Publications by authors named "Michael J Grusby"

45 Publications

ZBTB20 is required for anterior pituitary development and lactotrope specification.

Nat Commun 2016 Apr 15;7:11121. Epub 2016 Apr 15.

Department of Pathophysiology, Second Military Medical University, 800 Xiangyin Road, Shanghai 200433, China.

The anterior pituitary harbours five distinct hormone-producing cell types, and their cellular differentiation is a highly regulated and coordinated process. Here we show that ZBTB20 is essential for anterior pituitary development and lactotrope specification in mice. In anterior pituitary, ZBTB20 is highly expressed by all the mature endocrine cell types, and to some less extent by somatolactotropes, the precursors of prolactin (PRL)-producing lactotropes. Disruption of Zbtb20 leads to anterior pituitary hypoplasia, hypopituitary dwarfism and a complete loss of mature lactotropes. In ZBTB20-null mice, although lactotrope lineage commitment is normally initiated, somatolactotropes exhibit profound defects in lineage specification and expansion. Furthermore, endogenous ZBTB20 protein binds to Prl promoter, and its knockdown decreases PRL expression and secretion in a lactotrope cell line MMQ. In addition, ZBTB20 overexpression enhances the transcriptional activity of Prl promoter in vitro. In conclusion, our findings point to ZBTB20 as a critical regulator of anterior pituitary development and lactotrope specification.
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http://dx.doi.org/10.1038/ncomms11121DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4835541PMC
April 2016

IL-21 promotes CD8+ CTL activity via the transcription factor T-bet.

J Immunol 2013 Apr 11;190(8):3977-84. Epub 2013 Mar 11.

Center for Neurologic Diseases, Brigham and Women's Hospital and Harvard Medical School, Boston, MA 02115, USA.

CD8(+) T cells are fundamental for immune-mediated clearance of viral infections and contribute to immune pathology in autoimmune diseases such as type 1 diabetes. To execute these functions, CD8(+) T cells must differentiate into CTLs, a process that is precisely regulated by a variety of cytokines, costimulatory molecules, and transcription factors. IL-21 is an IL-2 family cytokine and a growth factor for multiple lymphocyte effector lineages, including cytotoxic CD8(+) T cells. Recent studies demonstrate that loss of IL-21 signaling results in reduced viral clearance in models of lymphocytic choriomeningitis virus infection, and also protection from type 1 diabetes in the NOD model. This is most likely the result of impaired CD8(+) CTL function in the absence of IL-21 signaling. Currently, the mechanisms by which IL-21 promotes CTL differentiation in CD8(+) T cells remain unclear, particularly the identity of the relevant transcription factor(s). We show that IL-21 promotes CTL function in vitro and killing of pancreatic islets in vivo via the use of transgenic mice expressing IL-21 in pancreatic β cells. We demonstrate that IL-21 induces the expression of the transcription factor T-bet in CD8(+) T cells, predominantly via STAT1, and that T-bet is required for the induction of cytolytic molecules, including perforin and granzyme B in response to IL-21. Finally, we show that IL-21-induced CTL function is T-bet dependent, as T-bet deficiency results in defective IL-21-dependent cytotoxicity in CD8(+) T cells in vitro and in vivo. Thus, IL-21 drives CD8(+) CTL differentiation via the actions of the transcription factor T-bet.
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http://dx.doi.org/10.4049/jimmunol.1201730DOI Listing
April 2013

PDLIM2 restricts Th1 and Th17 differentiation and prevents autoimmune disease.

Cell Biosci 2012 Jun 25;2(1):23. Epub 2012 Jun 25.

University of Pittsburgh Cancer Institute, Pittsburgh, PA, USA.

Unlabelled:

Background: PDLIM2 is essential for the termination of the inflammatory transcription factors NF-κB and STAT but is dispensable for the development of immune cells and immune tissues/organs. Currently, it remains unknown whether and how PDLIM2 is involved in physiologic and pathogenic processes.

Results: Here we report that naive PDLIM2 deficient CD4+ T cells were prone to differentiate into Th1 and Th17 cells. PDLIM2 deficiency, however, had no obvious effect on lineage commitment towards Th2 or Treg cells. Notably, PDLIM2 deficient mice exhibited increased susceptibility to experimental autoimmune encephalitis (EAE), a Th1 and/or Th17 cell-mediated inflammatory disease model of multiple sclerosis (MS). Mechanistic studies further indicate that PDLIM2 was required for restricting expression of Th1 and Th17 cytokines, which was in accordance with the role of PDLIM2 in the termination of NF-κB and STAT activation.

