein, Neuronal ,Apoptosis ,Breast Neoplasms ,macromolecular substances ,Biology ,Granzymes ,03 medical and health sciences ,0302 clinical medicine ,Immune system ,Cell Line, Tumor ,medicine ,Humans ,Cytotoxic T cell ,cdc42 GTP-Binding Protein ,Actin ,Actin remodeling ,Cancer ,Flow Cytometry ,medicine.disease ,Actin cytoskeleton ,Killer Cells, Natural ,Granzyme B ,Actin Cytoskeleton ,030104 developmental biology ,Oncology ,Drug Resistance, Neoplasm ,Cell culture ,030220 oncology & carcinogenesis ,Leukocytes, Mononuclear ,MCF-7 Cells ,Cancer research ,Female ,Peptide Hydrolases ,Signal Transduction - Abstract
Elucidation of the underlying molecular mechanisms of immune evasion in cancer is critical for the development of immunotherapies aimed to restore and stimulate effective antitumor immunity. Here, we evaluate the role of the actin cytoskeleton in breast cancer cell resistance to cytotoxic natural killer (NK) cells. A significant fraction of breast cancer cells responded to NK-cell attack via a surprisingly rapid and massive accumulation of F-actin near the immunologic synapse, a process we termed “actin response.” Live-cell imaging provided direct evidence that the actin response is associated with tumor cell resistance to NK-cell–mediated cell death. High-throughput imaging flow cytometry analyses showed that breast cancer cell lines highly resistant to NK cells were significantly enriched in actin response-competent cells as compared with susceptible cell lines. The actin response was not associated with a defect in NK-cell activation but correlated with reduced intracellular levels of the cytotoxic protease granzyme B and a lower rate of apoptosis in target cells. Inhibition of the actin response by knocking down CDC42 or N-WASP led to a significant increase in granzyme B levels in target cells and was sufficient to convert resistant breast cancer cell lines into a highly susceptible phenotype. The actin response and its protective effects were fully recapitulated using donor-derived primary NK cells as effector cells. Together, these findings establish the pivotal role of actin remodeling in breast cancer cell resistance to NK-cell–mediated killing. Significance: These findings establish the pivotal role of the actin cytoskeleton in driving breast cancer cell resistance to natural killer cells, a subset of cytotoxic lymphocytes with important roles in innate antitumor immunity. Cancer Res; 78(19); 5631–43. ©2018 AACR.
- Published
- 2018
14. CRP2, a new invadopodia actin bundling factor critically promotes breast cancer cell invasion and metastasis
- Author
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Monika Dieterle, Xianqing Mao, Antoun Al Absi, Céline Hoffmann, Bassam Janji, Anais Oudin, Guy Berchem, Flora Moreau, André Steinmetz, and Clément Thomas
- Subjects
0301 basic medicine ,Pathology ,Lung Neoplasms ,actin cytoskeleton ,Podosome ,Apoptosis ,Mice, SCID ,Metastasis ,Extracellular matrix ,Mice ,0302 clinical medicine ,Cell Movement ,Mice, Inbred NOD ,Tumor Cells, Cultured ,Pseudopodia ,Cytoskeleton ,invadopodia ,Cell migration ,LIM Domain Proteins ,invasion ,Extracellular Matrix ,Oncology ,030220 oncology & carcinogenesis ,Podosomes ,Invadopodia ,Female ,MMP-9 ,Research Paper ,medicine.medical_specialty ,Breast Neoplasms ,Biology ,03 medical and health sciences ,breast cancer ,Breast cancer ,Biomarkers, Tumor ,medicine ,Animals ,Humans ,Neoplasm Invasiveness ,Adaptor Proteins, Signal Transducing ,Cell Proliferation ,Cancer ,medicine.disease ,Actin cytoskeleton ,Xenograft Model Antitumor Assays ,Actins ,Matrix Metalloproteinases ,030104 developmental biology ,Cancer research ,LIM domain - Abstract
// Celine Hoffmann 1,* , Xianqing Mao 1,* , Monika Dieterle 1,2 , Flora Moreau 1 , Antoun Al Absi 1 , Andre Steinmetz 3 , Anais Oudin 2 , Guy Berchem 1 , Bassam Janji 1 and Clement Thomas 1 1 Laboratory of Experimental Cancer Research, Department of Oncology, Luxembourg Institute of Health, Luxembourg, Luxembourg 2 NorLux Neuro-Oncology Laboratory, Department of Oncology, Luxembourg Institute of Health, Luxembourg, Luxembourg 3 Department of Oncology, Luxembourg Institute of Health, Luxembourg, Luxembourg * These authors have contributed equally to this work Correspondence to: Clement Thomas, email: // Keywords : actin cytoskeleton, breast cancer, invadopodia, invasion, LIM domain, MMP-9 Received : October 26, 2015 Accepted : January 27, 2016 Published : February 11, 2016 Abstract A critical process underlying cancer metastasis is the acquisition by tumor cells of an invasive phenotype. At the subcellular level, invasion is facilitated by actin-rich protrusions termed invadopodia, which direct extracellular matrix (ECM) degradation. Here, we report the identification of a new cytoskeletal component of breast cancer cell invadopodia, namely cysteine-rich protein 2 (CRP2). We found that CRP2 was not or only weakly expressed in epithelial breast cancer cells whereas it was up-regulated in mesenchymal/invasive breast cancer cells. In addition, high expression of the CRP2 encoding gene CSRP2 was associated with significantly increased risk of metastasis in basal-like breast cancer patients. CRP2 knockdown significantly reduced the invasive potential of aggressive breast cancer cells, whereas it did not impair 2D cell migration. In keeping with this, CRP2-depleted breast cancer cells exhibited a reduced capacity to promote ECM degradation, and to secrete and express MMP-9, a matrix metalloproteinase repeatedly associated with cancer progression and metastasis. In turn, ectopic expression of CRP2 in weakly invasive cells was sufficient to stimulate cell invasion. Both GFP-fused and endogenous CRP2 localized to the extended actin core of invadopodia, a structure primarily made of actin bundles. Purified recombinant CRP2 autonomously crosslinked actin filaments into thick bundles, suggesting that CRP2 contributes to the formation/maintenance of the actin core. Finally, CRP2 depletion significantly reduced the incidence of lung metastatic lesions in two xenograft mouse models of breast cancer. Collectively, our data identify CRP2 as a new cytoskeletal component of invadopodia that critically promotes breast cancer cell invasion and metastasis.
