77 results on '"Carotta S"'
Search Results
2. NK cell development in bone marrow and liver: site matters
- Author
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Gotthardt, D, Prchal-Murphy, M, Seillet, C, Glasner, A, Mandelboim, O, Carotta, S, Sexl, V, and Putz, E M
- Published
- 2014
- Full Text
- View/download PDF
3. SCON - A Short Conditional intrON for conditional knockout with one-step zygote injection
- Author
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Hallay N, Hans-Christian Theussl, Heetak Lee, Andrea Pauli, Szép-Bakonyi R, Zhu Y, Jihoon Kim, Bon-Kyoung Koo, Boese A, Wu Ss, Gabriele Colozza, Pilat-Carotta S, and Krista R. Gert
- Subjects
Zygote ,Conditional gene knockout ,Intron ,CRISPR ,Computational biology ,Allele ,Biology ,Gene - Abstract
The generation of conditional alleles using CRISPR technology is still challenging. Here, we introduce a Short Conditional intrON (SCON, 189 bp) that enables rapid generation of conditional alleles via one-step zygote injection. SCON has conditional intronic function in various vertebrate species and its target insertion is as simple as CRISPR/Cas9-mediated gene tagging.
- Published
- 2021
4. Cell cycle dependence of gene transfer by lipoplex, polyplex and recombinant adenovirus
- Author
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Brunner, S, Sauer, T, Carotta, S, Cotten, M, Saltik, M, and Wagner, E
- Published
- 2000
- Full Text
- View/download PDF
5. Role of "Cancer Stem Cells" and Cell Survival in Tumor Development and Maintenance
- Author
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Adams, J.M, primary, Kelly, P.N., additional, Dakic, A., additional, Carotta, S., additional, Nutt, S.L., additional, and Strasser, A., additional
- Published
- 2008
- Full Text
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6. Red2Flpe-SCON: a versatile, multicolor strategy for generating mosaic conditional knockout mice.
- Author
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Wu SS, Kim S, Lee H, Lee JH, Park SY, Bakonyi R, Teriyapirom I, Hallay N, Pilat-Carotta S, Theussl HC, Kim J, Lee JH, Simons BD, Kim JK, Colozza G, and Koo BK
- Subjects
- Animals, Mice, Mosaicism, Cell Differentiation, Cell Proliferation genetics, Esophagus metabolism, Esophagus pathology, Cell Lineage genetics, Introns genetics, Female, Male, SOXB1 Transcription Factors genetics, SOXB1 Transcription Factors metabolism, Mice, Knockout, Red Fluorescent Protein, Luminescent Proteins genetics, Luminescent Proteins metabolism
- Abstract
Image-based lineage tracing enables tissue turnover kinetics and lineage potentials of different adult cell populations to be investigated. Previously, we reported a genetic mouse model system, Red2Onco, which ectopically expressed mutated oncogenes together with red fluorescent proteins (RFP). This system enabled the expansion kinetics and neighboring effects of oncogenic clones to be dissected. We now report Red2Flpe-SCON: a mosaic knockout system that uses multicolor reporters to label both mutant and wild-type cells. We develop the Red2Flpe mouse line for red clone-specific Flpe expression, as well as the FRT-based SCON (Short Conditional IntrON) method to facilitate tunable conditional mosaic knockouts in mice. We use the Red2Flpe-SCON method to study Sox2 mutant clonal analysis in the esophageal epithelium of adult mice which reveal that the stem cell gene, Sox2, is less essential for adult stem cell maintenance itself, but rather for stem cell proliferation and differentiation., (© 2024. The Author(s).)
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- 2024
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7. Author Correction: CIS is a potent checkpoint in NK cell-mediated tumor immunity.
- Author
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Delconte RB, Kolesnik TB, Dagley LF, Rautela J, Shi W, Putz EM, Stannard K, Zhang JG, Teh C, Firth M, Ushiki T, Andoniou CE, Degli-Esposti MA, Sharp PP, Sanvitale CE, Infusini G, Liau NPD, Linossi EM, Burns CJ, Carotta S, Gray DHD, Seillet C, Hutchinson DS, Belz GT, Webb AI, Alexander WS, Li SS, Bullock AN, Babon JJ, Smyth MJ, Nicholson SE, and Huntington ND
- Published
- 2024
- Full Text
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8. Discovery of BI 7446: A Potent Cyclic Dinucleotide STING Agonist with Broad-Spectrum Variant Activity for the Treatment of Cancer.
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Kuttruff CA, Fleck M, Carotta S, Arnhof H, Bretschneider T, Dahmann G, Gremel G, Grube A, Handschuh S, Heimann A, Hofmann MH, Impagnatiello MA, Nar H, Rast G, Schaaf O, Schmidt E, and Oost T
- Subjects
- Animals, Mice, Immunotherapy, Neoplasms pathology, Nucleotides, Cyclic pharmacology, Membrane Proteins agonists
- Abstract
Activating the stimulator of interferon genes (STING) pathway with STING agonists is an attractive immune oncology concept to treat patients with tumors that are refractory to single-agent anti-PD-1 therapy. For best clinical translatability and broad application to cancer patients, STING agonists with potent cellular activation of all STING variants are desired. Novel cyclic dinucleotide (CDN)-based selective STING agonists were designed and synthesized comprising noncanonical nucleobase, ribose, and phosphorothioate moieties. This strategy led to the discovery of 2',3'-CDN 13 (BI 7446), which features unprecedented potency and activates all five STING variants in cellular assays. ADME profiling revealed that CDN 13 has attractive drug-like properties for development as an intratumoral agent. Injection of low doses of CDN 13 into tumors in mice induced long-lasting, tumor-specific immune-mediated tumor rejection. Based on its compelling preclinical profile, BI 7446 has been advanced to clinical trials (monotherapy and in combination with anti-PD-1 antibody).
- Published
- 2023
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9. Author Correction: SCON-a Short Conditional intrON for conditional knockout with one-step zygote injection.
- Author
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Wu SS, Lee H, Szép-Bakonyi R, Colozza G, Boese A, Gert KR, Hallay N, Lee JH, Kim J, Zhu Y, Linssen MM, Pilat-Carotta S, Hohenstein P, Theussl HC, Pauli A, and Koo BK
- Published
- 2023
- Full Text
- View/download PDF
10. Tumor microenvironment mimicking 3D models unveil the multifaceted effects of SMAC mimetics.
- Author
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Pinto C, Slavic-Obradovic K, Fürweger D, Thaler B, Souabni A, Carotta S, Aichinger M, Reiser U, Impagnatiello MA, and Tirapu I
- Abstract
Small molecule IAP antagonists - SMAC mimetics (SM) - are being developed as an anticancer therapy. SM therapy was demonstrated not only to sensitize tumor cells to TNFα-mediated cell death but also to exert immunostimulatory properties. Their good safety and tolerability profile, plus promising preclinical data, warrants further investigation into their various effects within the tumor microenvironment. Using in vitro models of human tumor cells and fibroblast spheroids co-cultured with primary immune cells, we investigated the effects of SM on immune cell activation. SM treatment induces the maturation of human PBMC- and patient-derived dendritic cells (DC), and modulates cancer-associated fibroblasts towards an immune interacting phenotype. Finally, SM-induced tumor necroptosis further enhances DC activation, leading also to higher T-cell activation and infiltration into the tumor site. These results highlight the relevance of using heterotypic in vitro models to investigate the effects of targeted therapies on different components of the tumor microenvironment., Competing Interests: All authors were employees of Boehringer Ingelheim RCV GmbH & Co KG at the time the study was conducted. The SMAC mimetic compound used in this work is included in a patent filed by Boehringer Ingelheim International GmbH. The authors declare no other competing interests., (© 2023 The Author(s).)
- Published
- 2023
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11. SCON-a Short Conditional intrON for conditional knockout with one-step zygote injection.
- Author
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Wu SS, Lee H, Szép-Bakonyi R, Colozza G, Boese A, Gert KR, Hallay N, Lee JH, Kim J, Zhu Y, Linssen MM, Pilat-Carotta S, Hohenstein P, Theussl HC, Pauli A, and Koo BK
- Subjects
- Mice, Animals, Introns genetics, Gene Knockout Techniques, CRISPR-Cas Systems genetics, Zygote
- Abstract
The generation of conditional alleles using CRISPR technology is still challenging. Here, we introduce a Short Conditional intrON (SCON, 189 bp) that enables the rapid generation of conditional alleles via one-step zygote injection. In this study, a total of 13 SCON mouse lines were successfully generated by 2 different laboratories. SCON has conditional intronic functions in various vertebrate species, and its target insertion is as simple as CRISPR/Cas9-mediated gene tagging., (© 2022. The Author(s).)
- Published
- 2022
- Full Text
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12. p57 Kip2 imposes the reserve stem cell state of gastric chief cells.
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Lee JH, Kim S, Han S, Min J, Caldwell B, Bamford AD, Rocha ASB, Park J, Lee S, Wu SS, Lee H, Fink J, Pilat-Carotta S, Kim J, Josserand M, Szep-Bakonyi R, An Y, Ju YS, Philpott A, Simons BD, Stange DE, Choi E, Koo BK, and Kim JK
- Subjects
- Animals, Cell Lineage, Mice, Organoids, Stem Cells, Stomach, Chief Cells, Gastric metabolism, Cyclin-Dependent Kinase Inhibitor p57 metabolism
- Abstract
Adult stem cells constantly react to local changes to ensure tissue homeostasis. In the main body of the stomach, chief cells produce digestive enzymes; however, upon injury, they undergo rapid proliferation for prompt tissue regeneration. Here, we identified p57
Kip2 (p57) as a molecular switch for the reserve stem cell state of chief cells in mice. During homeostasis, p57 is constantly expressed in chief cells but rapidly diminishes after injury, followed by robust proliferation. Both single-cell RNA sequencing and dox-induced lineage tracing confirmed the sequential loss of p57 and activation of proliferation within the chief cell lineage. In corpus organoids, p57 overexpression induced a long-term reserve stem cell state, accompanied by altered niche requirements and a mature chief cell/secretory phenotype. Following the constitutive expression of p57 in vivo, chief cells showed an impaired injury response. Thus, p57 is a gatekeeper that imposes the reserve stem cell state of chief cells in homeostasis., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)- Published
- 2022
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13. Functional and molecular characterization of PD1 + tumor-infiltrating lymphocytes from lung cancer patients.