Conclusion: These findings suggest that PDLIM2 is a key modulator of T-cell-mediated immune responses that may be targeted for the therapy of human autoimmune diseases.
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http://dx.doi.org/10.1186/2045-3701-2-23DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3543335PMC
June 2012

The zinc finger protein ZBTB20 regulates transcription of fructose-1,6-bisphosphatase 1 and β cell function in mice.

Gastroenterology 2012 Jun 25;142(7):1571-1580.e6. Epub 2012 Feb 25.

Department of Pathophysiology, Second Military Medical University, Shanghai, China.

Background & Aims: Fructose-1,6-bisphosphatase (FBP)-1 is a gluconeogenic enzyme that regulates glucose metabolism and insulin secretion in β cells, but little is known about how its transcription is controlled. The zinc finger protein ZBTB20 regulates glucose homeostasis, so we investigated its effects on expression of FBP-1.

Methods: We analyzed gene expression using real-time reverse-transcription polymerase chain reaction, immunoblotting, and immunohistochemistry. We generated mice with β cell-specific disruption of Zbtb20 using Cre/LoxP technology. Expression of Zbtb20 in β cells was reduced using small interfering RNAs, and promoter occupancy and transcriptional regulation were analyzed by chromatin immunoprecipitation and reporter assays.

Results: ZBTB20 was expressed at high levels by β cells and other endocrine cells in islets of normal mice; expression levels were reduced in islets from diabetic db/db mice. Mice with β cell-specific knockout of Zbtb20 had normal development of β cells but had hyperglycemia, hypoinsulinemia, glucose intolerance, and impaired glucose-stimulated insulin secretion. Islets isolated from these mice had impaired glucose metabolism, adenosine triphosphate production, and insulin secretion after glucose stimulation in vitro, although insulin secretion returned to normal levels in the presence of KCl. ZBTB20 knockdown with small interfering RNAs impaired glucose-stimulated insulin secretion in the β cell line MIN6. Expression of Fbp1 was up-regulated in β cells with ZBTB20 knockout or knockdown; impairments to glucose-stimulated insulin secretion were restored by inhibition of FBPase activity. ZBTB20 was recruited to the Fbp1 promoter and repressed its transcription in β cells.

Conclusions: The transcription factor ZBTB20 regulates β cell function and glucose homeostasis in mice. It might be a therapeutic target for type 2 diabetes mellitus.
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http://dx.doi.org/10.1053/j.gastro.2012.02.043DOI Listing
June 2012

PDLIM2 inhibits T helper 17 cell development and granulomatous inflammation through degradation of STAT3.

Sci Signal 2011 Dec 6;4(202):ra85. Epub 2011 Dec 6.

Laboratory for Inflammatory Regulation, RIKEN Research Center for Allergy and Immunology, Yokohama, Kanagawa 230-0045, Japan.

Granuloma formation is an important host defense mechanism against intracellular bacteria; however, uncontrolled granulomatous inflammation is pathologic. T helper 17 (TH17) cells are thought to have a pathogenic role in autoimmune and inflammatory diseases, including in granulomas. Here, we report that the PDZ-LIM domain protein PDLIM2 inhibited TH17 cell development and granulomatous responses by acting as a nuclear ubiquitin E3 ligase that targeted signal transducer and activator of transcription 3 (STAT3), a transcription factor critical for the commitment of naïve CD4+ T cells to the TH17 lineage. PDLIM2 promoted the polyubiquitination and proteasomal degradation of STAT3, thereby disrupting STAT3-mediated gene activation. Deficiency in PDLIM2 resulted in the accumulation of STAT3 in the nucleus, enhanced the extent of TH17 cell differentiation, and exacerbated granuloma formation. This study delineates an essential role for PDLIM2 in inhibiting TH17 cell-mediated inflammatory responses by suppressing STAT3 signaling and provides a potential therapeutic target for the treatment of autoimmune diseases.
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http://dx.doi.org/10.1126/scisignal.2001637DOI Listing
December 2011

DNA methylation-dependent repression of PDZ-LIM domain-containing protein 2 in colon cancer and its role as a potential therapeutic target.

Cancer Res 2010 Mar 9;70(5):1766-72. Epub 2010 Feb 9.

University of Pittsburgh Cancer Institute, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA.