- Published
- 2016
15. A LIM Domain Protein from Tobacco Involved in Actin-Bundling and Histone Gene Transcription
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Céline Hoffmann, Erwin Grill, Clément Thomas, Monika Dieterle, André Steinmetz, Marc Diederich, Wen-Hui Shen, Katrin Neumann, Danièle Moes, Sabrina Gatti, Flora Moreau, and Marc Schumacher
- Subjects
DNA, Complementary ,Transcription, Genetic ,Active Transport, Cell Nucleus ,Arp2/3 complex ,macromolecular substances ,Plant Science ,Actin cytoskeleton organization ,Histones ,DNA-binding ,Genes, Reporter ,trans-acting factors ,Tobacco ,Actin-binding protein ,Cloning, Molecular ,Promoter Regions, Genetic ,Protein Structure, Quaternary ,Cytoskeleton ,Molecular Biology ,Actin ,Plant Proteins ,LIM domain ,Cell Nucleus ,LIM ,biology ,Nicotiana tabacum ,Protoplasts ,promoter regulation ,Actin remodeling ,cytoskeleton ,histone genes ,Actin cytoskeleton ,Molecular biology ,Actins ,Protein Structure, Tertiary ,Actin Cytoskeleton ,BY-2 ,biology.protein ,Protein Multimerization ,actin ,Research Article - Abstract
The two LIM domain-containing proteins from plants (LIMs) typically exhibit a dual cytoplasmic–nuclear distribution, suggesting that, in addition to their previously described roles in actin cytoskeleton organization, they participate in nuclear processes. Using a south-western blot-based screen aimed at identifying factors that bind to plant histone gene promoters, we isolated a positive clone containing the tobacco LIM protein WLIM2 (NtWLIM2) cDNA. Using both green fluorescent protein (GFP) fusion- and immunology-based strategies, we provide clear evidence that NtWLIM2 localizes to the actin cytoskeleton, the nucleus, and the nucleolus. Interestingly, the disruption of the actin cytoskeleton by latrunculin B significantly increases NtWLIM2 nuclear fraction, pinpointing a possible novel cytoskeletal–nuclear crosstalk. Biochemical and electron microscopy experiments reveal the ability of NtWLIM2 to directly bind to actin filaments and to crosslink the latter into thick actin bundles. Electrophoretic mobility shift assays show that NtWLIM2 specifically binds to the conserved octameric cis-elements (Oct) of the Arabidopsis histone H4A748 gene promoter and that this binding largely relies on both LIM domains. Importantly, reporter-based experiments conducted in Arabidopsis and tobacco protoplasts confirm the ability of NtWLIM2 to bind to and activate the H4A748 gene promoter in live cells. Expression studies indicate the constitutive presence of NtWLIM2 mRNA and NtWLIM2 protein during tobacco BY-2 cell proliferation and cell cycle progression, suggesting a role of NtWLIM2 in the activation of basal histone gene expression. Interestingly, both live cell and in vitro data support NtWLIM2 di/oligomerization. We propose that NtWLIM2 functions as an actin-stabilizing protein, which, upon cytoskeleton remodeling, shuttles to the nucleus in order to modify gene expression.
- Published
- 2013
16. The LIM Domains of WLIM1 Define a New Class of Actin Bundling Modules
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Christophe Ampe, Christina Hofmann, Clément Thomas, Monika Dieterle, Flora Moreau, André Steinmetz, Marleen Van Troys, Sabrina Gatti, and Céline Hoffmann
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Green Fluorescent Proteins ,Arp2/3 complex ,macromolecular substances ,Filamin ,Biochemistry ,Tobacco ,Actin-binding protein ,Molecular Biology ,Cytoskeleton ,Plant Proteins ,LIM domain ,Homeodomain Proteins ,Binding Sites ,Microscopy, Confocal ,biology ,Microfilament Proteins ,Actin remodeling ,Cell Biology ,Actin cytoskeleton ,Actins ,Protein Structure, Tertiary ,Cell biology ,Actin Cytoskeleton ,Kinetics ,biology.protein ,MDia1 ,Phosphotyrosine-binding domain ,Plasmids ,Protein Binding - Abstract
Actin filament bundling, i.e. the formation of actin cables, is an important process that relies on proteins able to directly bind and cross-link subunits of adjacent actin filaments. Animal cysteine-rich proteins and their plant counterparts are two LIM domain-containing proteins that were recently suggested to define a new family of actin cytoskeleton regulators involved in actin filament bundling. We here identified the LIM domains as responsible for F-actin binding and bundling activities of the tobacco WLIM1. The deletion of one of the two LIM domains reduced significantly, but did not entirely abolish, the ability of WLIM1 to bind actin filaments. Individual LIM domains were found to interact directly with actin filaments, although with a reduced affinity compared with the native protein. Variants lacking the C-terminal or the inter-LIM domain were only weakly affected in their F-actin stabilizing and bundling activities and trigger the formation of thick cables containing tightly packed actin filaments as does the native protein. In contrast, the deletion of one of the two LIM domains negatively impacted both activities and resulted in the formation of thinner and wavier cables. In conclusion, we demonstrate that the LIM domains of WLIM1 are new autonomous actin binding and bundling modules that cooperate to confer WLIM1 high actin binding and bundling activities.
- Published
- 2007
17. Tobacco WLIM1 Is a Novel F-Actin Binding Protein Involved in Actin Cytoskeleton Remodeling
- Author
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Christophe Ampe, Marleen Van Troys, Monika Dieterle, André Steinmetz, Céline Hoffmann, Clément Thomas, Institut de biologie moléculaire des plantes (IBMP), Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA), and Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)
- Subjects
0106 biological sciences ,CROSS-LINKING ,GROWING POLLEN TUBES ,Arp2/3 complex ,BUNDLING PROTEIN ,Plant Science ,01 natural sciences ,Actin remodeling of neurons ,LIM-DOMAIN PROTEIN ,Cells, Cultured ,Research Articles ,Plant Proteins ,0303 health sciences ,biology ,Microfilament Proteins ,ARABIDOPSIS ,Tubulin Modulators ,Cell biology ,Actin Cytoskeleton ,Dinitrobenzenes ,POLYMERIZATION ,Thiazolidines ,Recombinant Fusion Proteins ,Green Fluorescent Proteins ,Molecular Sequence Data ,ORGANIZATION ,[SDV.BC]Life Sciences [q-bio]/Cellular Biology ,macromolecular substances ,CELL EXPANSION ,Actin cytoskeleton organization ,03 medical and health sciences ,FLUORESCENT PROTEIN ,Sulfanilamides ,Tobacco ,Escherichia coli ,Actin-binding protein ,NUCLEUS ,030304 developmental biology ,Biology and Life Sciences ,Actin remodeling ,Cell Biology ,Bridged Bicyclo Compounds, Heterocyclic ,Actin cytoskeleton ,Actins ,Protein Structure, Tertiary ,Plant Leaves ,Thiazoles ,biology.protein ,Latrunculin ,MDia1 ,010606 plant biology & botany - Abstract
We used confocal microscopy and in vitro analyses to show that Nicotiana tabacum WLIM1, a LIM domain protein related to animal Cys-rich proteins, is a novel actin binding protein in plants. Green fluorescent protein (GFP)–tagged WLIM1 protein accumulated in the nucleus and cytoplasm of tobacco BY2 cells. It associated predominantly with actin cytoskeleton, as demonstrated by colabeling and treatment with actin-depolymerizing latrunculin B. High-speed cosedimentation assays revealed the ability of WLIM1 to bind directly to actin filaments with high affinity. Fluorescence recovery after photobleaching and fluorescence loss in photobleaching showed a highly dynamic in vivo interaction of WLIM1-GFP with actin filaments. Expression of WLIM1-GFP in BY2 cells significantly delayed depolymerization of the actin cytoskeleton induced by latrunculin B treatment. WLIM1 also stabilized actin filaments in vitro. Importantly, expression of WLIM1-GFP in Nicotiana benthamiana leaves induces significant changes in actin cytoskeleton organization, specifically, fewer and thicker actin bundles than in control cells, suggesting that WLIM1 functions as an actin bundling protein. This hypothesis was confirmed by low-speed cosedimentation assays and direct observation of F-actin bundles that formed in vitro in the presence of WLIM1. Taken together, these data identify WLIM1 as a novel actin binding protein that increases actin cytoskeleton stability by promoting bundling of actin filaments.