- Author
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Lipp JJ, Wang L, Yang H, Yao F, Harrer N, Müller S, Berezowska S, Dorn P, Marti TM, Schmid RA, Hegedüs B, Souabni A, Carotta S, Pearson MA, Sommergruber W, Kocher GJ, and Hall SRR
- Subjects
- CTLA-4 Antigen, Humans, Lymphocytes, Tumor-Infiltrating metabolism, Lymphocytes, Tumor-Infiltrating pathology, Programmed Cell Death 1 Receptor genetics, Carcinoma, Non-Small-Cell Lung genetics, Carcinoma, Non-Small-Cell Lung therapy, Lung Neoplasms genetics, Lung Neoplasms therapy
- Abstract
Antibody-mediated cancer immunotherapy targets inhibitory surface molecules, such as PD1, PD-L1, and CTLA-4, aiming to re-invigorate dysfunctional T cells. We purified and characterized tumor-infiltrating lymphocytes (TILs) and their patient-matched non-tumor counterparts from treatment-naïve NSCLC patient biopsies to evaluate the effect of PD1 expression on the functional and molecular profiles of tumor-resident T cells. We show that PD1+ CD8+ TILs have elevated expression of the transcriptional regulator ID3 and that the cytotoxic potential of CD8 T cells can be improved by knocking down ID3, defining it as a potential regulator of T cell effector function. PD1+ CD4+ memory TILs display transcriptional patterns consistent with both helper and regulator function, but can robustly facilitate B cell activation and expansion. Furthermore, we show that expanding ex vivo-prepared TILs in vitro broadly preserves their functionality with respect to tumor cell killing, B cell help, and TCR repertoire. Although purified PD1+ CD8+ TILs generally maintain an exhausted phenotype upon expansion in vitro, transcriptional analysis reveals a downregulation of markers of T-cell dysfunction, including the co-inhibitory molecules PD1 and CTLA-4 and transcription factors ID3, TOX and TOX2, while genes involved in cell cycle and DNA repair are upregulated. We find reduced expression of WNT signaling components to be a hallmark of PD1+ CD8+ exhausted T cells in vivo and in vitro and demonstrate that restoring WNT signaling, by pharmacological blockade of GSK3β, can improve effector function. These data unveil novel targets for tumor immunotherapy and have promising implications for the development of a personalized TIL-based cell therapy for lung cancer., Competing Interests: No potential conflict of interest was reported by the author(s)., (© 2022 The Author(s). Published with license by Taylor & Francis Group, LLC.)
- Published
- 2022
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14. Acquired resistance to anti-MAPK targeted therapy confers an immune-evasive tumor microenvironment and cross-resistance to immunotherapy in melanoma.
- Author
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Haas L, Elewaut A, Gerard CL, Umkehrer C, Leiendecker L, Pedersen M, Krecioch I, Hoffmann D, Novatchkova M, Kuttke M, Neumann T, da Silva IP, Witthock H, Cuendet MA, Carotta S, Harrington KJ, Zuber J, Scolyer RA, Long GV, Wilmott JS, Michielin O, Vanharanta S, Wiesner T, and Obenauf AC
- Subjects
- Animals, Humans, Immune Evasion, Immunologic Factors therapeutic use, Immunotherapy, Mice, Neoplasm Recurrence, Local, Protein Kinase Inhibitors pharmacology, Melanoma drug therapy, Tumor Microenvironment
- Abstract
How targeted therapies and immunotherapies shape tumors, and thereby influence subsequent therapeutic responses, is poorly understood. In the present study, we show, in melanoma patients and mouse models, that when tumors relapse after targeted therapy with MAPK pathway inhibitors, they are cross-resistant to immunotherapies, despite the different modes of action of these therapies. We find that cross-resistance is mediated by a cancer cell-instructed, immunosuppressive tumor microenvironment that lacks functional CD103
+ dendritic cells, precluding an effective T cell response. Restoring the numbers and functionality of CD103+ dendritic cells can re-sensitize cross-resistant tumors to immunotherapy. Cross-resistance does not arise from selective pressure of an immune response during evolution of resistance, but from the MAPK pathway, which not only is reactivated, but also exhibits an increased transcriptional output that drives immune evasion. Our work provides mechanistic evidence for cross-resistance between two unrelated therapies, and a scientific rationale for treating patients with immunotherapy before they acquire resistance to targeted therapy., (© 2021. The Author(s), under exclusive licence to Springer Nature America, Inc.)- Published
- 2021
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15. Cutibacterium acnes Infection Induces Type I Interferon Synthesis Through the cGAS-STING Pathway.
- Author
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Fischer K, Tschismarov R, Pilz A, Straubinger S, Carotta S, McDowell A, and Decker T
- Subjects
- Adaptor Proteins, Vesicular Transport metabolism, Cyclic AMP metabolism, Humans, Immunity, Innate, Inflammasomes metabolism, NF-kappa B metabolism, Signal Transduction, THP-1 Cells, Acne Vulgaris immunology, Gram-Positive Bacterial Infections immunology, Interferon Type I metabolism, Macrophages immunology, Membrane Proteins metabolism, Nucleotidyltransferases metabolism, Propionibacteriaceae physiology
- Abstract
Cutibacterium (previously Propionibacterium ) acnes is an anaerobic, Gram-positive commensal of the human body. The bacterium has been associated with a variety of diseases, including acne vulgaris, prosthetic joint infections, prostate cancer, and sarcoidosis. The accumulation of C. acnes in diseases such as acne and prostate cancer has been shown to correlate with enhanced inflammation. While the C. acnes -induced proinflammatory axis, via NF-κB and MAPK signaling and inflammasome activation, has been investigated over the last few decades, the potential role of C. acnes in triggering the type I interferon (IFN-I) pathway has not been addressed. Our results show that C. acnes induces the IFN-I signaling axis in human macrophages by triggering the cGAS-STING pathway. In addition, IFN-I signaling induced by C. acnes strongly depends on the adapter protein TRIF in a non-canonical manner; these signaling events occurred in the absence of any detectable intracellular replication of the bacterium. Collectively, our results provide important insight into C. acnes -induced intracellular signaling cascades in human macrophages and suggest IFN-I as a factor in the etiology of C. acnes -induced diseases. This knowledge may be valuable for developing novel therapies targeting C. acnes in diseases where the accumulation of the bacterium leads to an inflammatory pathology., (Copyright © 2020 Fischer, Tschismarov, Pilz, Straubinger, Carotta, McDowell and Decker.)
- Published
- 2020
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16. Selective and Potent CDK8/19 Inhibitors Enhance NK-Cell Activity and Promote Tumor Surveillance.
- Author
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Hofmann MH, Mani R, Engelhardt H, Impagnatiello MA, Carotta S, Kerenyi M, Lorenzo-Herrero S, Böttcher J, Scharn D, Arnhof H, Zoephel A, Schnitzer R, Gerstberger T, Sanderson MP, Rajgolikar G, Goswami S, Vasu S, Ettmayer P, Gonzalez S, Pearson M, McConnell DB, Kraut N, Muthusamy N, and Moll J
- Subjects
- Animals, Apoptosis, Breast Neoplasms enzymology, Breast Neoplasms immunology, Breast Neoplasms pathology, Cell Proliferation, Cytotoxicity, Immunologic immunology, Female, Humans, Killer Cells, Natural drug effects, Leukemia, Myeloid, Acute enzymology, Leukemia, Myeloid, Acute immunology, Leukemia, Myeloid, Acute pathology, Melanoma, Experimental enzymology, Melanoma, Experimental immunology, Melanoma, Experimental pathology, Mice, Mice, Inbred C57BL, Mice, Inbred NOD, Mice, SCID, Phosphorylation, STAT1 Transcription Factor metabolism, Tumor Cells, Cultured, Xenograft Model Antitumor Assays, Breast Neoplasms drug therapy, Cyclin-Dependent Kinase 8 antagonists & inhibitors, Cyclin-Dependent Kinases antagonists & inhibitors, Killer Cells, Natural immunology, Leukemia, Myeloid, Acute drug therapy, Melanoma, Experimental drug therapy, Protein Kinase Inhibitors pharmacology
- Abstract
Natural killer (NK) cells play a pivotal role in controlling cancer. Multiple extracellular receptors and internal signaling nodes tightly regulate NK activation. Cyclin-dependent kinases of the mediator complex (CDK8 and CDK19) were described as a signaling intermediates in NK cells. Here, we report for the first time the development and use of CDK8/19 inhibitors to suppress phosphorylation of STAT1
S727 in NK cells and to augment the production of the cytolytic molecules perforin and granzyme B (GZMB). Functionally, this resulted in enhanced NK-cell-mediated lysis of primary leukemia cells. Treatment with the CDK8/19 inhibitor BI-1347 increased the response rate and survival of mice bearing melanoma and breast cancer xenografts. In addition, CDK8/19 inhibition augmented the antitumoral activity of anti-PD-1 antibody and SMAC mimetic therapy, both agents that promote T-cell-mediated antitumor immunity. Treatment with the SMAC mimetic compound BI-8382 resulted in an increased number of NK cells infiltrating EMT6 tumors. Combination of the CDK8/19 inhibitor BI-1347, which augments the amount of degranulation enzymes, with the SMAC mimetic BI-8382 resulted in increased survival of mice carrying the EMT6 breast cancer model. The observed survival benefit was dependent on an intermittent treatment schedule of BI-1347, suggesting the importance of circumventing a hyporesponsive state of NK cells. These results suggest that CDK8/19 inhibitors can be combined with modulators of the adaptive immune system to inhibit the growth of solid tumors, independent of their activity on cancer cells, but rather through promoting NK-cell function., (©2020 American Association for Cancer Research.)- Published
- 2020
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17. Radiotherapy as a Backbone for Novel Concepts in Cancer Immunotherapy.
- Author
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Kabiljo J, Harpain F, Carotta S, and Bergmann M
- Abstract
Radiation-induced immunogenic cell death has been described to contribute to the efficacy of external beam radiotherapy in local treatment of solid tumors. It is well established that radiation therapy can induce immunogenic cell death in cancer cells under certain conditions. Initial clinical studies combining radiotherapy with immunotherapies suggest a synergistic potential of this approach. Improving our understanding of how radiation reconditions the tumor immune microenvironment should pave the way for designing rational and robust combinations with immunotherapeutic drugs that enhance both local and systemic anti-cancer immune effects. In this review, we summarize irradiation-induced types of immunogenic cell death and their effects on the tumor microenvironment. We discuss preclinical insights on mechanisms and benefits of combining radiotherapy with immunotherapy, focusing on immune checkpoint inhibitors. In addition, we elaborate how these observations were translated into clinical studies and which parameters may be optimized to achieve best results in future clinical trials.
- Published
- 2019
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18. Landscape and Dynamics of Single Immune Cells in Hepatocellular Carcinoma.