Constitutive activation of the nuclear factor-kappaB (NF-kappaB) transcription factor plays a key role in chronic colonic inflammation and colon tumorigenesis. However, the mechanisms by which the tightly regulated NF-kappaB pathway becomes constitutively activated during colonic pathogenesis remain obscure. Here, we report that PDLIM2, an essential terminator of NF-kappaB activation, is repressed in various human colorectal cancer cell lines, suggesting one important mechanism for the constitutive activation of NF-kappaB. Indeed, expression of exogenous PDLIM2 inhibited constitutive NF-kappaB activation in these colorectal cancer cells. Importantly, the PDLIM2 expression was sufficient to suppress in vitro anchorage-independent growth and in vivo tumor formation of these malignant cells. We have further shown that the PDLIM2 repression involves promoter methylation. Accordingly, treatment of the colorectal tumor cell lines with the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine restored PDLIM2 expression and resulted in growth arrest. These studies thus provide new mechanistic insights into colon tumorigenesis by identifying a novel tumor suppressor role for PDLIM2.
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http://dx.doi.org/10.1158/0008-5472.CAN-09-3263DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3003295PMC
March 2010

NIP45 controls the magnitude of the type 2 T helper cell response.

Proc Natl Acad Sci U S A 2010 Feb 4;107(8):3663-8. Epub 2010 Feb 4.

Department of Chemical Physiology, The Scripps Research Institute, La Jolla, CA 92037, USA.

Nuclear factor of activated T cell (NFAT) transcription factors are key regulators of gene transcription within immune cells. The NFAT-interacting protein, (NIP45), augments NFAT-driven IL-4 expression by a mechanism that relies on arginine methylation. To establish the function of NIP45 in vivo, we generated mice with a targeted deletion of the gene encoding this cofactor. NIP45-deficient T helper cells displayed profound defects in the expression of NFAT-regulated cytokine genes, including IL-4. Whereas NIP45 deficiency does not interfere with T helper cell NFAT activation or lineage-specific transcription-factor expression, NIP45 acts as an enhancer for the assembly of protein arginine methyltransferase 1 and the protein arginine methyltransferase 1-linked histone 4 arginine 3 methylation with the IL-4 promoter. Our study reveals an essential role for NIP45 in promoting robust cytokine expression in vivo, which is required for the efficient handling of parasites. We propose that NIP45 acts as a molecular rheostat serving to amplify the type-2 immune response.
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http://dx.doi.org/10.1073/pnas.0914700107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2840445PMC
February 2010

c-Maf activates the promoter and enhancer of the IL-21 gene, and TGF-beta inhibits c-Maf-induced IL-21 production in CD4+ T cells.

J Leukoc Biol 2010 Apr 30;87(4):703-12. Epub 2009 Dec 30.

Department of Molecular Genetics, Graduate School of Medicine, Chiba University, Chiba 260-8670, Japan.

Previous studies have shown that IL-6 potently induces IL-21 production in CD4(+) T cells, whereas TGF-beta inhibits IL-6-induced IL-21 production in CD4(+) T cells. In this study, we addressed the mechanisms underlying the transcriptional regulation of IL-21 production in CD4(+) T cells. We found that IL-6 induced c-Maf expression in CD4(+) T cells and that the enforced expression of c-Maf induced IL-21 production in CD4(+) T cells without IL-6, IL-4/STAT6 signaling, or an autocrine effect of IL-21. Moreover, we found that c-Maf directly bound to and activated IL-21P and the CNS-2 enhancer through MARE sites. On the other hand, we also found that although TGF-beta up-regulated IL-6-induced c-Maf expression in CD4(+) T cells, TGF-beta inhibited c-Maf-induced IL-21 production in CD4(+) T cells. Finally, we found that Foxp3 bound to IL-21P and the CNS-2 enhancer and inhibited c-Maf-induced IL-21 production modestly but significantly in CD4(+) T cells. Taken together, these results suggest that c-Maf induces IL-21 production directly in CD4(+) T cells by activating IL-21P and the CNS-2 enhancer and that TGF-beta suppresses c-Maf-induced IL-21 production in CD4(+) T cells.
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http://dx.doi.org/10.1189/jlb.0909639DOI Listing
April 2010

Zinc finger protein Zbtb20 is essential for postnatal survival and glucose homeostasis.

Mol Cell Biol 2009 May 9;29(10):2804-15. Epub 2009 Mar 9.

Department of Immunology and Infectious Diseases, Harvard School of Public Health, 651 Huntington Ave., Boston, MA 02115, USA.