- Published
- 2006
18. F-box-like domain in the polerovirus protein P0 is required for silencing suppressor function
- Author
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Pascal Genschik, Thomas Kretsch, Maghsoud Pazhouhandeh, Katia Marrocco, Bassam Berry, Esther Lechner, Monika Dieterle, Véronique Brault, Kenneth Richards, Odile Hemmer, and Véronique Ziegler-Graff
- Subjects
0106 biological sciences ,food.ingredient ,Molecular Sequence Data ,Arabidopsis ,Nicotiana benthamiana ,F-Box Motifs ,Saccharomyces cerevisiae ,01 natural sciences ,Plant Viruses ,Polerovirus ,Viral Proteins ,03 medical and health sciences ,food ,Ubiquitin ,Gene Expression Regulation, Plant ,Tobacco ,Skp1 ,Gene silencing ,Amino Acid Sequence ,Gene Silencing ,Conserved Sequence ,030304 developmental biology ,Genetics ,0303 health sciences ,Gene knockdown ,Multidisciplinary ,biology ,Arabidopsis Proteins ,fungi ,food and beverages ,Biological Sciences ,Argonaute ,biology.organism_classification ,Protein Structure, Tertiary ,RNA silencing ,Mutation ,biology.protein ,Sequence Alignment ,Protein Binding ,010606 plant biology & botany - Abstract
Plants employ small RNA-mediated posttranscriptional gene silencing as a virus defense mechanism. In response, plant viruses encode proteins that can suppress RNA silencing, but the mode of action of most such proteins is poorly understood. Here, we show that the silencing suppressor protein P0 of two Arabidopsis -infecting poleroviruses interacts by means of a conserved minimal F-box motif with Arabidopsis thaliana orthologs of S-phase kinase-related protein 1 (SKP1), a component of the SCF family of ubiquitin E3 ligases. Point mutations in the F-box-like motif abolished the P0–SKP1 ortholog interaction, diminished virus pathogenicity, and inhibited the silencing suppressor activity of P0. Knockdown of expression of a SKP1 ortholog in Nicotiana benthamiana rendered the plants resistant to polerovirus infection. Together, the results support a model in which P0 acts as an F-box protein that targets an essential component of the host posttranscriptional gene silencing machinery.
- Published
- 2006
19. Arabidopsis CUL3A and CUL3B genes are essential for normal embryogenesis
- Author
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Alexis Thomann, Pascal Genschik, Monika Dieterle, Ueli Grossniklaus, Marylin Vantard, Jacqueline Gheyeselinck, and Vladimir Brukhin
- Subjects
0106 biological sciences ,Genetics ,0303 health sciences ,Embryo ,Cell Biology ,Plant Science ,Protein degradation ,Biology ,biology.organism_classification ,01 natural sciences ,Ubiquitin ligase ,03 medical and health sciences ,Essential gene ,Arabidopsis ,biology.protein ,Arabidopsis thaliana ,Gene ,Cullin ,030304 developmental biology ,010606 plant biology & botany - Abstract
Cullin (CUL)-dependent ubiquitin ligases form a class of structurally related multisubunit enzymes that control the rapid and selective degradation of important regulatory proteins involved in cell cycle progression and development, among others. The CUL3-BTB ligases belong to this class of enzymes and despite recent findings on their molecular composition, our knowledge on their functions and substrates remains still very limited. In contrast to budding and fission yeast, CUL3 is an essential gene in metazoans. The model plant Arabidopsis thaliana encodes two related CUL3 genes, called CUL3A and CUL3B. We recently reported that cul3a loss-of-function mutants are viable but exhibit a mild flowering and light sensitivity phenotype. We investigated the spatial and temporal expression of the two CUL3 genes in reproductive tissues and found that their expression patterns are largely overlapping suggesting possible functional redundancy. Thus, we investigated the consequences on plant development of combined Arabidopsis cul3a cul3b loss-of-function mutations. Homozygous cul3b mutant plants developed normally and were fully fertile. However, the disruption of both the CUL3A and CUL3B genes reduced gametophytic transmission and caused embryo lethality. The observed embryo abortion was found to be under maternal control. Arrest of embryogenesis occurred at multiple stages of embryo development, but predominantly at the heart stage. At the cytological level, CUL3 loss-of-function mutations affected both embryo pattern formation and endosperm development.
- Published
- 2005
20. Arabidopsis AtCUL3a and AtCUL3b Form Complexes with Members of the BTB/POZ-MATH Protein Family
- Author
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Henriette Weber, Perdita Hano, Anne Bernhardt, Mark Estelle, Hanjo Hellmann, Monika Dieterle, Pascal Genschik, and Aysegül Mutlu
- Subjects
Protein family ,Physiology ,Molecular Sequence Data ,Arabidopsis ,Plant Science ,Protein structure ,Gene Expression Regulation, Plant ,Genetics ,Amino Acid Sequence ,Phylogeny ,Zinc finger ,Sequence Homology, Amino Acid ,biology ,Arabidopsis Proteins ,Cullin Proteins ,biology.organism_classification ,Protein Structure, Tertiary ,Ubiquitin ligase ,Biochemistry ,Proteasome ,Multiprotein Complexes ,biology.protein ,Sequence Alignment ,Cullin ,Research Article - Abstract
The ubiquitin proteasome pathway in plants has been shown to be important for many developmental processes. The E3 ubiquitin-protein ligases facilitate transfer of the ubiquitin moiety to substrate proteins. Many E3 ligases contain cullin proteins as core subunits. Here, we show that Arabidopsis (Arabidopsis thaliana) AtCUL3 proteins interact in yeast two-hybrid and in vitro pull-down assays with proteins containing a BTB/POZ (broad complex, tramtrack, bric-a-brac/pox virus and zinc finger) motif. By changing specific amino acid residues within the proteins, critical parts of the cullin and BTB/POZ proteins are defined that are required for these kinds of interactions. In addition, we show that AtCUL3 proteins assemble with the RING-finger protein AtRBX1 and are targets for the RUB-conjugation pathway. The analysis of AtCUL3a and AtCUL3b expression as well as several BTB/POZ-MATH genes indicates that these genes are expressed in all parts of the plant. The results presented here provide strong evidence that AtCUL3a and AtCUL3b can assemble in Arabidopsis with BTB/POZ-MATH and AtRBX1 proteins to form functional E3 ligases.