- Author
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Zhang Q, He Y, Luo N, Patel SJ, Han Y, Gao R, Modak M, Carotta S, Haslinger C, Kind D, Peet GW, Zhong G, Lu S, Zhu W, Mao Y, Xiao M, Bergmann M, Hu X, Kerkar SP, Vogt AB, Pflanz S, Liu K, Peng J, Ren X, and Zhang Z
- Subjects
- Carcinoma, Hepatocellular genetics, Carcinoma, Hepatocellular pathology, Cell Communication genetics, Cell Communication immunology, Cell Lineage genetics, Cell Lineage immunology, Dendritic Cells immunology, Dendritic Cells pathology, Gene Expression Regulation, Neoplastic, Humans, Inflammation genetics, Inflammation pathology, Leukocyte Common Antigens immunology, Liver immunology, Liver pathology, Liver Neoplasms genetics, Liver Neoplasms pathology, Lymph Nodes immunology, Lymph Nodes pathology, Lymphocytes immunology, Lymphocytes pathology, Lysosomal Membrane Proteins genetics, Macrophages immunology, Macrophages pathology, Myeloid Cells immunology, Myeloid Cells pathology, Neoplasm Proteins genetics, Sequence Analysis, RNA, Single-Cell Analysis, Transcriptome genetics, Transcriptome immunology, Tumor Microenvironment genetics, Tumor Microenvironment immunology, Carcinoma, Hepatocellular immunology, Cation Transport Proteins genetics, Inflammation immunology, Liver Neoplasms immunology, Membrane Glycoproteins genetics
- Abstract
The immune microenvironment of hepatocellular carcinoma (HCC) is poorly characterized. Combining two single-cell RNA sequencing technologies, we produced transcriptomes of CD45
+ immune cells for HCC patients from five immune-relevant sites: tumor, adjacent liver, hepatic lymph node (LN), blood, and ascites. A cluster of LAMP3+ dendritic cells (DCs) appeared to be the mature form of conventional DCs and possessed the potential to migrate from tumors to LNs. LAMP3+ DCs also expressed diverse immune-relevant ligands and exhibited potential to regulate multiple subtypes of lymphocytes. Of the macrophages in tumors that exhibited distinct transcriptional states, tumor-associated macrophages (TAMs) were associated with poor prognosis, and we established the inflammatory role of SLC40A1 and GPNMB in these cells. Further, myeloid and lymphoid cells in ascites were predominantly linked to tumor and blood origins, respectively. The dynamic properties of diverse CD45+ cell types revealed by this study add new dimensions to the immune landscape of HCC., (Copyright © 2019 Elsevier Inc. All rights reserved.)- Published
- 2019
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19. LMP2 immunoproteasome promotes lymphocyte survival by degrading apoptotic BH3-only proteins.
- Author
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Zanker D, Pang K, Oveissi S, Lu C, Faou P, Nowell C, Mbogo GW, Carotta S, Quillici C, Karupiah G, Hibbs ML, Nutt SL, Neeson P, Puthalakath H, and Chen W
- Subjects
- Animals, Blotting, Western, Cell Line, Cell Survival, Cells, Cultured, Cysteine Endopeptidases deficiency, Female, Mice, Mice, Inbred C57BL, Mice, Knockout, Proteasome Endopeptidase Complex deficiency, BH3 Interacting Domain Death Agonist Protein immunology, Cysteine Endopeptidases immunology, Lymphocytes immunology, Proteasome Endopeptidase Complex immunology
- Abstract
The role of the immunoproteasome is perceived as confined to adaptive immune responses given its ability to produce peptides ideal for MHC Class-I binding. Here, we demonstrate that the immunoproteasome subunit, LMP2, has functions beyond its immunomodulatory role. Using LMP2-deficient mice, we demonstrate that LMP2 is crucial for lymphocyte development and survival in the periphery. Moreover, LMP2-deficient lymphocytes show impaired degradation of key BH3-only proteins, resulting in elevated levels of pro-apoptotic BIM and increased cell death. Interestingly, LMP2 is the sole immunoproteasome subunit required for BIM degradation. Together, our results suggest LMP2 has important housekeeping functions and represents a viable therapeutic target for cancer., (© 2018 Australasian Society for Immunology Inc.)
- Published
- 2018
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20. Publisher Correction: Mutations in Vps15 perturb neuronal migration in mice and are associated with neurodevelopmental disease in humans.
- Author
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Gstrein T, Edwards A, Přistoupilová A, Leca I, Breuss M, Pilat-Carotta S, Hansen AH, Tripathy R, Traunbauer AK, Hochstoeger T, Rosoklija G, Repic M, Landler L, Stránecký V, Dürnberger G, Keane TM, Zuber J, Adams DJ, Flint J, Honzik T, Gut M, Beltran S, Mechtler K, Sherr E, Kmoch S, Gut I, and Keays DA
- Abstract
In the supplementary information PDF originally posted, there were discrepancies from the integrated supplementary information that appeared in the HTML; the former has been corrected as follows. In the legend to Supplementary Fig. 2c, "major organs of the mouse" has been changed to "major organs of the adult mouse." In the legend to Supplementary Fig. 6d,h, "At E14.5 Mbe/Mbe mutants have a smaller percentage of Brdu positive cells in bin 3" has been changed to "At E14.5 Mbe/Mbe mutants have a higher percentage of Brdu positive cells in bin 3."
- Published
- 2018
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21. PU.1 Is Required for the Developmental Progression of Multipotent Progenitors to Common Lymphoid Progenitors.
- Author
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Pang SHM, de Graaf CA, Hilton DJ, Huntington ND, Carotta S, Wu L, and Nutt SL
- Subjects
- Animals, Biomarkers, Colony-Forming Units Assay, Gene Expression, Gene Expression Regulation, Developmental, Genes, Reporter, Hematopoietic Stem Cells cytology, Hematopoietic Stem Cells metabolism, Immunophenotyping, Lymphopoiesis genetics, Mice, Mice, Knockout, Proto-Oncogene Proteins metabolism, Trans-Activators metabolism, Transcriptional Activation, Cell Differentiation genetics, Lymphoid Progenitor Cells cytology, Lymphoid Progenitor Cells metabolism, Multipotent Stem Cells cytology, Multipotent Stem Cells metabolism, Proto-Oncogene Proteins genetics, Trans-Activators genetics
- Abstract
The transcription factor PU.1 is required for the development of mature myeloid and lymphoid cells. Due to this essential role and the importance of PU.1 in regulating several signature markers of lymphoid progenitors, its precise function in early lymphopoiesis has been difficult to define. Here, we demonstrate that PU.1 was required for efficient generation of lymphoid-primed multipotent progenitors (LMPPs) from hematopoietic stem cells and was essential for the subsequent formation of common lymphoid progenitors (CLPs). By contrast, further differentiation into the B-cell lineage was independent of PU.1. Examination of the transcriptional changes in conditional progenitors revealed that PU.1 activates lymphoid genes in LMPPs, while repressing genes normally expressed in neutrophils. These data identify PU.1 as a critical regulator of lymphoid priming and the transition between LMPPs and CLPs.
- Published
- 2018
- Full Text
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22. HOXB4 Promotes Hemogenic Endothelium Formation without Perturbing Endothelial Cell Development.
- Author
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Teichweyde N, Kasperidus L, Carotta S, Kouskoff V, Lacaud G, Horn PA, Heinrichs S, and Klump H
- Subjects
- Animals, Cell Differentiation physiology, Cells, Cultured, Core Binding Factor Alpha 2 Subunit metabolism, Embryonic Stem Cells metabolism, Embryonic Stem Cells physiology, Gene Expression Profiling methods, Hematopoietic Stem Cells metabolism, Hematopoietic Stem Cells physiology, Mice, Pluripotent Stem Cells metabolism, Pluripotent Stem Cells physiology, Transcription, Genetic physiology, Up-Regulation physiology, Endothelial Cells metabolism, Endothelial Cells physiology, Endothelium metabolism, Endothelium physiology, Hematopoiesis physiology, Homeodomain Proteins metabolism, Transcription Factors metabolism
- Abstract
Generation of hematopoietic stem cells (HSCs) from pluripotent stem cells, in vitro, holds great promise for regenerative therapies. Primarily, this has been achieved in mouse cells by overexpression of the homeotic selector protein HOXB4. The exact cellular stage at which HOXB4 promotes hematopoietic development, in vitro, is not yet known. However, its identification is a prerequisite to unambiguously identify the molecular circuits controlling hematopoiesis, since the activity of HOX proteins is highly cell and context dependent. To identify that stage, we retrovirally expressed HOXB4 in differentiating mouse embryonic stem cells (ESCs). Through the use of Runx1
(-/-) ESCs containing a doxycycline-inducible Runx1 coding sequence, we uncovered that HOXB4 promoted the formation of hemogenic endothelium cells without altering endothelial cell development. Whole-transcriptome analysis revealed that its expression mediated the upregulation of transcription of core transcription factors necessary for hematopoiesis, culminating in the formation of blood progenitors upon initiation of Runx1 expression., (Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.)- Published
- 2018
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23. Mutations in Vps15 perturb neuronal migration in mice and are associated with neurodevelopmental disease in humans.
- Author
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Gstrein T, Edwards A, Přistoupilová A, Leca I, Breuss M, Pilat-Carotta S, Hansen AH, Tripathy R, Traunbauer AK, Hochstoeger T, Rosoklija G, Repic M, Landler L, Stránecký V, Dürnberger G, Keane TM, Zuber J, Adams DJ, Flint J, Honzik T, Gut M, Beltran S, Mechtler K, Sherr E, Kmoch S, Gut I, and Keays DA
- Subjects
- Alkylating Agents toxicity, Animals, Animals, Newborn, Atrophy chemically induced, Atrophy genetics, Atrophy pathology, Autophagy drug effects, Autophagy genetics, Brain drug effects, Brain pathology, Cell Movement drug effects, Disease Models, Animal, Embryo, Mammalian, Ethylnitrosourea toxicity, Female, Gene Expression Regulation, Developmental genetics, Humans, Male, Mice, Mice, Inbred C57BL, Mice, Transgenic, Neurons drug effects, Neurons ultrastructure, Signal Transduction drug effects, Signal Transduction genetics, Vacuolar Proton-Translocating ATPases drug effects, Cell Movement genetics, Gene Expression Regulation, Developmental drug effects, Mutation drug effects, Neurodevelopmental Disorders chemically induced, Neurodevelopmental Disorders diagnostic imaging, Neurodevelopmental Disorders genetics, Neurodevelopmental Disorders pathology, Neurons pathology, Vacuolar Proton-Translocating ATPases genetics
- Abstract
The formation of the vertebrate brain requires the generation, migration, differentiation and survival of neurons. Genetic mutations that perturb these critical cellular events can result in malformations of the telencephalon, providing a molecular window into brain development. Here we report the identification of an N-ethyl-N-nitrosourea-induced mouse mutant characterized by a fractured hippocampal pyramidal cell layer, attributable to defects in neuronal migration. We show that this is caused by a hypomorphic mutation in Vps15 that perturbs endosomal-lysosomal trafficking and autophagy, resulting in an upregulation of Nischarin, which inhibits Pak1 signaling. The complete ablation of Vps15 results in the accumulation of autophagic substrates, the induction of apoptosis and severe cortical atrophy. Finally, we report that mutations in VPS15 are associated with cortical atrophy and epilepsy in humans. These data highlight the importance of the Vps15-Vps34 complex and the Nischarin-Pak1 signaling hub in the development of the telencephalon.