Zbtb20 is a member of the POK family of proteins, which function primarily as transcriptional repressors via interactions mediated by their conserved C(2)H(2) Krüppel type zinc finger and BTB/POZ domains. To define the function of Zbtb20 in vivo, we generated knockout mice by homologous recombination. Zbtb20 null mice display a stark phenotype characterized by postnatal growth retardation, metabolic dysfunction, and lethality. Zbtb20 knockout mice displayed abnormal glucose homeostasis, hormonal responses, and depletion of energy stores, consistent with an energetic deficit. Additionally, increased serum bilirubin and alanine aminotransferase levels were suggestive of liver dysfunction. To identify potential liver-specific Zbtb20 target genes, we performed transcript profiling studies on liver tissue from Zbtb20 knockout mice and wild-type littermate controls. These studies identified sets of genes involved in growth, metabolism, and detoxification that were differentially regulated in Zbtb20 knockout liver. Transgenic mice expressing Zbtb20 in the liver were generated and crossed onto the Zbtb20 knockout background, which resulted in no significant normalization of growth or glucose metabolism but a significant increase in life span compared to controls. These data indicate that the phenotype of Zbtb20 knockout mice results from liver-dependent and -independent defects, suggesting that Zbtb20 plays nonredundant roles in multiple organ systems.
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http://dx.doi.org/10.1128/MCB.01667-08DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2682054PMC
May 2009

Identification of the small protein rich in arginine and glycine (SRAG): a newly identified nucleolar protein that can regulate cell proliferation.

J Biol Chem 2009 May 2;284(18):12504-11. Epub 2009 Mar 2.

Department of Immunology and Infectious Disease, Harvard School of Public Health, Boston, Massachusetts 02115, USA.

The characterization of new proteins will aid in our explanation of normal biology and disease. Toward that goal, we describe the initial characterization of the small protein rich in arginine and glycine (SRAG). Human and mouse SRAG are 248/249-amino acid arginine- and glycine-rich proteins that are widely expressed in tissues and cell lines. Two SRAG isoforms, SRAG-5 and SRAG-3, which are truncations of full-length SRAG, were also identified. Although all SRAG proteins reside in the nucleus, they were also found within the nucleolus. Localization within the nucleolus was regulated by the N terminus of the protein. Our initial studies indicated that SRAG can interact with RNA. Full-length SRAG protein levels were highest in resting cells and were reduced in proliferating cells. The reduction in SRAG protein that occurs in proliferating cells was mapped with inhibitors to the G(2)/M phase of the cell cycle. As expected, the overexpression of SRAG reduced the percentage of cells in the G(2)/M phase and increased cell death. In sum, we have identified a new and intriguing member of the nucleolar proteome.
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http://dx.doi.org/10.1074/jbc.M809436200DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2673316PMC
May 2009

Interleukin-21 is required for the development of type 1 diabetes in NOD mice.

Diabetes 2009 May 10;58(5):1144-55. Epub 2009 Feb 10.

Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, Massachusetts, USA.

Objective: Interleukin (IL)-21 is a type 1 cytokine that has been implicated in the pathogenesis of type 1 diabetes via the unique biology of the nonobese diabetic (NOD) mouse strain. The aim of this study was to investigate a causal role for IL-21 in type 1 diabetes.

Research Design And Methods: We generated IL-21R-deficient NOD mice and C57Bl/6 mice expressing IL-21 in pancreatic beta-cells, allowing the determination of the role of insufficient and excessive IL-21 signaling in type 1 diabetes.

Results: Deficiency in IL-21R expression renders NOD mice resistant to insulitis, production of insulin autoantibodies, and onset of type 1 diabetes. The lymphoid compartment in IL-21R-/- NOD is normal and does not contain an increased regulatory T-cell fraction or diminished effector cytokine responses. However, we observed a clear defect in autoreactive effector T-cells in IL-21R-/- NOD by transfer experiments. Conversely, overexpression of IL-21 in pancreatic beta-cells induced inflammatory cytokine and chemokines, including IL-17A, IL17F, IFN-gamma, monocyte chemoattractant protein (MCP)-1, MCP-2, and interferon-inducible protein-10 in the pancreas. The ensuing leukocytic infiltration in the islets resulted in destruction of beta-cells and spontaneous type 1 diabetes in the normally diabetes-resistant C57Bl/6 and NOD x C57Bl/6 backgrounds.