- Published
- 2005
21. Molecular and functional characterization of Arabidopsis Cullin 3A
- Author
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Monika Dieterle, Valérie Cognat, Alexis Thomann, Yves Parmentier, Rebecca Müller, Thomas Kretsch, Jean-Pierre Renou, Wen-Hui Shen, Pascal Genschik, and Gaetan Lemonnier
- Subjects
0106 biological sciences ,Genetics ,0303 health sciences ,RBX1 ,Cullin Proteins ,fungi ,Ubiquitin-Protein Ligases ,Mutant ,food and beverages ,Cell Biology ,Plant Science ,Biology ,biology.organism_classification ,01 natural sciences ,Ubiquitin ligase ,03 medical and health sciences ,Arabidopsis ,biology.protein ,Arabidopsis thaliana ,Cullin ,030304 developmental biology ,010606 plant biology & botany - Abstract
Cullin proteins, which belong to multigenic families in all eukaryotes, associate with other proteins to form ubiquitin protein ligases (E3s) that target substrates for proteolysis by the 26S proteasome. Here, we present the molecular and genetic characterization of a plant Cullin3. In contrast to fungi and animals, the genome of the model plant Arabidopsis thaliana contains two related CUL3 genes, called CUL3A and CUL3B. We found that CUL3A is ubiquitously expressed in plants and is able to interact with the ring-finger protein RBX1. A genomic search revealed the existence of at least 76 BTB-domain proteins in Arabidopsis belonging to 11 major families. Yeast two-hybrid experiments indicate that representative members of certain families are able to physically interact with both CUL3A and CUL3B, suggesting that Arabidopsis CUL3 forms E3 protein complexes with certain BTB domain proteins. In order to determine the function of CUL3A, we used a reverse genetic approach. The cul3a null mutant flowers slightly later than the control plants. Furthermore, this mutant exhibits a reduced sensitivity of the inhibition of hypocotyl growth in far-red light and miss-expresses COP1. The viability of the mutant plants suggests functional redundancy between the two CUL3 genes in Arabidopsis.
- Published
- 2004
22. A new type of mutation in phytochrome A causes enhanced light sensitivity and alters the degradation and subcellular partitioning of the photoreceptor
- Author
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Martina Krenz, Claudia Büche, Monika Dieterle, Diana Bauer, Eberhard Schäfer, and Thomas Kretsch
- Subjects
Genetics ,Phytochrome ,Mutant ,Cell Biology ,Plant Science ,Protein degradation ,Biology ,Fusion protein ,Cell biology ,Green fluorescent protein ,Phytochrome A ,Missense mutation ,Photomorphogenesis - Abstract
A specific light program consisting of multiple treatments with alternating red and far-red light pulses was used to isolate mutants in phytochrome A-dependent signal transduction pathways in Arabidopsis. Because of their phenotype, the mutants were called eid for empfindlicher im dunkelroten Licht, which means hypersensitive in far-red light. One of the isolated mutants, eid4, is a novel semi-dominant allele of the phytochrome A gene that carries a missense mutation in the chromophore-binding domain. The mutation did not change the photochemical properties of the photoreceptor, but it leads to an increased stability under light conditions that induce its rapid degradation. Fusion proteins with the green fluorescent protein exhibited clear alterations in subcellular localization of the mutated photoreceptor: The fusion protein was impaired in the formation of sequestered areas of phytochrome in the cytosol, which can explain its reduced light-dependent degradation. In contrast, the mutation stabilizes nuclear speckles (NUS) that appear late under continuous far-red light, whereas the formation of early, transiently appearing NUS remained more or less unaltered.
- Published
- 2004
23. The Negatively Acting Factors EID1 and SPA1 Have Distinct Functions in Phytochrome A-Specific Light Signaling
- Author
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Thomas Kretsch, Monika Dieterle, Claudia Büche, and Yong-Chun Zhou
- Subjects
Chlorophyll ,Light ,Physiology ,Photosynthetic Reaction Center Complex Proteins ,Mutant ,Arabidopsis ,Light-Harvesting Protein Complexes ,Cell Cycle Proteins ,Plant Science ,Biology ,Hypocotyl ,Anthocyanins ,Phytochrome A ,Genetics ,Arabidopsis thaliana ,Alleles ,Crosses, Genetic ,Plant Proteins ,Phytochrome ,Arabidopsis Proteins ,F-Box Proteins ,Nuclear Proteins ,Photosystem II Protein Complex ,biology.organism_classification ,Phenotype ,Cell biology ,DNA-Binding Proteins ,Mutation ,Photomorphogenesis ,Signal transduction ,Carrier Proteins ,Signal Transduction ,Research Article - Abstract
EID1 (empfindlicher im dunkelroten Licht) and SPA1 (suppressor of phytochrome A[phyA]-105) function as negatively acting components in phyA-specific light signaling. Mutants in the respective genes led to very similar phenotypes under weak-light conditions. To examine whether both genes are functionally redundant, detailed physiological and genetic analyses were performed witheid1 and spa1 mutants isolated from the same wild-type background. Measurements of hypocotyl elongation, anthocyanin accumulation, and Lhcb1-transcript accumulation under different light treatments demonstrated that SPA1 has a strong influence on the regulation of very low fluence responses and a weaker influence on high-irradiance responses. In contrast, EID1 severely altered high-irradiance responses and caused almost no change on very low fluence responses. Analyses on eid1 phyA-105double mutants demonstrated that EID1 could not suppress the phenotype of the weak phyA allele under continuous far-red light. Measurements on eid1 spa1 double mutants exhibited a strong interference of both genes in the regulation of hypocotyl elongation. These results indicate that EID1 and SPA1 are involved in different but interacting phyA-dependent signal transduction chains.
- Published
- 2002
24. Phytochrome E Controls Light-Induced Germination of Arabidopsis
- Author
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Monika Dieterle, Wendy M. Stoddart, Garry C. Whitelam, Eberhard Schäfer, and Lars Hennig
- Subjects
food.ingredient ,Phytochrome ,Physiology ,Plant Science ,Biology ,biology.organism_classification ,Cell biology ,Hypocotyl ,Phytochrome A ,food ,Germination ,Arabidopsis ,Botany ,Genetics ,Arabidopsis thaliana ,Photomorphogenesis ,Cotyledon - Abstract
Germination of Arabidopsis seeds is light dependent and under phytochrome control. Previously, phytochromes A and B and at least one additional, unspecified phytochrome were shown to be involved in this process. Here, we used a set of photoreceptor mutants to test whether phytochrome D and/or phytochrome E can control germination of Arabidopsis. The results show that only phytochromes B and E, but not phytochrome D, participate directly in red/far-red light (FR)-reversible germination. Unlike phytochromes B and D, phytochrome E did not inhibit phytochrome A-mediated germination. Surprisingly, phytochrome E was required for germination of Arabidopsis seeds in continuous FR. However, inhibition of hypocotyl elongation by FR, induction of cotyledon unfolding, and induction of agravitropic growth were not affected by loss of phytochrome E. Therefore, phytochrome E is not required per se for phytochrome A-mediated very low fluence responses and the high irradiance response. Immunoblotting revealed that the need of phytochrome E for germination in FR was not caused by altered phytochrome A levels. These results uncover a novel role of phytochrome E in plant development and demonstrate the considerable functional diversification of the closely related phytochromes B, D, and E.