- Published
- 2018
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24. MOZ (KAT6A) is essential for the maintenance of classically defined adult hematopoietic stem cells.
- Author
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Sheikh BN, Yang Y, Schreuder J, Nilsson SK, Bilardi R, Carotta S, McRae HM, Metcalf D, Voss AK, and Thomas T
- Subjects
- Animals, Biomarkers metabolism, Bone Marrow Cells pathology, Cell Count, Cell Differentiation, Cellular Senescence, Colony-Forming Units Assay, Gene Deletion, Integrases metabolism, Mice, Inbred C57BL, Phenotype, Resting Phase, Cell Cycle, Stem Cell Transplantation, Adult Stem Cells cytology, Adult Stem Cells metabolism, Hematopoietic Stem Cells cytology, Hematopoietic Stem Cells metabolism, Histone Acetyltransferases metabolism
- Abstract
Hematopoietic stem cells (HSCs) are conventionally thought to be at the apex of a hierarchy that produces all mature cells of the blood. The quintessential property of these cells is their ability to reconstitute the entire hematopoietic system of hemoablated recipients. This characteristic has enabled HSCs to be used to replenish the hematopoietic system of patients after chemotherapy or radiotherapy. Here, we use deletion of the monocytic leukemia zinc finger gene (Moz/Kat6a/Myst3) to examine the effects of removing HSCs. Loss of MOZ in adult mice leads to the rapid loss of HSCs as defined by transplantation. This is accompanied by a reduction of the LSK-CD48
- CD150+ and LSK-CD34- Flt3- populations in the bone marrow and a reduction in quiescent cells in G0 Surprisingly, the loss of classically defined HSCs did not affect mouse viability, and there was no recovery of the LSK-CD48- CD150+ and LSK-CD34- Flt3- populations 15 to 18 months after Moz deletion. Clonal analysis of myeloid progenitors, which produce short-lived granulocytes, demonstrate that these are derived from cells that had undergone recombination at the Moz locus up to 2 years earlier, suggesting that early progenitors have acquired extended self-renewal. Our results establish that there are essential differences in HSC requirement for steady-state blood cell production compared with the artificial situation of reconstitution after transplantation into a hemoablated host. A better understanding of steady-state hematopoiesis may facilitate the development of novel therapies engaging hematopoietic cell populations with previously unrecognized traits, as well as characterizing potential vulnerability to oncogenic transformation., (© 2016 by The American Society of Hematology.)- Published
- 2016
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- View/download PDF
25. Deciphering the Innate Lymphoid Cell Transcriptional Program.
- Author
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Seillet C, Mielke LA, Amann-Zalcenstein DB, Su S, Gao J, Almeida FF, Shi W, Ritchie ME, Naik SH, Huntington ND, Carotta S, and Belz GT
- Subjects
- Animals, Basic-Leucine Zipper Transcription Factors immunology, Bone Marrow Cells immunology, Cell Differentiation genetics, Cell Differentiation immunology, Cell Lineage immunology, Gene Expression Regulation, Hepatocyte Nuclear Factor 1-alpha immunology, Immunity, Innate genetics, Killer Cells, Natural immunology, Mice, Programmed Cell Death 1 Receptor immunology, Transcription Factors genetics, Transcription Factors immunology, Basic-Leucine Zipper Transcription Factors genetics, Hepatocyte Nuclear Factor 1-alpha genetics, Immunity, Innate immunology, Lymphocytes immunology, Programmed Cell Death 1 Receptor genetics
- Abstract
Innate lymphoid cells (ILCs) are enriched at mucosal surfaces, where they provide immune surveillance. All ILC subsets develop from a common progenitor that gives rise to pre-committed progenitors for each of the ILC lineages. Currently, the temporal control of gene expression that guides the emergence of these progenitors is poorly understood. We used global transcriptional mapping to analyze gene expression in different ILC progenitors. We identified PD-1 to be specifically expressed in PLZF
+ ILCp and revealed that the timing and order of expression of the transcription factors NFIL3, ID2, and TCF-1 was critical. Importantly, induction of ILC lineage commitment required only transient expression of NFIL3 prior to ID2 and TCF-1 expression. These findings highlight the importance of the temporal program that permits commitment of progenitors to the ILC lineage, and they expand our understanding of the core transcriptional program by identifying potential regulators of ILC development., (Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.)- Published
- 2016
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- View/download PDF
26. CIS is a potent checkpoint in NK cell-mediated tumor immunity.
- Author
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Delconte RB, Kolesnik TB, Dagley LF, Rautela J, Shi W, Putz EM, Stannard K, Zhang JG, Teh C, Firth M, Ushiki T, Andoniou CE, Degli-Esposti MA, Sharp PP, Sanvitale CE, Infusini G, Liau NP, Linossi EM, Burns CJ, Carotta S, Gray DH, Seillet C, Hutchinson DS, Belz GT, Webb AI, Alexander WS, Li SS, Bullock AN, Babon JJ, Smyth MJ, Nicholson SE, and Huntington ND
- Subjects
- Animals, Cell Proliferation genetics, Cytotoxicity, Immunologic genetics, Immunologic Surveillance, Interferon-gamma metabolism, Interleukin-15 metabolism, Janus Kinase 1 metabolism, Lymphocyte Activation genetics, Melanoma, Experimental, Mice, Mice, Inbred C57BL, Mice, Knockout, Molecular Targeted Therapy, Neoplasms immunology, Signal Transduction genetics, Suppressor of Cytokine Signaling Proteins genetics, Immunotherapy methods, Killer Cells, Natural immunology, Neoplasms therapy, Suppressor of Cytokine Signaling Proteins metabolism
- Abstract
The detection of aberrant cells by natural killer (NK) cells is controlled by the integration of signals from activating and inhibitory ligands and from cytokines such as IL-15. We identified cytokine-inducible SH2-containing protein (CIS, encoded by Cish) as a critical negative regulator of IL-15 signaling in NK cells. Cish was rapidly induced in response to IL-15, and deletion of Cish rendered NK cells hypersensitive to IL-15, as evidenced by enhanced proliferation, survival, IFN-γ production and cytotoxicity toward tumors. This was associated with increased JAK-STAT signaling in NK cells in which Cish was deleted. Correspondingly, CIS interacted with the tyrosine kinase JAK1, inhibiting its enzymatic activity and targeting JAK for proteasomal degradation. Cish(-/-) mice were resistant to melanoma, prostate and breast cancer metastasis in vivo, and this was intrinsic to NK cell activity. Our data uncover a potent intracellular checkpoint in NK cell-mediated tumor immunity and suggest possibilities for new cancer immunotherapies directed at blocking CIS function.
- Published
- 2016
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- View/download PDF
27. Novel non-canonical role of STAT1 in Natural Killer cell cytotoxicity.
- Author
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Putz EM, Majoros A, Gotthardt D, Prchal-Murphy M, Zebedin-Brandl EM, Fux DA, Schlattl A, Schreiber RD, Carotta S, Müller M, Gerner C, Decker T, and Sexl V
- Abstract
STAT1 is an important regulator of NK cell maturation and cytotoxicity. Although the consequences of Stat1 -deficiency have been described in detail the underlying molecular functions of STAT1 in NK cells are only partially understood. Here, we describe a novel non-canonical role of STAT1 that was unmasked in NK cells expressing a Stat1-Y701F mutant. This mutation prevents JAK-dependent phosphorylation, subsequent nuclear translocation and cytokine-induced transcriptional activity as verified by RNA-seq analysis. As expected Stat1-Y701F mice displayed impaired NK cell maturation comparable to Stat1
-/- animals. In contrast Stat1-Y701F NK cells exerted a significantly enhanced cytotoxicity in vitro and in vivo compared to Stat1-/- NK cells in the absence of detectable transcriptional activity. We thus investigated the STAT1 interactome using primary NK cells derived from Stat1ind mice that inducibly express a FLAG-tagged STAT1. Mass spectrometry revealed that STAT1 directly binds proteins involved in cell junction formation and proteins associated to membrane or membrane-bound vesicles. In line, immunofluorescence studies uncovered the recruitment of STAT1 to the target-cell interphase during NK cell killing. This led us to propose a novel function for STAT1 at the immunological synapse in NK cells regulating tumor surveillance and cytotoxicity.- Published
- 2016
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- View/download PDF
28. Targeting NK Cells for Anticancer Immunotherapy: Clinical and Preclinical Approaches.
- Author
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Carotta S
- Abstract
The recent success of checkpoint blockade has highlighted the potential of immunotherapy approaches for cancer treatment. Although the majority of approved immunotherapy drugs target T cell subsets, it is appreciated that other components of the immune system have important roles in tumor immune surveillance as well and thus represent promising additional targets for immunotherapy. Natural killer (NK) cells are the body's first line of defense against infected or transformed cells, as they kill target cells in an antigen-independent manner. Although several studies have clearly demonstrated the active role of NK cells in cancer immune surveillance, only few clinically approved therapies currently exist that harness their potential. Our increased understanding of NK cell biology over the past few years has renewed the interest in NK cell-based anticancer therapies, which has lead to a steady increase of NK cell-based clinical and preclinical trials. Here, the role of NK cells in cancer immune surveillance is summarized, and several novel approaches to enhance NK cell cytotoxicity against cancer are discussed.