Conclusions: This work provides demonstration of the essential prodiabetogenic activities of IL-21 on diverse genetic backgrounds (NOD and C57BL/6) and indicates that IL-21 blockade could be a promising strategy for interventions in human type 1 diabetes.
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http://dx.doi.org/10.2337/db08-0882DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2671036PMC
May 2009

PDLIM2 suppresses human T-cell leukemia virus type I Tax-mediated tumorigenesis by targeting Tax into the nuclear matrix for proteasomal degradation.

Blood 2009 Apr 8;113(18):4370-80. Epub 2009 Jan 8.

University of Pittsburgh Cancer Institute, University of Pittsburgh Medical Center, PA, USA.

The mechanisms by which the human T-cell leukemia virus type I (HTLV-I) Tax oncoprotein deregulates cellular signaling for oncogenesis have been extensively studied, but how Tax itself is regulated remains largely unknown. Here we report that Tax was negatively regulated by PDLIM2, which promoted Tax K48-linked polyubiquitination. In addition, PDLIM2 recruited Tax from its functional sites into the nuclear matrix where the polyubiquitinated Tax was degraded by the proteasome. Consistently, PDLIM2 suppressed Tax-mediated signaling activation, cell transformation, and oncogenesis both in vitro and in animal. Notably, PDLIM2 expression was down-regulated in HTLV-I-transformed T cells, and PDLIM2 reconstitution reversed the tumorigenicity of the malignant cells. These studies indicate that the counterbalance between HTLV-I/Tax and PDLIM2 may determine the outcome of HTLV-I infection. These studies also suggest a potential therapeutic strategy for cancers and other diseases associated with HTLV-I infection and/or PDLIM2 deregulation.
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http://dx.doi.org/10.1182/blood-2008-10-185660DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2676091PMC
April 2009

IL-21R is essential for epicutaneous sensitization and allergic skin inflammation in humans and mice.

J Clin Invest 2009 Jan 15;119(1):47-60. Epub 2008 Dec 15.

Division of Immunology, Children's Hospital and Harvard Medical SChool, Boston, MA 02115, USA.

Atopic dermatitis (AD) is a common allergic inflammatory skin disease caused by a combination of intense pruritus, scratching, and epicutaneous (e.c.) sensitization with allergens. To explore the roles of IL-21 and IL-21 receptor (IL-21R) in AD, we examined skin lesions from patients with AD and used a mouse model of allergic skin inflammation. IL-21 and IL-21R expression was upregulated in acute skin lesions of AD patients and in mouse skin subjected to tape stripping, a surrogate for scratching. The importance of this finding was highlighted by the fact that both Il21r-/- mice and WT mice treated with soluble IL-21R-IgG2aFc fusion protein failed to develop skin inflammation after e.c. sensitization of tape-stripped skin. Adoptively transferred OVA-specific WT CD4+ T cells accumulated poorly in draining LNs (DLNs) of e.c. sensitized Il21r-/- mice. This was likely caused by both DC-intrinsic and nonintrinsic effects, because trafficking of skin DCs to DLNs was defective in Il21r-/- mice and, to a lesser extent, in WT mice reconstituted with Il21r-/- BM. More insight into this defect was provided by the observation that skin DCs from tape-stripped WT mice, but not Il21r-/- mice, upregulated CCR7 and migrated toward CCR7 ligands. Treatment of epidermal and dermal cells with IL-21 activated MMP2, which has been implicated in trafficking of skin DCs. These results suggest an important role for IL-21R in the mobilization of skin DCs to DLNs and the subsequent allergic response to e.c. introduced antigen.
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http://dx.doi.org/10.1172/JCI32310DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2613448PMC
January 2009

Zinc finger protein ZBTB20 is a key repressor of alpha-fetoprotein gene transcription in liver.

Proc Natl Acad Sci U S A 2008 Aug 31;105(31):10859-64. Epub 2008 Jul 31.

Department of Pathophysiology, Second Military Medical University, 800 Xiangyin Road, Shanghai 200433, China.

The alpha-fetoprotein (AFP) gene is highly activated in fetal liver but is dramatically repressed shortly after birth. The mechanisms that underlie AFP transcriptional repression in postpartum liver are not well understood. AFP enhancer, repressor region, and promoter are implicated to be involved in AFP postnatal repression, but the major transcriptional repressor remains undefined. We previously identified a zinc finger protein gene ZBTB20. To determine its physiological functions in vivo, we have generated hepatocyte-specific ZBTB20 knockout mice by the Cre/loxP approach and demonstrated here that ZBTB20 ablation in liver led to dramatic derepression of the AFP gene in entire liver throughout adult life, although the hepatocytes were normally under nonproliferating status. Furthermore, we found that ZBTB20 was a transcriptional repressor capable of specifically inhibiting AFP promoter-driven transcriptional activity. Liver chromatin immunoprecipitation and mobility shift assays showed that ZBTB20 bound to AFP promoter directly. ZBTB20 was developmentally activated in liver after birth and inversely correlated with its AFP gene expression, suggesting that activated ZBTB20 expression in liver mediated AFP gene repression. Our data point to ZBTB20 as a key regulator governing AFP gene transcription and postulate a new model for the postnatal gene repression of AFP in liver.
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http://dx.doi.org/10.1073/pnas.0800647105DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2504784PMC
August 2008