- Published
- 2002
25. Human muscle LIM protein dimerizes along the actin cytoskeleton and cross-links actin filaments
- Author
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Flora Moreau, Michèle Moes, Katrin Neumann, Emeline Goretti, Carole Luthold, Monika Dieterle, André Steinmetz, Clément Thomas, and Céline Hoffmann
- Subjects
Sarcomeres ,Arp2/3 complex ,Muscle Proteins ,macromolecular substances ,Cell Line ,Actin remodeling of neurons ,Mice ,Stress Fibers ,Animals ,Humans ,Myocytes, Cardiac ,Actin-binding protein ,Molecular Biology ,Actin ,LIM domain ,Muscle Cells ,biology ,Microfilament Proteins ,Actin remodeling ,Cell Biology ,Articles ,LIM Domain Proteins ,Actin cytoskeleton ,Cell biology ,Actin Cytoskeleton ,biology.protein ,MDia1 ,Protein Multimerization ,Cardiomyopathies ,Protein Binding ,Signal Transduction - Abstract
The muscle LIM protein (MLP) is a nucleocytoplasmic shuttling protein playing important roles in the regulation of myocyte remodeling and adaptation to hypertrophic stimuli. Missense mutations in human MLP or its ablation in transgenic mice promotes cardiomyopathy and heart failure. The exact function(s) of MLP in the cytoplasmic compartment and the underlying molecular mechanisms remain largely unknown. Here, we provide evidence that MLP autonomously binds to, stabilizes, and bundles actin filaments (AFs) independently of calcium and pH. Using total internal reflection fluorescence microscopy, we have shown how MLP cross-links actin filaments into both unipolar and mixed-polarity bundles. Quantitative analysis of the actin cytoskeleton configuration confirmed that MLP substantially promotes actin bundling in live myoblasts. In addition, bimolecular fluorescence complementation (BiFC) assays revealed MLP self-association. Remarkably, BiFC complexes mostly localize along actin filament-rich structures, such as stress fibers and sarcomeres, supporting a functional link between MLP self-association and actin cross-linking. Finally, we have demonstrated that MLP self-associates through its N-terminal LIM domain, whereas it binds to AFs through its C-terminal LIM domain. Together our data support that MLP contributes to the maintenance of cardiomyocyte cytoarchitecture by a mechanism involving its self-association and actin filament cross-linking.
- Published
- 2014
26. Live cell imaging reveals actin-cytoskeleton-induced self-association of the actin-bundling protein WLIM1
- Author
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Céline Hoffmann, Daniéle Moes, Monika Dieterle, Katrin Neumann, Flora Moreau, Angela Tavares Furtado, Dominique Dumas, André Steinmetz, Clément Thomas, Centre de Recherche Publique- Santé, Université du Luxembourg (Uni.lu), Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), and Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS)
- Subjects
0303 health sciences ,Binding Sites ,030302 biochemistry & molecular biology ,Green Fluorescent Proteins ,Arabidopsis ,Cell Biology ,macromolecular substances ,LIM Domain Proteins ,Plants, Genetically Modified ,Cell Line ,03 medical and health sciences ,Actin Cytoskeleton ,Tobacco ,[SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM] ,[SDV.MHEP]Life Sciences [q-bio]/Human health and pathology ,030304 developmental biology ,Protein Binding - Abstract
International audience; Crosslinking of actin filaments into bundles is essential for the assembly and stabilization of specific cytoskeletal structures. However, relatively little is known about the molecular mechanisms underlying actin bundle formation. The two LIM-domain-containing proteins define a novel and evolutionarily conserved family of actin-bundling proteins whose actin-binding and -crosslinking activities primarily rely on their LIM domains. Using TIRF microscopy, we describe real-time formation of actin bundles induced by tobacco NtWLIM1 in vitro. We show that NtWLIM1 binds to single filaments and subsequently promotes their interaction and zippering into tight bundles of mixed polarity. NtWLIM1-induced bundles grew by both elongation of internal filaments and addition of preformed fragments at their extremities. Importantly, these data are highly consistent with the modes of bundle formation and growth observed in transgenic Arabidopsis plants expressing a GFP-fused Arabidopsis AtWLIM1 protein. Using two complementary live cell imaging approaches, a close relationship between NtWLIM1 subcellular localization and self-association was established. Indeed, both BiFC and FLIM-FRET data revealed that, although unstable NtWLIM1 complexes can sporadically form in the cytosol, stable complexes concentrate along the actin cytoskeleton. Remarkably, disruption of the actin cytoskeleton significantly impaired self-association of NtWLIM1. In addition, biochemical analyses support the idea that F-actin facilitates the switch of purified recombinant NtWLIM1 from a monomeric to a di- or oligomeric state. On the basis of our data, we propose a model in which actin binding promotes the formation and stabilization of NtWLIM1 complexes, which in turn might drive the crosslinking of actin filaments.
- Published
- 2014
27. Live cell imaging approaches reveal actin cytoskeleton-induced self-association of the actin-bundling protein WLIM1
- Author
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Flora Moreau, Angela Tavares Furtado, Dominique Dumas, Danièle Moes, Katrin Neumann, Monika Dieterle, André Steinmetz, Clément Thomas, and Céline Hoffmann
- Subjects
0106 biological sciences ,0303 health sciences ,Actin remodeling ,Arp2/3 complex ,macromolecular substances ,Cell Biology ,Biology ,Filamin ,Actin cytoskeleton ,01 natural sciences ,Cell biology ,03 medical and health sciences ,Actin remodeling of neurons ,biology.protein ,MDia1 ,Actin-binding protein ,Cytoskeleton ,030304 developmental biology ,010606 plant biology & botany - Abstract
Crosslinking of actin filaments into bundles is essential for the assembly and stabilization of specific cytoskeletal structures. However, relatively little is known about the molecular mechanisms underlying actin bundle formation. The two LIM-domain-containing proteins define a novel and evolutionarily conserved family of actin-bundling proteins whose actin-binding and -crosslinking activities primarily rely on their LIM domains. Using TIRF microscopy, we describe real-time formation of actin bundles induced by tobacco NtWLIM1 in vitro. We show that NtWLIM1 binds to single filaments and subsequently promotes their interaction and zippering into tight bundles of mixed polarity. NtWLIM1-induced bundles grew by both elongation of internal filaments and addition of preformed fragments at their extremities. Importantly, these data are highly consistent with the modes of bundle formation and growth observed in transgenic Arabidopsis plants expressing a GFP-fused Arabidopsis AtWLIM1 protein. Using two complementary live cell imaging approaches, a close relationship between NtWLIM1 subcellular localization and self-association was established. Indeed, both BiFC and FLIM-FRET data revealed that, although unstable NtWLIM1 complexes can sporadically form in the cytosol, stable complexes concentrate along the actin cytoskeleton. Remarkably, disruption of the actin cytoskeleton significantly impaired self-association of NtWLIM1. In addition, biochemical analyses support the idea that F-actin facilitates the switch of purified recombinant NtWLIM1 from a monomeric to a di- or oligomeric state. On the basis of our data, we propose a model in which actin binding promotes the formation and stabilization of NtWLIM1 complexes, which in turn might drive the crosslinking of actin filaments.