- Published
- 2016
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- View/download PDF
29. Complementarity and redundancy of IL-22-producing innate lymphoid cells.
- Author
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Rankin LC, Girard-Madoux MJ, Seillet C, Mielke LA, Kerdiles Y, Fenis A, Wieduwild E, Putoczki T, Mondot S, Lantz O, Demon D, Papenfuss AT, Smyth GK, Lamkanfi M, Carotta S, Renauld JC, Shi W, Carpentier S, Soos T, Arendt C, Ugolini S, Huntington ND, Belz GT, and Vivier E
- Subjects
- Animals, Citrobacter rodentium immunology, Cluster Analysis, Disease Models, Animal, Enterobacteriaceae Infections genetics, Enterobacteriaceae Infections immunology, Enterobacteriaceae Infections metabolism, Enterobacteriaceae Infections mortality, Enterobacteriaceae Infections pathology, Female, Gene Expression Profiling, Gene Expression Regulation, Homeostasis, Lymphocyte Subsets immunology, Lymphocyte Subsets metabolism, Male, Mice, Mice, Knockout, Mice, Transgenic, Myeloid Cell Leukemia Sequence 1 Protein deficiency, Myeloid Cell Leukemia Sequence 1 Protein genetics, Myeloid Cell Leukemia Sequence 1 Protein metabolism, Natural Cytotoxicity Triggering Receptor 1 metabolism, Signal Transduction, T-Box Domain Proteins deficiency, T-Box Domain Proteins genetics, T-Box Domain Proteins metabolism, Transcriptome, Interleukin-22, Immunity, Innate, Interleukins biosynthesis, Lymphocytes immunology, Lymphocytes metabolism
- Abstract
Intestinal T cells and group 3 innate lymphoid cells (ILC3 cells) control the composition of the microbiota and gut immune responses. Within the gut, ILC3 subsets coexist that either express or lack the natural cytoxicity receptor (NCR) NKp46. We identified here the transcriptional signature associated with the transcription factor T-bet-dependent differentiation of NCR(-) ILC3 cells into NCR(+) ILC3 cells. Contrary to the prevailing view, we found by conditional deletion of the key ILC3 genes Stat3, Il22, Tbx21 and Mcl1 that NCR(+) ILC3 cells were redundant for the control of mouse colonic infection with Citrobacter rodentium in the presence of T cells. However, NCR(+) ILC3 cells were essential for cecal homeostasis. Our data show that interplay between intestinal ILC3 cells and adaptive lymphocytes results in robust complementary failsafe mechanisms that ensure gut homeostasis.
- Published
- 2016
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- View/download PDF
30. The Helix-Loop-Helix Protein ID2 Governs NK Cell Fate by Tuning Their Sensitivity to Interleukin-15.
- Author
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Delconte RB, Shi W, Sathe P, Ushiki T, Seillet C, Minnich M, Kolesnik TB, Rankin LC, Mielke LA, Zhang JG, Busslinger M, Smyth MJ, Hutchinson DS, Nutt SL, Nicholson SE, Alexander WS, Corcoran LM, Vivier E, Belz GT, Carotta S, and Huntington ND
- Subjects
- Animals, Cell Lineage immunology, Cells, Cultured, Female, Flow Cytometry, Male, Mice, Mice, Mutant Strains, Receptors, Interleukin-15 immunology, Receptors, Interleukin-15 metabolism, Reverse Transcriptase Polymerase Chain Reaction, Transcription Factors immunology, Transcription Factors metabolism, Cell Differentiation immunology, Inhibitor of Differentiation Protein 2 immunology, Interleukin-15 immunology, Killer Cells, Natural cytology, Killer Cells, Natural immunology
- Abstract
The inhibitor of DNA binding 2 (Id2) is essential for natural killer (NK) cell development with its canonical role being to antagonize E-protein function and alternate lineage fate. Here we have identified a key role for Id2 in regulating interleukin-15 (IL-15) receptor signaling and homeostasis of NK cells by repressing multiple E-protein target genes including Socs3. Id2 deletion in mature NK cells was incompatible with their homeostasis due to impaired IL-15 receptor signaling and metabolic function and this could be rescued by strong IL-15 receptor stimulation or genetic ablation of Socs3. During NK cell maturation, we observed an inverse correlation between E-protein target genes and Id2. These results shift the current paradigm on the role of ID2, indicating that it is required not only to antagonize E-proteins during NK cell commitment, but constantly required to titrate E-protein activity to regulate NK cell fitness and responsiveness to IL-15., (Copyright © 2016 Elsevier Inc. All rights reserved.)
- Published
- 2016
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31. Regulation of early T-lineage gene expression and developmental progression by the progenitor cell transcription factor PU.1.
- Author
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Champhekar A, Damle SS, Freedman G, Carotta S, Nutt SL, and Rothenberg EV
- Subjects
- Animals, Cell Survival, Cells, Cultured, Mice, Inbred C57BL, Proto-Oncogene Proteins genetics, Receptors, Notch metabolism, Stem Cells, Trans-Activators genetics, Transcriptome, Cell Differentiation, Gene Expression Regulation, Developmental, Proto-Oncogene Proteins metabolism, T-Lymphocytes cytology, T-Lymphocytes metabolism, Trans-Activators metabolism
- Abstract
The ETS family transcription factor PU.1 is essential for the development of several blood lineages, including T cells, but its function in intrathymic T-cell precursors has been poorly defined. In the thymus, high PU.1 expression persists through multiple cell divisions in early stages but then falls sharply during T-cell lineage commitment. PU.1 silencing is critical for T-cell commitment, but it has remained unknown how PU.1 activities could contribute positively to T-cell development. Here we employed conditional knockout and modified antagonist PU.1 constructs to perturb PU.1 function stage-specifically in early T cells. We show that PU.1 is needed for full proliferation, restricting access to some non-T fates, and controlling the timing of T-cell developmental progression such that removal or antagonism of endogenous PU.1 allows precocious access to T-cell differentiation. Dominant-negative effects reveal that this repression by PU.1 is mediated indirectly. Genome-wide transcriptome analysis identifies novel targets of PU.1 positive and negative regulation affecting progenitor cell signaling and cell biology and indicating distinct regulatory effects on different subsets of progenitor cell transcription factors. Thus, in addition to supporting early T-cell proliferation, PU.1 regulates the timing of activation of the core T-lineage developmental program., (© 2015 Champhekar et al.; Published by Cold Spring Harbor Laboratory Press.)
- Published
- 2015
- Full Text
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32. Peripheral natural killer cell maturation depends on the transcription factor Aiolos.
- Author
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Holmes ML, Huntington ND, Thong RP, Brady J, Hayakawa Y, Andoniou CE, Fleming P, Shi W, Smyth GK, Degli-Esposti MA, Belz GT, Kallies A, Carotta S, Smyth MJ, and Nutt SL
- Subjects
- Animals, CD11b Antigen genetics, CD11b Antigen immunology, Cell Differentiation genetics, Gene Regulatory Networks immunology, Ikaros Transcription Factor, Killer Cells, Natural cytology, Mice, Mice, Knockout, Neoplasms, Experimental genetics, Neoplasms, Experimental immunology, Positive Regulatory Domain I-Binding Factor 1, T-Box Domain Proteins genetics, T-Box Domain Proteins immunology, Trans-Activators genetics, Transcription Factors genetics, Transcription Factors immunology, Tumor Necrosis Factor Receptor Superfamily, Member 7 genetics, Tumor Necrosis Factor Receptor Superfamily, Member 7 immunology, Virus Diseases genetics, Virus Diseases immunology, Cell Differentiation immunology, Immunity, Cellular, Killer Cells, Natural immunology, Trans-Activators immunology
- Abstract
Natural killer (NK) cells are an innate lymphoid cell lineage characterized by their capacity to provide rapid effector functions, including cytokine production and cytotoxicity. Here, we identify the Ikaros family member, Aiolos, as a regulator of NK-cell maturation. Aiolos expression is initiated at the point of lineage commitment and maintained throughout NK-cell ontogeny. Analysis of cell surface markers representative of distinct stages of peripheral NK-cell maturation revealed that Aiolos was required for the maturation in the spleen of CD11b(high)CD27(-) NK cells. The differentiation block was intrinsic to the NK-cell lineage and resembled that found in mice lacking either T-bet or Blimp1; however, genetic analysis revealed that Aiolos acted independently of all other known regulators of NK-cell differentiation. NK cells lacking Aiolos were strongly hyper-reactive to a variety of NK-cell-mediated tumor models, yet impaired in controlling viral infection, suggesting a regulatory function for CD27(-) NK cells in balancing these two arms of the immune response. These data place Aiolos in the emerging gene regulatory network controlling NK-cell maturation and function., (© 2014 The Authors.)
- Published
- 2014
- Full Text
- View/download PDF
33. The transcription factors IRF8 and PU.1 negatively regulate plasma cell differentiation.
- Author
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Carotta S, Willis SN, Hasbold J, Inouye M, Pang SH, Emslie D, Light A, Chopin M, Shi W, Wang H, Morse HC 3rd, Tarlinton DM, Corcoran LM, Hodgkin PD, and Nutt SL
- Subjects
- Animals, Cell Line, Cluster Analysis, Gene Expression Profiling, Gene Expression Regulation, Humans, Immunoglobulin Class Switching genetics, Interferon Regulatory Factors metabolism, Mice, Mice, Transgenic, Plasma Cells immunology, Protein Binding, Proto-Oncogene Proteins metabolism, Trans-Activators metabolism, Cell Differentiation genetics, Interferon Regulatory Factors genetics, Plasma Cells cytology, Plasma Cells metabolism, Proto-Oncogene Proteins genetics, Trans-Activators genetics
- Abstract
Activated B cells undergo immunoglobulin class-switch recombination (CSR) and differentiate into antibody-secreting plasma cells. The distinct transcriptomes of B cells and plasma cells are maintained by the antagonistic influences of two groups of transcription factors: those that maintain the B cell program, including BCL6 and PAX5, and plasma cell-promoting factors, such as IRF4 and BLIMP-1. We show that the complex of IRF8 and PU.1 controls the propensity of B cells to undergo CSR and plasma cell differentiation by concurrently promoting the expression of BCL6 and PAX5 and repressing AID and BLIMP-1. As the PU.1-IRF8 complex functions in a reciprocal manner to IRF4, we propose that concentration-dependent competition between these factors controls B cell terminal differentiation., (© 2014 Carotta et al.)