Development and characterization of IL-21-producing CD4+ T cells.

J Exp Med 2008 Jun 12;205(6):1369-79. Epub 2008 May 12.

Department of Molecular Genetics, Graduate School of Medicine, Chiba University, Chiba, 260-8670 Japan.

It has recently been shown that interleukin (IL)-21 is produced by Th17 cells, functions as an autocrine growth factor for Th17 cells, and plays critical roles in autoimmune diseases. In this study, we investigated the differentiation and characteristics of IL-21-producing CD4(+) T cells by intracellular staining. Unexpectedly, we found that under Th17-polarizing conditions, the majority of IL-21-producing CD4(+) T cells did not produce IL-17A and -17F. We also found that IL-6 and -21 potently induced the development of IL-21-producing CD4(+) T cells without the induction of IL-4, IFN-gamma, IL-17A, or IL-17F production. On the other hand, TGF-beta inhibited IL-6- and IL-21-induced development of IL-21-producing CD4(+) T cells. IL-2 enhanced the development of IL-21-producing CD4(+) T cells under Th17-polarizing conditions. Finally, IL-21-producing CD4(+) T cells exhibited a stable phenotype of IL-21 production in the presence of IL-6, but retained the potential to produce IL-4 under Th2-polarizing conditions and IL-17A under Th17-polarizing conditions. These results suggest that IL-21-producing CD4(+) T cells exhibit distinct characteristics from Th17 cells and develop preferentially in an IL-6-rich environment devoid of TGF-beta, and that IL-21 functions as an autocrine growth factor for IL-21-producing CD4(+) T cells.
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http://dx.doi.org/10.1084/jem.20072057DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2413034PMC
June 2008

IL-13 receptor alpha2 selectively inhibits IL-13-induced responses in the murine lung.

J Immunol 2008 Jan;180(1):522-9

Department of Medicine, Division of Allergy and Clinical Immunology, Johns Hopkins University School of Medicine, Baltimore, MD 21224, USA.

IL-13 is a critical cytokine at sites of Th2 inflammation. In these locations it mediates its effects via a receptor complex, which contains IL-4Ralpha and IL-13Ralpha1. A third, high-affinity IL-13 receptor, IL-13Ralpha2, also exists. Although it was initially felt to be a decoy receptor, this has not been formally demonstrated and the role(s) of this receptor has recently become controversial. To define the role(s) of IL-13Ralpha2 in IL-13-induced pulmonary inflammation and remodeling, we compared the effects of lung-targeted transgenic IL-13 in mice with wild-type and null IL-13Ralpha2 loci. We also investigated the effect of IL-13Ralpha2 deficiency on the OVA-induced inflammatory response. In this study, we show that in the absence of IL-13Ralpha2, IL-13-induced pulmonary inflammation, mucus metaplasia, subepithelial fibrosis, and airway remodeling are significantly augmented. These changes were accompanied by increased expression and production of chemokines, proteases, mucin genes, and TGF-beta1. Similarly, an enhanced inflammatory response was observed in an OVA-induced phenotype. In contrast, disruption of IL-13Ralpha2 had no effect on the tissue effects of lung-targeted transgenic IL-4. Thus, IL-13Ralpha2 is a selective and powerful inhibitor of IL-13-induced inflammatory, remodeling, and physiologic responses in the murine lung.
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http://dx.doi.org/10.4049/jimmunol.180.1.522DOI Listing
January 2008

Contribution of IL-12R mediated feedback loop to Th1 cell differentiation.

FEBS Lett 2007 Nov 12;581(27):5199-206. Epub 2007 Oct 12.

Institute of Molecular Biology, University of Zurich, Winterthurerstrasse, 190, 8057 Zurich, Switzerland.