- Published
- 2013
28. Arabidopsis actin-depolymerizing factors (ADFs) 1 and 9 display antagonist activities
- Author
-
Clément Thomas, Danièle Moes, Karthik Arumugam, Stéphane Tholl, Flora Moreau, Katrin Neumann, Céline Hoffmann, Monika Dieterle, and André Steinmetz
- Subjects
Biophysics ,Arabidopsis ,Arp2/3 complex ,macromolecular substances ,Biochemistry ,Actin remodeling of neurons ,Structural Biology ,Tobacco ,Genetics ,Actin-binding protein ,Cytoskeleton ,Molecular Biology ,Actin ,biology ,Arabidopsis Proteins ,Actin remodeling ,Cell Biology ,Cofilin ,Actin cytoskeleton ,Cell biology ,Actin Cytoskeleton ,Actin Depolymerizing Factors ,biology.protein ,MDia1 ,Actin bundling ,ADF - Abstract
We provide evidence that one of the 11 Arabidopsis actin-depolymerizing factors (ADFs), namely ADF9, does not display typical F-actin depolymerizing activity. Instead, ADF9 effectively stabilizes actin filaments in vitro and concomitantly bundles actin filaments with the highest efficiency under acidic conditions. Competition experiments show that ADF9 antagonizes ADF1 activity by reducing its ability to potentiate F-actin depolymerization. Accordingly, ectopic expression of ADF1 and ADF9 in tobacco cells has opposite effects. ADF1 severs actin filaments/bundles and promotes actin cytoskeleton disassembly, whereas ADF9 induces the formation of long bundles. Together these data reveal an additional degree of complexity in the comprehension of the biological functions of the ADF family and illustrate that antagonist activities can be displayed by seemingly equivalent actin-binding proteins.
- Published
- 2011
29. Arabidopsis LIM Proteins: A Family of Actin Bundlers with Distinct Expression Patterns and Modes of Regulation[W][OA]
- Author
-
Céline Hoffmann, Danièle Moes, Flora Moreau, Stéphane Tholl, Jessica Papuga, Clément Thomas, Monika Dieterle, and André Steinmetz
- Subjects
animal structures ,Arabidopsis ,Arp2/3 complex ,Plant Science ,macromolecular substances ,Pollen Tube ,Filamentous actin ,Actin cytoskeleton organization ,Gene Expression Regulation, Plant ,Actin-binding protein ,Research Articles ,biology ,Arabidopsis Proteins ,Microfilament Proteins ,Actin remodeling ,Cell Biology ,Hydrogen-Ion Concentration ,Actin cytoskeleton ,Plants, Genetically Modified ,Actins ,Cell biology ,body regions ,Actin Cytoskeleton ,Profilin ,embryonic structures ,biology.protein ,Calcium ,MDia1 ,Carrier Proteins - Abstract
Recently, a number of two LIM-domain containing proteins (LIMs) have been reported to trigger the formation of actin bundles, a major higher-order cytoskeletal assembly. Here, we analyzed the six Arabidopsis thaliana LIM proteins. Promoter-β-glucuronidase reporter studies revealed that WLIM1, WLIM2a, and WLIM2b are widely expressed, whereas PLIM2a, PLIM2b, and PLIM2c are predominantly expressed in pollen. LIM-green fluorescent protein (GFP) fusions all decorated the actin cytoskeleton and increased actin bundle thickness in transgenic plants and in vitro, although with different affinities and efficiencies. Remarkably, the activities of WLIMs were calcium and pH independent, whereas those of PLIMs were inhibited by high pH and, in the case of PLIM2c, by high [Ca2+]. Domain analysis showed that the C-terminal domain is key for the responsiveness of PLIM2c to pH and calcium. Regulation of LIM by pH was further analyzed in vivo by tracking GFP-WLIM1 and GFP-PLIM2c during intracellular pH modifications. Cytoplasmic alkalinization specifically promoted release of GFP-PLIM2c but not GFP-WLIM1, from filamentous actin. Consistent with these data, GFP-PLIM2c decorated long actin bundles in the pollen tube shank, a region of relatively low pH. Together, our data support a prominent role of Arabidopsis LIM proteins in the regulation of actin cytoskeleton organization and dynamics in sporophytic tissues and pollen.
- Published
- 2010
30. Actin bundling in plants
- Author
-
Monika Dieterle, André Steinmetz, Danièle Moes, Stéphane Tholl, Clément Thomas, Jessica Papuga, and Flora Moreau
- Subjects
biology ,fungi ,Microfilament Proteins ,food and beverages ,Arp2/3 complex ,Actin remodeling ,Cytoplasmic Streaming ,macromolecular substances ,Cell Biology ,Cell Enlargement ,Plants ,Microfilament ,Actin cytoskeleton ,Actin cytoskeleton organization ,Actins ,Cell biology ,Profilin ,Structural Biology ,Formins ,biology.protein ,Protein Multimerization ,Cytoskeleton ,Protein Structure, Quaternary ,Cell Division - Abstract
Tight regulation of plant actin cytoskeleton organization and dynamics is crucial for numerous cellular processes including cell division, expansion and intracellular trafficking. Among the various actin regulatory proteins, actin-bundling proteins trigger the formation of bundles composed of several parallel actin filaments closely packed together. Actin bundles are present in virtually all plant cells, but their biological roles have rarely been addressed directly. However, decades of research in the plant cytoskeleton field yielded a bulk of data from which an overall picture of the functions supplied by actin bundles in plant cells emerges. Although plants lack several equivalents of animal actin-bundling proteins, they do possess major bundler classes including fimbrins, villins and formins. The existence of additional players is not excluded as exemplified by the recent characterization of plant LIM proteins, which trigger the formation of actin bundles both in vitro and in vivo. This apparent functional redundancy likely reflects the need for plant cells to engineer different types of bundles that act at different sub-cellular locations and exhibit specific function-related properties. By surveying information regarding the properties of plant actin bundles and their associated bundling proteins, the present review aims at clarifying why and how plants make actin bundles.
- Published
- 2009
31. Actin bundling via LIM domains
- Author
-
Céline Hoffmann, Flora Moreau, Clément Thomas, Sabrina Gatti, Monika Dieterle, André Steinmetz, and Jessica Papuga
- Subjects
biology ,Cytoskeleton organization ,Plant Science ,macromolecular substances ,Bioinformatics ,Actin cytoskeleton ,Cell biology ,Article Addendum ,Gene expression ,biology.protein ,Actin-binding protein ,Cytoskeleton ,Actin ,Function (biology) ,LIM domain - Abstract
The LIM domain is defined as a protein-protein interaction module involved in the regulation of diverse cellular processes including gene expression and cytoskeleton organization. We have recently shown that the tobacco WLIM1, a two LIM domain-containing protein, is able to bind to, stabilize and bundle actin filaments, suggesting that it participates to the regulation of actin cytoskeleton structure and dynamics. In the December issue of the Journal of Biological Chemistry we report a domain analysis that specifically ascribes the actin-related activities of WLIM1 to its two LIM domains. Results suggest that LIM domains function synergistically in the full-length protein to achieve optimal activities. Here we briefly summarize relevant data regarding the actin-related properties/functions of two LIM domain-containing proteins in plants and animals. In addition, we provide further evidence of cooperative effects between LIM domains by transiently expressing a chimeric multicopy WLIM1 protein in BY2 cells.