- Published
- 2014
- Full Text
- View/download PDF
34. Nfil3 is required for the development of all innate lymphoid cell subsets.
- Author
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Seillet C, Rankin LC, Groom JR, Mielke LA, Tellier J, Chopin M, Huntington ND, Belz GT, and Carotta S
- Subjects
- Animals, Basic-Leucine Zipper Transcription Factors deficiency, Basic-Leucine Zipper Transcription Factors genetics, Cell Differentiation genetics, Cell Differentiation immunology, Citrobacter rodentium immunology, Citrobacter rodentium pathogenicity, Enterobacteriaceae Infections genetics, Enterobacteriaceae Infections immunology, Gene Expression, Immunity, Mucosal genetics, Killer Cells, Natural immunology, Lung cytology, Lung immunology, Mice, Mice, Inbred C57BL, Mice, Knockout, Peyer's Patches cytology, Peyer's Patches immunology, Transplantation Chimera genetics, Transplantation Chimera immunology, Basic-Leucine Zipper Transcription Factors immunology, Immunity, Innate genetics, Lymphocyte Subsets cytology, Lymphocyte Subsets immunology
- Abstract
Innate lymphoid cell (ILC) populations protect against infection and are essential for lymphoid tissue formation and tissue remodeling after damage. Nfil3 is implicated in the function of adaptive immune lineages and NK cell development, but it is not yet known if Nfil3 regulates other innate lymphoid lineages. Here, we identify that Nfil3 is essential for the development of Peyer's patches and ILC2 and ILC3 subsets. Loss of Nfil3 selectively reduced Peyer's patch formation and was accompanied by impaired recruitment and distribution of lymphocytes within the patches. ILC subsets exhibited high Nfil3 expression and genetic deletion of Nfil3 severely compromised the development of all subsets. Subsequently, Nfil3(-/-) mice were highly susceptible to disease when challenged with inflammatory or infectious agents. Thus, we demonstrate that Nfil3 is a key regulator of the development of ILC subsets essential for immune protection in the lung and gut., (© 2014 Seillet et al.)
- Published
- 2014
- Full Text
- View/download PDF
35. Innate immunodeficiency following genetic ablation of Mcl1 in natural killer cells.
- Author
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Sathe P, Delconte RB, Souza-Fonseca-Guimaraes F, Seillet C, Chopin M, Vandenberg CJ, Rankin LC, Mielke LA, Vikstrom I, Kolesnik TB, Nicholson SE, Vivier E, Smyth MJ, Nutt SL, Glaser SP, Strasser A, Belz GT, Carotta S, and Huntington ND
- Subjects
- Animals, Apoptosis drug effects, Apoptosis physiology, Cytokines pharmacology, Disease Models, Animal, Dose-Response Relationship, Drug, Female, Immune System Diseases physiopathology, Immunity, Innate physiology, Interleukin-15 pharmacology, Interleukin-15 physiology, Killer Cells, Natural drug effects, Lymphopenia genetics, Lymphopenia pathology, Lymphopenia physiopathology, Male, Mice, Mice, Inbred C57BL, STAT5 Transcription Factor physiology, Signal Transduction physiology, Gene Deletion, Immune System Diseases genetics, Immunity, Innate genetics, Killer Cells, Natural metabolism, Killer Cells, Natural pathology, Myeloid Cell Leukemia Sequence 1 Protein genetics, Myeloid Cell Leukemia Sequence 1 Protein metabolism
- Abstract
The cytokine IL-15 is required for natural killer (NK) cell homeostasis; however, the intrinsic mechanism governing this requirement remains unexplored. Here we identify the absolute requirement for myeloid cell leukaemia sequence-1 (Mcl1) in the sustained survival of NK cells in vivo. Mcl1 is highly expressed in NK cells and regulated by IL-15 in a dose-dependent manner via STAT5 phosphorylation and subsequent binding to the 3'-UTR of Mcl1. Specific deletion of Mcl1 in NK cells results in the absolute loss of NK cells from all tissues owing to a failure to antagonize pro-apoptotic proteins in the outer mitochondrial membrane. This NK lymphopenia results in mice succumbing to multiorgan melanoma metastases, being permissive to allogeneic transplantation and being resistant to toxic shock following polymicrobial sepsis challenge. These results clearly demonstrate a non-redundant pathway linking IL-15 to Mcl1 in the maintenance of NK cells and innate immune responses in vivo.
- Published
- 2014
- Full Text
- View/download PDF
36. Differential requirement for Nfil3 during NK cell development.
- Author
-
Seillet C, Huntington ND, Gangatirkar P, Axelsson E, Minnich M, Brady HJ, Busslinger M, Smyth MJ, Belz GT, and Carotta S
- Subjects
- Animals, Animals, Newborn, Basic-Leucine Zipper Transcription Factors genetics, Basic-Leucine Zipper Transcription Factors metabolism, Cell Differentiation genetics, Cell Lineage genetics, Cells, Cultured, Flow Cytometry, Gene Expression immunology, Killer Cells, Natural metabolism, Liver cytology, Liver immunology, Liver metabolism, Mice, Mice, Knockout, Mice, Transgenic, Reverse Transcriptase Polymerase Chain Reaction, T-Box Domain Proteins genetics, T-Box Domain Proteins immunology, T-Box Domain Proteins metabolism, TNF-Related Apoptosis-Inducing Ligand genetics, TNF-Related Apoptosis-Inducing Ligand immunology, TNF-Related Apoptosis-Inducing Ligand metabolism, Thymus Gland cytology, Thymus Gland immunology, Thymus Gland metabolism, Basic-Leucine Zipper Transcription Factors immunology, Cell Differentiation immunology, Cell Lineage immunology, Killer Cells, Natural immunology
- Abstract
NK cells can be grouped into distinct subsets that are localized to different organs and exhibit a different capacity to secrete cytokines and mediate cytotoxicity. Despite these hallmarks that reflect tissue-specific specialization in NK cells, little is known about the factors that control the development of these distinct subsets. The basic leucine zipper transcription factor Nfil3 (E4bp4) is essential for bone marrow-derived NK cell development, but it is not clear whether Nfil3 is equally important for all NK cell subsets or how it induces NK lineage commitment. In this article, we show that Nfil3 is required for the formation of Eomes-expressing NK cells, including conventional medullary and thymic NK cells, whereas TRAIL(+) Eomes(-) NK cells develop independently of Nfil3. Loss of Nfil3 during the development of bone marrow-derived NK cells resulted in reduced expression of Eomes and, conversely, restoration of Eomes expression in Nfil3(-/-) progenitors rescued NK cell development and maturation. Collectively, these findings demonstrate that Nfil3 drives the formation of mature NK cells by inducing Eomes expression and reveal the differential requirements of NK cell subsets for Nfil3.
- Published
- 2014
- Full Text
- View/download PDF
37. Transcriptional control of pre-B cell development and leukemia prevention.
- Author
-
Pang SH, Carotta S, and Nutt SL
- Subjects
- Animals, B-Lymphocytes cytology, B-Lymphocytes metabolism, Humans, Leukemia genetics, Leukemia metabolism, Precursor Cells, B-Lymphoid metabolism, Transcription Factors genetics, Transcription Factors metabolism, Gene Expression Regulation, Developmental, Leukemia prevention & control, Precursor Cells, B-Lymphoid cytology, Transcription, Genetic
- Abstract
The differentiation of early B cell progenitors is controlled by multiple transcriptional regulators and growth-factor receptors. The triad of DNA-binding proteins, E2A, EBF1, and PAX5 is critical for both the early specification and commitment of B cell progenitors, while a larger number of secondary determinants, such as members of the Ikaros, ETS, Runx, and IRF families have more direct roles in promoting stage-specific pre-B gene-expression program. Importantly, it is now apparent that mutations in many of these transcription factors are associated with the progression to acute lymphoblastic leukemia. In this review, we focus on recent studies that have shed light on the transcriptional hierarchy that controls efficient B cell commitment and differentiation as well as focus on the oncogenic consequences of the loss of many of the same factors.
- Published
- 2014
- Full Text
- View/download PDF
38. The transcription factor T-bet is essential for the development of NKp46+ innate lymphocytes via the Notch pathway.
- Author
-
Rankin LC, Groom JR, Chopin M, Herold MJ, Walker JA, Mielke LA, McKenzie AN, Carotta S, Nutt SL, and Belz GT
- Subjects
- Animals, Cell Differentiation genetics, Cell Differentiation immunology, Citrobacter rodentium immunology, Enterobacteriaceae Infections immunology, Lymphocyte Subsets cytology, Mice, Mice, Knockout, T-Box Domain Proteins genetics, Antigens, Ly metabolism, Immunity, Innate, Lymphocyte Subsets immunology, Lymphocyte Subsets metabolism, Natural Cytotoxicity Triggering Receptor 1 metabolism, Receptors, Notch metabolism, Signal Transduction, T-Box Domain Proteins metabolism
- Abstract
NKp46+ innate lymphoid cells (ILCs) serve important roles in regulating the intestinal microbiota and defense against pathogens. Whether NKp46+ ILCs arise directly from lymphoid tissue-inducer (LTi) cells or represent a separate lineage remains controversial. We report here that the transcription factor T-bet (encoded by Tbx21) was essential for the development of NKp46+ ILCs but not of LTi cells or nuocytes. Deficiency in interleukin 22 (IL-22)-producing NKp46+ ILCs resulted in greater susceptibility of Tbx21-/- mice to intestinal infection. Haploinsufficient T-bet expression resulted in lower expression of the signaling molecule Notch, and Notch signaling was necessary for the transition of LTi cells into NKp46+ ILCs. Furthermore, NKp46+ ILCs differentiated solely from the CD4- LTi population, not the CD4+ LTi population. Our results pinpoint the regulation of Notch signaling by T-bet as a distinct molecular pathway that guides the development of NKp46+ ILCs.
- Published
- 2013
- Full Text
- View/download PDF
39. Regulation of murine natural killer cell commitment.
- Author
-
Huntington ND, Nutt SL, and Carotta S
- Abstract
Natural killer (NK) cells can derive from the same precursors as B and T cells, however, to achieve lineage specificity, several transcription factors need to be activated or annulled. While a few important transcription factors have been identified for NK genesis the mechanisms of how this is achieved is far from resolved. Adding to the complexity of this, NK cells are found and potentially develop in diverse locations in vivo and it remains to be addressed if a common NK cell precursor seeds diverse niches and how transcription factors may differentially regulate NK cell commitment in distinct microenvironments. Here we will summarize some recent findings in NK cell commitment and discuss how a NK cell transcriptional network might be organized, while addressing some misconceptions and anomalies along the way.
- Published
- 2013
- Full Text
- View/download PDF
40. Development of hematopoietic stem and progenitor cells from mouse embryonic stem cells, in vitro, supported by ectopic human HOXB4 expression.