T helper 1 (Th1) cell fate is induced by overlapping signaling pathways, whose kinetic principles and regulatory motifs are largely unknown. We identified a simple positive feedback loop in the STAT4 signaling pathway, whereby activation by IL-12 leads to the increased expression in IL-12 receptor. A computational analysis shows that this feedback loop synergizes with the one mediated by the IFN-gamma secreted by differentiating cells, when the induction of Th1 cell fate is weak. Positive feedback loops are often utilized to enhance phenotypic differentiation. This effect was confirmed by experiments showing that stochastic fluctuations in the expression of IL-12 receptor gene were amplified, leading to two discrete levels of expression in a cell population.
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http://dx.doi.org/10.1016/j.febslet.2007.10.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2757731PMC
November 2007

Functional role for I kappa BNS in T cell cytokine regulation as revealed by targeted gene disruption.

J Immunol 2007 Aug;179(3):1681-92

Laboratory of Immunobiology, Department of Medical Oncology, Dana Farber Cancer Institute, Harvard Medical School, Boston, MA 02115, USA.

Triggering of the TCR by cognate peptide/MHC ligands induces expression of I kappa BNS, a member of the I kappa B family of NF-kappaB inhibitors whose expression is associated with apoptosis of immature thymocytes. To understand the role of I kappa BNS in TCR triggering, we created a targeted disruption of the I kappa BNS gene. Surprisingly, mice lacking I kappa BNS show normal thymic progression but both thymocytes and T cells manifest reduced TCR-stimulated proliferation. Moreover, I kappa BNS knockout thymocytes and T cells produce significantly less IL-2 and IFN-gamma than wild-type cells. Transfection analysis demonstrates that I kappa BNS and c-Rel individually increase IL-2 promoter activity. The effect of I kappa BNS on the IL-2 promoter, unlike c-Rel, is dependent on the NF-kappaB rather than the CD28RE site; mutation of the NF-kappaB site extinguishes the induction of transcription by I kappa BNS in transfectants and prevents association of I kappa BNS with IL-2 promoter DNA. Microarray analyses confirm the reduction in IL-2 production and some IFN-gamma-linked transcripts in I kappa BNS knockout T cells. Collectively, our findings demonstrate that I kappa BNS regulates production of IL-2 and other cytokines induced via "strong" TCR ligation.
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http://dx.doi.org/10.4049/jimmunol.179.3.1681DOI Listing
August 2007

PDLIM2-mediated termination of transcription factor NF-kappaB activation by intranuclear sequestration and degradation of the p65 subunit.

Nat Immunol 2007 Jun 29;8(6):584-91. Epub 2007 Apr 29.

Laboratory for Host Defense, RIKEN Research Center for Allergy and Immunology, Yokohama, Kanagawa 230-0045, Japan.

Activation of transcription factor NF-kappaB in the innate immune system is tightly regulated to prevent excessive inflammatory responses. How NF-kappaB activation is terminated, however, is not fully understood. Here we report that PDLIM2 negatively regulated NF-kappaB activity, acting as a nuclear ubiquitin E3 ligase targeting the p65 subunit of NF-kappaB. PDLIM2 bound to p65 and promoted p65 polyubiquitination. In addition, PDLIM2 targeted p65 to discrete intranuclear compartments where polyubiquitinated p65 was degraded by the proteasome. PDLIM2 deficiency resulted in larger amounts of nuclear p65, defective p65 ubiquitination and augmented production of proinflammatory cytokines in response to innate stimuli. Our findings delineate a pathway by which PDLIM2 terminates NF-kappaB activation through intranuclear sequestration and subsequent degradation.
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http://dx.doi.org/10.1038/ni1464DOI Listing
June 2007

Regulation of signal transducer and activator of transcription signaling by the tyrosine phosphatase PTP-BL.

Immunity 2007 Feb;26(2):163-76

Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, MA 02115, USA.

Signal Transducer and Activator of Transcription (STAT) proteins are a family of latent cytoplasmic transcription factors that are activated by tyrosine phosphorylation after cytokine stimulation. One mechanism by which STAT signaling is regulated is by dephosphorylation through the action of protein tyrosine phosphatases (PTP). We have identified PTP-Basophil like (PTP-BL) as a STAT PTP. PTP-BL dephosphorylates STAT proteins in vitro and in vivo, resulting in attenuation of STAT-mediated gene activation. In CD4(+) T cells, PTP-BL deficiency leads to increased and prolonged activation of STAT4 and STAT6, and consequently enhanced T helper 1 (Th1) and Th2 cell differentiation. Taken together, our findings demonstrate that PTP-BL is a physiologically important negative regulator of the STAT signaling pathway.
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http://dx.doi.org/10.1016/j.immuni.2007.01.010DOI Listing
February 2007

Osteopontin induces ubiquitin-dependent degradation of STAT1 in RAW264.7 murine macrophages.