- Published
- 2007
32. CUL4 associates with DDB1 and DET1 and its downregulation affects diverse aspects of development in Arabidopsis thaliana
- Author
-
Monika Dieterle, Pascal Genschik, Chris Bowler, Anne Bernhardt, Verena Schade, Marco Anders, Giovanna Benvenuto, Esther Lechner, Hanjo Hellmann, Manu J. Dubin, and Perdita Hano
- Subjects
Light ,RBX1 ,Arabidopsis ,Down-Regulation ,Plant Science ,Plant Roots ,DDB1 ,Gene Expression Regulation, Plant ,Genetics ,Arabidopsis thaliana ,biology ,Arabidopsis Proteins ,Cullin Proteins ,Intracellular Signaling Peptides and Proteins ,Gene Expression Regulation, Developmental ,Nuclear Proteins ,Cell Biology ,biology.organism_classification ,Ubiquitin ligase ,DNA-Binding Proteins ,Plant Leaves ,Proteasome ,biology.protein ,Carrier Proteins ,Cullin - Abstract
Cullins are central scaffolding subunits in eukaryotic E3 ligases that facilitate the ubiquitination of target proteins. Arabidopsis contains at least 11 cullin proteins but only a few of them have been assigned biological roles. In this work Arabidopsis cullin 4 is shown to assemble with DDB1, RBX1, DET1 and DDB2 in vitro and in planta. In addition, by using T-DNA insertion and CUL4 antisense lines we demonstrate that corresponding mutants are severely affected in different aspects of development. Reduced CUL4 expression leads to a reduced number of lateral roots, and to abnormal vascular tissue and stomatal development. Furthermore, cul4 mutants display a weak constitutive photomorphogenic phenotype. These results therefore assign an important function to CUL4 during plant development and provide strong evidence that CUL4 assembles together with RBX1 and DDB1 proteins to form a functional E3 ligase in Arabidopsis.
- Published
- 2006
33. Functional analysis of EID1, an F-box protein involved in phytochrome A-dependent light signal transduction
- Author
-
Thorsten Stolpe, Martina Krenz, Markus Funk, Katia Marrocco, Erik Bury, Pascal Genschik, Thomas Kretsch, Yong-Chun Zhou, Monika Dieterle, Institut de biologie moléculaire des plantes (IBMP), and Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)
- Subjects
0106 biological sciences ,Light ,Recombinant Fusion Proteins ,Molecular Sequence Data ,Arabidopsis ,Plant Science ,Flowers ,Saccharomyces cerevisiae ,01 natural sciences ,F-box protein ,03 medical and health sciences ,Phytochrome A ,Bimolecular fluorescence complementation ,SCF complex ,Two-Hybrid System Techniques ,Genetics ,Amino Acid Sequence ,Nuclear protein ,Alleles ,030304 developmental biology ,0303 health sciences ,biology ,Phytochrome ,Sequence Homology, Amino Acid ,Arabidopsis Proteins ,F-Box Proteins ,Nuclear Proteins ,Cell Biology ,biology.organism_classification ,3. Good health ,Cell biology ,Protein Structure, Tertiary ,DNA-Binding Proteins ,Plant Leaves ,Protein Subunits ,Biochemistry ,Microscopy, Fluorescence ,Seedlings ,biology.protein ,Photomorphogenesis ,Sequence Alignment ,010606 plant biology & botany ,Signal Transduction - Abstract
Empfindlicher im Dunkelroten Licht 1 (EID1) is an F-box protein that functions as a negative regulator in phytochrome A (phyA)-specific light signalling. F-box proteins are components of SCF ubiquitin ligase complexes that target proteins for degradation in the proteasome. Here we present further characterization of EID1 at the expression level, and show that it regulates photomorphogenesis in seedlings, rosette leaf development and flowering. Data on transcript expression patterns indicate that EID1 is expressed during all stages of Arabidopsis development and exhibits no light response. Microscope studies demonstrate that EID1 is localized to the nucleus, where it can form speckles under continuous far-red light that resemble clastosomes. To characterize the composition and formation of SCF(EID1) complexes further, we used two-hybrid and bridge assays in yeast and in planta. EID1 interacts specifically with several Arabidopsis Skp1-like (ASK) proteins and Cullin1 to form stable dimeric and trimeric complexes. Our results support a two-step association process in which the F-box protein binds first to the ASK adaptor, forming a unit which then associates with the catalytic core of the SCF complex. Finally, our data indicate that the EID1 target interaction domain is composed of two independent modules.
- Published
- 2006
34. Molecular and functional characterization of Arabidopsis Cullin 3A
- Author
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Monika, Dieterle, Alexis, Thomann, Jean-Pierre, Renou, Yves, Parmentier, Valerie, Cognat, Gaetan, Lemonnier, Rebecca, Müller, Wen-Hui, Shen, Thomas, Kretsch, and Pascal, Genschik
- Subjects
Light ,Arabidopsis Proteins ,Arabidopsis ,Genes, Homeobox ,Flowers ,Saccharomyces cerevisiae ,Cullin Proteins ,Hypocotyl ,Protein Structure, Tertiary ,Phenotype ,Gene Expression Regulation, Plant ,Multigene Family ,Two-Hybrid System Techniques ,Mutation ,Carrier Proteins ,Phylogeny - Abstract
Cullin proteins, which belong to multigenic families in all eukaryotes, associate with other proteins to form ubiquitin protein ligases (E3s) that target substrates for proteolysis by the 26S proteasome. Here, we present the molecular and genetic characterization of a plant Cullin3. In contrast to fungi and animals, the genome of the model plant Arabidopsis thaliana contains two related CUL3 genes, called CUL3A and CUL3B. We found that CUL3A is ubiquitously expressed in plants and is able to interact with the ring-finger protein RBX1. A genomic search revealed the existence of at least 76 BTB-domain proteins in Arabidopsis belonging to 11 major families. Yeast two-hybrid experiments indicate that representative members of certain families are able to physically interact with both CUL3A and CUL3B, suggesting that Arabidopsis CUL3 forms E3 protein complexes with certain BTB domain proteins. In order to determine the function of CUL3A, we used a reverse genetic approach. The cul3a null mutant flowers slightly later than the control plants. Furthermore, this mutant exhibits a reduced sensitivity of the inhibition of hypocotyl growth in far-red light and miss-expresses COP1. The viability of the mutant plants suggests functional redundancy between the two CUL3 genes in Arabidopsis.
- Published
- 2005
35. A new type of mutation in phytochrome A causes enhanced light sensitivity and alters the degradation and subcellular partitioning of the photoreceptor
- Author
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Monika, Dieterle, Diana, Bauer, Claudia, Büche, Martina, Krenz, Eberhard, Schäfer, and Thomas, Kretsch
- Subjects
Phenotype ,Base Sequence ,Light ,Arabidopsis Proteins ,Phytochrome A ,Recombinant Fusion Proteins ,Molecular Sequence Data ,Photosynthetic Reaction Center Complex Proteins ,Arabidopsis ,Mutation, Missense ,Amino Acid Sequence ,Phytochrome ,Protein Serine-Threonine Kinases - Abstract
A specific light program consisting of multiple treatments with alternating red and far-red light pulses was used to isolate mutants in phytochrome A-dependent signal transduction pathways in Arabidopsis. Because of their phenotype, the mutants were called eid for empfindlicher im dunkelroten Licht, which means hypersensitive in far-red light. One of the isolated mutants, eid4, is a novel semi-dominant allele of the phytochrome A gene that carries a missense mutation in the chromophore-binding domain. The mutation did not change the photochemical properties of the photoreceptor, but it leads to an increased stability under light conditions that induce its rapid degradation. Fusion proteins with the green fluorescent protein exhibited clear alterations in subcellular localization of the mutated photoreceptor: The fusion protein was impaired in the formation of sequestered areas of phytochrome in the cytosol, which can explain its reduced light-dependent degradation. In contrast, the mutation stabilizes nuclear speckles (NUS) that appear late under continuous far-red light, whereas the formation of early, transiently appearing NUS remained more or less unaltered.