- Author
-
Pilat S, Carotta S, and Klump H
- Subjects
- Animals, Cell Differentiation, Cell Proliferation, Embryoid Bodies cytology, Embryoid Bodies metabolism, Embryonic Stem Cells metabolism, Green Fluorescent Proteins metabolism, Hematopoietic Stem Cells metabolism, Humans, Mice, Platelet Membrane Glycoprotein IIb metabolism, Retroviridae metabolism, Transduction, Genetic, Cell Culture Techniques methods, Embryonic Stem Cells cytology, Hematopoietic Stem Cells cytology, Homeodomain Proteins metabolism, Transcription Factors metabolism
- Abstract
Differentiation of pluripotent embryonic stem (ES) cells can recapitulate many aspects of hematopoiesis, in vitro, and can even generate cells capable of long-term multilineage repopulation after transplantation into recipient mice, when the homeodomain transcription factor HOXB4 is ectopically expressed. Thus, the ES-cell differentiation system is of great value for a detailed understanding of the process of blood formation. Furthermore, it is also promising for future application in hematopoietic cell and gene therapy. Since the arrival of techniques which allow the reprogramming of somatic cells back to an ES cell-like state, the generation of hematopoietic stem cells from patient-specific so-called induced pluripotent stem cells shows great promise for future therapeutic applications. In this chapter, we describe how to cultivate a certain feeder cell-independent mouse embryonic stem cell line, to manipulate these cells by retroviral gene transfer to ectopically express HOXB4, to differentiate these ES cells via embryoid body formation, and to selectively expand the arising, HOXB4-expressing hematopoietic stem and progenitor cells.
- Published
- 2013
- Full Text
- View/download PDF
41. Serum- and stromal cell-free hypoxic generation of embryonic stem cell-derived hematopoietic cells in vitro, capable of multilineage repopulation of immunocompetent mice.
- Author
-
Lesinski DA, Heinz N, Pilat-Carotta S, Rudolph C, Jacobs R, Schlegelberger B, Klump H, and Schiedlmeier B
- Subjects
- Animals, Bone Marrow metabolism, Cells, Cultured, Coculture Techniques, Culture Media, Serum-Free pharmacology, Cytokines pharmacology, Embryonic Stem Cells metabolism, Flow Cytometry, Hematopoietic Stem Cells metabolism, Mice, Mice, SCID, Pluripotent Stem Cells metabolism, Regeneration physiology, Stromal Cells metabolism, Cell Differentiation, Cell Lineage, Embryonic Stem Cells cytology, Hematopoietic Stem Cells cytology, Hypoxia metabolism, Pluripotent Stem Cells cytology, Stromal Cells cytology
- Abstract
Induced pluripotent stem cells (iPSCs) may become a promising source for the generation of patient-specific hematopoietic stem cells (HSCs) in vitro. A crucial prerequisite will be the availability of reliable protocols for the directed and efficient differentiation toward HSCs. So far, the most robust strategy for generating HSCs from pluripotent cells in vitro has been established in the mouse model involving ectopic expression of the human transcription factor HOXB4. However, most differentiation protocols include coculture on a xenogenic stroma cell line and the use of animal serum. Involvement of any of both would pose a major barrier to the translation of those protocols to human autologous iPSCs intended for clinical use. Therefore, we asked whether long-term repopulating HSCs can, in principle, be generated from embryonic stem cells without stroma cells or serum. Here, we showed that long-term multilineage engraftment could be accomplished in immunocompetent mice when HSCs were generated in serum-free medium without stroma cell support and when hypoxic conditions were used. Under those conditions, HOXB4(+) embryonic stem cell-derived hematopoietic stem and progenitor cells were immunophenotypically similar to definitive bone marrow resident E-SLAM(+) (CD150(+)CD48(-)CD45(+)CD201(+)) HSCs. Thus, our findings may ease the development of definitive, adult-type HSCs from pluripotent stem cells, entirely in vitro.
- Published
- 2012
- Full Text
- View/download PDF
42. Identification of the earliest NK-cell precursor in the mouse BM.
- Author
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Carotta S, Pang SH, Nutt SL, and Belz GT
- Subjects
- Animals, Cell Differentiation, Cell Lineage, Cells, Cultured, Inhibitor of Differentiation Protein 2 genetics, Inhibitor of Differentiation Protein 2 metabolism, Interleukin-7 Receptor alpha Subunit metabolism, Killer Cells, Natural immunology, Killer Cells, Natural metabolism, Lymphoid Progenitor Cells immunology, Lymphoid Progenitor Cells metabolism, Mice, Mice, Transgenic, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Killer Cells, Natural cytology, Lymphoid Progenitor Cells cytology
- Abstract
Natural killer (NK) cells are generated in the bone marrow (BM) from lymphoid progenitors. Although several different maturation states of committed NK cells have been described, the initial stages of NK-cell differentiation from the common lymphoid progenitor are not well understood. Here we describe the identification of the earliest committed NK-cell precursors in the BM. These precursors, termed pre-pro NK cells, lack the expression of most canonical NK cell-specific surface markers but express the transcription factor inhibitor of DNA binding 2 and high levels of the IL-7 receptor. In vitro differentiation studies demonstrate that pre-pro NK cells are committed to NK-cell lineage and appear to be upstream of the previously identified NK-cell progenitor population.
- Published
- 2011
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43. Id2 expression delineates differential checkpoints in the genetic program of CD8α+ and CD103+ dendritic cell lineages.
- Author
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Jackson JT, Hu Y, Liu R, Masson F, D'Amico A, Carotta S, Xin A, Camilleri MJ, Mount AM, Kallies A, Wu L, Smyth GK, Nutt SL, and Belz GT
- Subjects
- Animals, Cell Differentiation genetics, Cell Differentiation physiology, Cells, Cultured, Dendritic Cells metabolism, Gene Expression physiology, Genes, cdc physiology, Inhibitor of Differentiation Protein 2 physiology, Mice, Mice, Inbred BALB C, Mice, Inbred C57BL, Mice, Transgenic, Models, Biological, Antigens, CD metabolism, CD8 Antigens metabolism, Cell Lineage genetics, Dendritic Cells physiology, Inhibitor of Differentiation Protein 2 genetics, Integrin alpha Chains metabolism
- Abstract
Dendritic cells (DCs) have critical roles in the induction of the adaptive immune response. The transcription factors Id2, Batf3 and Irf-8 are required for many aspects of murine DC differentiation including development of CD8α(+) and CD103(+) DCs. How they regulate DC subset specification is not completely understood. Using an Id2-GFP reporter system, we show that Id2 is broadly expressed in all cDC subsets with the highest expression in CD103(+) and CD8α(+) lineages. Notably, CD103(+) DCs were the only DC able to constitutively cross-present cell-associated antigens in vitro. Irf-8 deficiency affected loss of development of virtually all conventional DCs (cDCs) while Batf3 deficiency resulted in the development of Sirp-α(-) DCs that had impaired survival. Exposure to GM-CSF during differentiation induced expression of CD103 in Id2-GFP(+) DCs. It did not restore cross-presenting capacity to Batf3(-/-) or CD103(-)Sirp-α(-)DCs in vitro. Thus, Irf-8 and Batf3 regulate distinct stages in DC differentiation during the development of cDCs. Genetic mapping DC subset differentiation using Id2-GFP may have broad implications in understanding the interplay of DC subsets during protective and pathological immune responses.
- Published
- 2011
- Full Text
- View/download PDF
44. A role for Blimp1 in the transcriptional network controlling natural killer cell maturation.
- Author
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Kallies A, Carotta S, Huntington ND, Bernard NJ, Tarlinton DM, Smyth MJ, and Nutt SL
- Subjects
- Animals, Base Sequence, Cell Differentiation genetics, Cell Differentiation immunology, Cell Proliferation, Cytokines biosynthesis, Cytotoxicity, Immunologic, DNA Primers genetics, Gene Expression Regulation, Developmental, Gene Regulatory Networks, Homeostasis immunology, Humans, In Vitro Techniques, Interleukin-15 immunology, Killer Cells, Natural metabolism, Mice, Mice, Inbred C57BL, Mice, Knockout, Mice, Transgenic, Positive Regulatory Domain I-Binding Factor 1, Repressor Proteins genetics, Repressor Proteins immunology, Signal Transduction immunology, T-Box Domain Proteins immunology, Transcription Factors genetics, Up-Regulation, Killer Cells, Natural cytology, Killer Cells, Natural immunology, Transcription Factors immunology
- Abstract
Natural killer (NK) cells are innate lymphocytes capable of immediate effector functions including cytokine production and cytotoxicity. Compared with B and T cells, the factors that control the peripheral maturation of NK cells are poorly understood. We show that Blimp1, a transcriptional repressor required for the differentiation of plasma cells and short-lived effector T cells, is expressed by NK cells throughout their development. Interleukin 15 (IL-15) is required for the early induction of Blimp1 in NK cells, with expression increasing in the most mature subsets of mouse and human NK cells. We show that Blimp1 is required for NK-cell maturation and homeostasis and for regulating their proliferative potential. It is also essential for high granzyme B expression, but not for most cytokine production and cytotoxicity. Surprisingly, interferon regulatory factor 4 (IRF4) and B-cell lymphoma 6 (Bcl6), 2 transcription factors crucial for the regulation of Blimp1 in B and T cells, are largely dispensable for Blimp1 expression in NK cells. T-bet deficiency, however, leads to attenuated Blimp1 expression. We have identified NK cells as the first hematopoietic cell type in which the IRF4-Blimp1-Bcl6 regulatory axis is not in operation, highlighting the distinct nature of the NK-cell gene-regulatory network.
- Published
- 2011
- Full Text
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45. The interactions of multiple cytokines control NK cell maturation.
- Author
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Brady J, Carotta S, Thong RP, Chan CJ, Hayakawa Y, Smyth MJ, and Nutt SL
- Subjects
- Animals, Cell Differentiation genetics, Cells, Cultured, Cytokines antagonists & inhibitors, Cytokines physiology, Cytotoxicity Tests, Immunologic, Homeodomain Proteins genetics, Humans, Immunophenotyping, Inflammation Mediators physiology, Killer Cells, Natural cytology, Membrane Proteins metabolism, Membrane Proteins physiology, Mice, Mice, Inbred C57BL, Mice, Knockout, Signal Transduction genetics, Signal Transduction immunology, Cell Differentiation immunology, Cytokines metabolism, Inflammation Mediators metabolism, Killer Cells, Natural immunology, Killer Cells, Natural metabolism
- Abstract
Although NK cells are well known for their cytotoxic functions, they also produce an array of immunoregulatory cytokines and chemokines. During an immune response, NK cells are exposed to complex combinations of cytokines that influence their differentiation and function. In this study, we have examined the phenotypic and functional consequences of exposing mouse NK cells to IL-4, IL-12, IL-15, IL-18, and IL-21 and found that although all factors induced signs of maturation, characterized by decreased proliferation and IFN-γ secretion, distinct combinations induced unique cytokine secretion profiles. In contrast, the immunosuppressive factors IL-10 and TGF-β had little direct effect on NK cell effector functions. Sustained IL-18 signals resulted in IL-13 and GM-CSF production, whereas IL-12 and IL-21 induced IL-10 and TNF-α. Surprisingly, with the exception of IL-21, all cytokines suppressed cytotoxic function of NK cells at the expense of endogenous cytokine production suggesting that "helper-type" NK cells were generated. The cytokine signals also profoundly altered the cell surface phenotype of the NK cells-a striking example being the downregulation of the activating receptor NKG2D by IL-4 that resulted in decreased NKG2D-dependent killing. IL-4 exposure also modulated NKG2D expression in vivo suggesting it is functionally important during immune responses. This study highlights the plasticity of NK cell differentiation and suggests that the relative abundance of cytokines at sites of inflammation will lead to diverse outcomes in terms of NK cell phenotype and interaction with the immune system.