- Published
- 2004
36. Characterization of a novel non-constitutive photomorphogenic cop1 allele
- Author
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Monika Dieterle, Eberhard Schäfer, Thomas Kretsch, and Claudia Büche
- Subjects
Light ,Physiology ,Ubiquitin-Protein Ligases ,Mutant ,Molecular Sequence Data ,Arabidopsis ,Plant Science ,Polymerase Chain Reaction ,Consensus Sequence ,Genetics ,Ring finger ,medicine ,Morphogenesis ,Arabidopsis thaliana ,Amino Acid Sequence ,Allele ,Gene ,DNA Primers ,Phytochrome ,biology ,Base Sequence ,Arabidopsis Proteins ,fungi ,Darkness ,biology.organism_classification ,RING finger domain ,Focus Issue on Light Signaling ,medicine.anatomical_structure ,Amino Acid Substitution ,Mutagenesis, Site-Directed ,Photomorphogenesis - Abstract
A specific light program consisting of multiple treatments with alternating red and far-red light pulses was used to isolate mutants in phytochrome A-dependent signal transduction in Arabidopsis seedlings. Because of their phenotype, the mutants were called eid (empfindlicher im dunkelroten Licht, which means hypersensitive in far-red light). One of the isolated mutants, eid6, is a novel recessive allele of the COP1 gene (constitutive photomorphogenic 1) that carries an amino acid transition in a conserved histidine residue of the RING finger domain. Mutant seedlings exhibited an extreme hypersensitivity towards all tested light qualities, but in contrast to known cop1 alleles, no constitutive photomorphogenic phenotype was detectable in darkness. Thus, the novel cop1eid6 allele seems to encode for a protein whose remaining activity is sufficient for the suppression of photomorphogenesis in dark-grown plants. In adult cop1eid6 plants, the development of the Cop1 phenotype is dominated by phytochrome B. Comparison of the phenotype of the novel cop1eid6 and the weak cop1-4 allele under continuous far-red light indicates that the RING finger and coiled-coil domains of COP1 are sufficient for some specific regulatory function in phytochrome A-dependent high irradiance responses.
- Published
- 2003
37. EID1, an F-box protein involved in phytochrome A-specific light signaling
- Author
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Yong-Chun Zhou, Markus Funk, Eberhard Schäfer, Monika Dieterle, and Thomas Kretsch
- Subjects
Leucine zipper ,Light ,Green Fluorescent Proteins ,Molecular Sequence Data ,Genes, Recessive ,Plasma protein binding ,F-box protein ,DNA-binding protein ,Phytochrome A ,Research Communication ,Transformation, Genetic ,Two-Hybrid System Techniques ,Genetics ,Tissue Distribution ,Amino Acid Sequence ,Nuclear protein ,Cloning, Molecular ,Alleles ,Plant Proteins ,Cell Nucleus ,Leucine Zippers ,SKP Cullin F-Box Protein Ligases ,biology ,Phytochrome ,Base Sequence ,Sequence Homology, Amino Acid ,Arabidopsis Proteins ,F-Box Proteins ,Nuclear Proteins ,Cell biology ,DNA-Binding Proteins ,Luminescent Proteins ,Phenotype ,Microscopy, Fluorescence ,Mutation ,biology.protein ,Signal transduction ,Developmental Biology ,Protein Binding ,Signal Transduction - Abstract
To perceive red and far-red light, plants have evolved specific photoreceptors called phytochromes. Even though the spectral properties of all phytochromes are very similar, they show a distinct mode of action. Here we describe EID1, a negatively acting component of the signaling cascade that shifts the responsiveness of the phytochrome A (phyA) signaling system associated with hypocotyl elongation from red to far-red wavelengths. EID1 is a novel nuclear F-box protein that contains a leucine zipper whose integrity is necessary for its biological function. EID1 most probably acts by targeting activated components of the phyA signaling pathway to ubiquitin-dependent proteolysis.
- Published
- 2001
38. The pH sensibility of actin-bundling LIM proteins is governed by the acidic properties of their C-terminal domain
- Author
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Monika Dieterle, Céline Hoffmann, Flora Moreau, Clément Thomas, Angela Tavares Furtado, Katrin Neumann, André Steinmetz, Danièle Moes, and Jessica Papuga
- Subjects
Amino Acid Motifs ,Molecular Sequence Data ,Arabidopsis ,Biophysics ,Peptide ,Biochemistry ,Proto-Oncogene Proteins c-myc ,pH responsiveness ,Gene Expression Regulation, Plant ,Structural Biology ,Genetics ,Protein Isoforms ,Amino Acid Sequence ,Cytoskeleton ,Molecular Biology ,Sequence Deletion ,LIM domain ,chemistry.chemical_classification ,biology ,Arabidopsis Proteins ,C-terminus ,Cell Biology ,Hydrogen-Ion Concentration ,LIM Domain Proteins ,biology.organism_classification ,Actins ,Actin bundling ,Protein Structure, Tertiary ,c-Myc ,chemistry ,Domain (ring theory) ,Pollen ,Trans-acting ,Actin-bundling - Abstract
Actin-bundling Arabidopsis LIM proteins are subdivided into two subfamilies differing in their pH sensitivity. Widely-expressed WLIMs are active under low and high physiologically-relevant pH conditions, whereas pollen-enriched PLIMs are inactivated by pH values above 6.8. By a domain swapping approach we identified the C-terminal (Ct) domain of PLIMs as the domain responsible for pH responsiveness. Remarkably, this domain conferred pH sensitivity to LIM proteins, when provided “in trans” (i.e., as a single, independent, peptide), indicating that it operates through the interaction with another domain. An acidic 6xc-Myc peptide functionally mimicked the Ct domain of PLIMs and efficiently inhibited LIM actin bundling activity under high pH conditions. Together, our data suggest a model where PLIMs are regulated by an intermolecular interaction between their acidic Ct domain and another, yet unidentified, domain.
- Full Text
- View/download PDF
39. Plant CULLIN-based E3s: Phytohormones come first
- Author
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Alexis Thomann, Pascal Genschik, and Monika Dieterle
- Subjects
Ubiquitin-Protein Ligases ,Biophysics ,Arabidopsis ,Signalling ,Biochemistry ,Ubiquitin ,Plant Growth Regulators ,Structural Biology ,Phytohormone ,Plant Cells ,Genetics ,Arabidopsis thaliana ,Molecular Biology ,biology ,Cell Cycle ,fungi ,CUL-dependent ubiquitin ligase ,food and beverages ,Cell Biology ,Plants ,biology.organism_classification ,Cullin Proteins ,Ubiquitin ligase ,Cell biology ,biology.protein ,Signal transduction ,Ubiquitin protein ligases ,Cullin ,Function (biology) ,Signal Transduction - Abstract
CULLIN (CUL)-dependent ubiquitin ligases form a class of structurally related multi-subunit enzymes that control the rapid and selective degradation of important regulatory proteins involved in cell cycle progression and development, among others. Several classes of these E3s are also conserved in plants and genetic analyses, using Arabidopsis thaliana, indicate that they play an important function during plant development and responses to the environment. In this review, we will discuss the molecular composition and function of these enzymes in plants with a major emphasis on phytohormone signal transduction pathways.
- Full Text
- View/download PDF
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