- Published
- 2010
- Full Text
- View/download PDF
46. Mcl-1 is essential for germinal center formation and B cell memory.
- Author
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Vikstrom I, Carotta S, Lüthje K, Peperzak V, Jost PJ, Glaser S, Busslinger M, Bouillet P, Strasser A, Nutt SL, and Tarlinton DM
- Subjects
- Animals, Antibody Affinity, Cell Survival, Gene Deletion, Germinal Center cytology, Mice, Mice, Inbred C57BL, Molecular Sequence Data, Myeloid Cell Leukemia Sequence 1 Protein, Proto-Oncogene Proteins c-bcl-2 genetics, bcl-X Protein genetics, bcl-X Protein immunology, B-Lymphocytes immunology, Germinal Center immunology, Immunologic Memory, Proto-Oncogene Proteins c-bcl-2 immunology
- Abstract
Lymphocyte survival during immune responses is controlled by the relative expression of pro- and anti-apoptotic molecules, regulating the magnitude, quality, and duration of the response. We investigated the consequences of deleting genes encoding the anti-apoptotic molecules Mcl1 and Bcl2l1 (Bcl-x(L)) from B cells using an inducible system synchronized with expression of activation-induced cytidine deaminase (Aicda) after immunization. This revealed Mcl1 and not Bcl2l1 to be indispensable for the formation and persistence of germinal centers (GCs). Limiting Mcl1 expression reduced the magnitude of the GC response with an equivalent, but not greater, effect on memory B cell formation and no effect on persistence. Our results identify Mcl1 as the main anti-apoptotic regulator of activated B cell survival and suggest distinct mechanisms controlling survival of GC and memory B cells.
- Published
- 2010
- Full Text
- View/download PDF
47. Surprising new roles for PU.1 in the adaptive immune response.
- Author
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Carotta S, Wu L, and Nutt SL
- Subjects
- Animals, Hematopoiesis genetics, Hematopoiesis immunology, Humans, Immunoglobulin Class Switching genetics, Interleukin-9 metabolism, Membrane Proteins immunology, Mice, Mutation genetics, Proto-Oncogene Proteins genetics, Th1-Th2 Balance, Trans-Activators genetics, Adaptive Immunity genetics, B-Lymphocytes immunology, CD4-Positive T-Lymphocytes immunology, Dendritic Cells immunology, Proto-Oncogene Proteins immunology, Trans-Activators immunology
- Abstract
The ETS family transcription factor PU.1 is one of the best-studied regulators of hematopoiesis. While research over the past two decades has established that PU.1 is essential for many aspects of lymphoid and myeloid cell development, the more recent development of the tools that enable PU.1 function to be assessed in adult mice and in specific cell lineages has led to the discovery of some surprising new roles of this versatile factor in the adaptive immune response. Despite being required for fetal lymphopoiesis, PU.1 is dispensable for the differentiation of committed B cells. There is, however, an emerging and still uncharacterized function of PU.1 as a repressor for late B-cell differentiation. In contrast, PU.1 is required at every point for the differentiation of all dendritic cells, in part, although its regulation of the crucial receptor Flt3. Within the T-cell lineage, PU.1 is required for the earliest thymic development, although the mechanism remains unknown, while recent studies have shown a previously unknown function of PU.1 in peripheral T-cell differentiation. Here, we review insights derived from these mouse models of PU.1 deficiency, with particular emphasis on these functions of PU.1 in the lymphocyte and dendritic cell lineages., (© 2010 John Wiley & Sons A/S.)
- Published
- 2010
- Full Text
- View/download PDF
48. The transcription factor PU.1 controls dendritic cell development and Flt3 cytokine receptor expression in a dose-dependent manner.
- Author
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Carotta S, Dakic A, D'Amico A, Pang SH, Greig KT, Nutt SL, and Wu L
- Subjects
- Animals, Base Sequence, Cell Differentiation, Cells, Cultured, Dendritic Cells drug effects, Dose-Response Relationship, Drug, Flow Cytometry, Membrane Proteins drug effects, Membrane Proteins genetics, Mice, Mice, Transgenic, Models, Immunological, Molecular Sequence Data, Polymerase Chain Reaction, Promoter Regions, Genetic, Proto-Oncogene Proteins pharmacology, Trans-Activators pharmacology, Dendritic Cells immunology, Membrane Proteins metabolism, Proto-Oncogene Proteins metabolism, Trans-Activators metabolism
- Abstract
The transcription factor PU.1 plays multiple context and concentration dependent roles in lymphoid and myeloid cell development. Here we showed that PU.1 (encoded by Sfpi1) was essential for dendritic cell (DC) development in vivo and that conditional ablation of PU.1 in defined precursors, including the common DC progenitor, blocked Flt3 ligand-induced DC generation in vitro. PU.1 was also required for the parallel granulocyte-macrophage colony stimulating factor-induced DC pathway from early hematopoietic progenitors. Molecular studies demonstrated that PU.1 directly regulated Flt3 in a concentration-dependent manner, as Sfpi1(+/-) cells displayed reduced expression of Flt3 and impaired DC formation. These studies identify PU.1 as a critical regulator of both conventional and plasmacytoid DC development and provide one mechanism how altered PU.1 concentration can have profound functional consequences for hematopoietic cell development., (Copyright 2010 Elsevier Inc. All rights reserved.)
- Published
- 2010
- Full Text
- View/download PDF
49. Analysis of interleukin-21-induced Prdm1 gene regulation reveals functional cooperation of STAT3 and IRF4 transcription factors.
- Author
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Kwon H, Thierry-Mieg D, Thierry-Mieg J, Kim HP, Oh J, Tunyaplin C, Carotta S, Donovan CE, Goldman ML, Tailor P, Ozato K, Levy DE, Nutt SL, Calame K, and Leonard WJ
- Subjects
- Animals, B-Lymphocytes immunology, Base Sequence, Binding Sites, CD4-Positive T-Lymphocytes immunology, Cell Differentiation, Genome-Wide Association Study, Interferon Regulatory Factors genetics, Introns, Mice, Mice, Inbred C57BL, Mice, Knockout, Molecular Sequence Data, Positive Regulatory Domain I-Binding Factor 1, STAT3 Transcription Factor genetics, Gene Expression Regulation, Interferon Regulatory Factors metabolism, Interleukins metabolism, STAT3 Transcription Factor metabolism, Transcription Factors genetics
- Abstract
Interleukin-21 (IL-21) is a pleiotropic cytokine that induces expression of transcription factor BLIMP1 (encoded by Prdm1), which regulates plasma cell differentiation and T cell homeostasis. We identified an IL-21 response element downstream of Prdm1 that binds the transcription factors STAT3 and IRF4, which are required for optimal Prdm1 expression. Genome-wide ChIP-Seq mapping of STAT3- and IRF4-binding sites showed that most regions with IL-21-induced STAT3 binding also bound IRF4 in vivo and furthermore revealed that the noncanonical TTCnnnTAA GAS motif critical in Prdm1 was broadly used for STAT3 binding. Comparing genome-wide expression array data to binding sites revealed that most IL-21-regulated genes were associated with combined STAT3-IRF4 sites rather than pure STAT3 sites. Correspondingly, ChIP-Seq analysis of Irf4(-/-) T cells showed greatly diminished STAT3 binding after IL-21 treatment, and Irf4(-/-) mice showed impaired IL-21-induced Tfh cell differentiation in vivo. These results reveal broad cooperative gene regulation by STAT3 and IRF4., (Copyright 2009 Elsevier Inc. All rights reserved.)
- Published
- 2009
- Full Text
- View/download PDF
50. PU.1 regulates TCR expression by modulating GATA-3 activity.
- Author
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Chang HC, Han L, Jabeen R, Carotta S, Nutt SL, and Kaplan MH
- Subjects
- Animals, CD4-Positive T-Lymphocytes metabolism, Cell Differentiation immunology, Cytokines biosynthesis, Cytokines immunology, Female, GATA3 Transcription Factor genetics, GATA3 Transcription Factor immunology, Gene Knockdown Techniques, Mice, Mice, Inbred C57BL, Mice, Transgenic, Proto-Oncogene Proteins genetics, Proto-Oncogene Proteins immunology, RNA, Small Interfering immunology, RNA, Small Interfering metabolism, Receptors, Antigen, T-Cell metabolism, Th2 Cells metabolism, Trans-Activators genetics, Trans-Activators immunology, CD4-Positive T-Lymphocytes immunology, GATA3 Transcription Factor metabolism, Proto-Oncogene Proteins metabolism, Receptors, Antigen, T-Cell immunology, Th2 Cells immunology, Trans-Activators metabolism
- Abstract
The Ets transcription factor PU.1 is a master regulator for the development of multiple lineages during hematopoiesis. The expression pattern of PU.1 is dynamically regulated during early T lineage development in the thymus. We previously revealed that PU.1 delineates heterogeneity of effector Th2 populations. In this study, we further define the function of PU.1 on the Th2 phenotype using mice that specifically lack PU.1 in T cells using an lck-Cre transgene with a conditional Sfpi1 allele (Sfpi1(lck-/-)). Although deletion of PU.1 by the lck-Cre transgene does not affect T cell development, Sfpi1(lck-/-) T cells have a lower activation threshold than wild-type T cells. When TCR engagement is limiting, Sfpi1(lck-/-) T cells cultured in Th2 polarizing conditions secrete higher levels of Th2 cytokines and have greater cytokine homogeneity than wild-type cells. We show that PU.1 modulates the levels of TCR expression in CD4(+) T cells by regulating the DNA-binding activity of GATA-3 and limiting GATA-3 regulation of TCR gene expression. GATA-3-dependent regulation of TCR expression is also observed in Th1 and Th2 cells. In CD4(+) T cells, PU.1 expression segregates into subpopulations of cells that have lower levels of surface TCR, suggesting that PU.1 contributes to the heterogeneity of TCR expression. Thus, we have identified a mechanism whereby increased GATA-3 function in the absence of the antagonizing activity of PU.1 leads to increased TCR expression, a reduced activation threshold, and increased homogeneity in Th2 populations.
- Published
- 2009
- Full Text
- View/download PDF
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