J Immunol 2007 Feb;178(3):1870-81

Department of Surgery, Duke University Medical Center, Durham, NC 27710, USA.

In systemic inflammation induced by endotoxin (LPS), the macrophage produces the majority of the circulating NO metabolites. However, while the molecular pathways which up-regulate iNOS expression have been extensively studied in the macrophage, little is known of the parallel counterregulatory pathways which repress or inhibit macrophage iNOS expression. Using both in vivo and in vitro murine models of endotoxin (LPS) stimulation, we have previously demonstrated that NO feedback inhibits its own synthesis by increasing transcription of osteopontin (OPN), a potent transrepressor of inducible NO synthase expression. In this current study, using a system of LPS-treated RAW264.7 macrophages, we go on to demonstrate that OPN increases STAT1 ubiquitination and subsequent 26s proteasome-mediated degradation to inhibit STAT1 dependent iNOS promoter activity, transcription, and protein expression. In addition, we identify STAT-interacting LIM protein as the critical STAT ubiquitin E3 ligase critical for STAT1 degradation in this setting. OPN has not been linked previously to STAT1 degradation. This regulation of STAT1 degradation underlies OPN's effect as an inhibitor of iNOS gene transcription. These are novel findings and define OPN as a unique and as yet, poorly characterized, transactivator of STAT1 degradation by the ubiquitin-proteasome system.
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http://dx.doi.org/10.4049/jimmunol.178.3.1870DOI Listing
February 2007

IL-13 is pivotal in the fibro-obliterative process of bronchiolitis obliterans syndrome.

J Immunol 2007 Jan;178(1):511-9

Department of Medicine, Division of Pulmonary and Critical Care Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA 90095, USA.

Acute allograft rejection is considered to be a predominately type 1 immune mediated response to the donor alloantigen. However, the type 2 immune mediated response has been implicated in multiple fibroproliferative diseases. Based on the fibro-obliterative lesion found during bronchiolitis obliterans syndrome (BOS), we hypothesized that the type 2 immune mediated response is involved in chronic lung allograft rejection. Specifically, whereas acute rejection is, in part, a type 1 immune response, chronic rejection is, in part, a type 2 immune response. We found the type 2 cytokine, IL-13, to be elevated and biologically active in human bronchoalveolar lavage fluid during BOS. Translational studies using a murine model of BOS demonstrated increased expression of IL-13 and its receptors that paralleled fibro-obliteration. In addition, in vivo neutralization of IL-13 reduced airway allograft matrix deposition and murine BOS, by a mechanism that was independent of IL-4. Furthermore, using IL-13Ralpha2(-/-) mice, we found increased fibro-obliteration. Moreover, anti-IL-13 therapy in combination with cyclosporin A had profound effects on reducing murine BOS. This supports the notion that IL-13 biological axis plays an important role during the pathogenesis of BOS independent of the IL-4 biological axis.
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http://dx.doi.org/10.4049/jimmunol.178.1.511DOI Listing
January 2007

IL-21 inhibits IFN-gamma production in developing Th1 cells through the repression of Eomesodermin expression.

J Immunol 2006 Sep;177(6):3721-7

Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, MA 02115, USA.

Exposure of naive Th cell precursors (Thp) to IL-21 inhibits IFN-gamma production from developing Th1 cells. The inhibition of IFN-gamma seen in IL-21-treated Thp cells is specific as the expression of other Th1 cytokines is unaffected. Recently, it has been reported that Eomesodermin (Eomes), a member of the T-box gene family, is expressed in developing CD8+ T cells and plays an important role in regulating IFN-gamma production and cytolytic effector function. In this study, we show that Eomes mRNA and protein are also expressed in developing Th1 cells, and exposure of naive Thp cells to IL-21 results in a decrease in Eomes expression. Moreover, the repression of Eomes expression by IL-21 is not due to an indirect effect of IL-21 on the expression of IFN-gamma or STAT4 and is independent of STAT1 and T-bet expression. Finally, we show that ectopic expression of Eomes prevents the inhibition of IFN-gamma production from IL-21-treated Thp cells. Taken together, these results demonstrate that Eomes plays a role in regulating IFN-gamma production in CD4+ T cells and IL-21 inhibits IFN-gamma production in developing Th1 cells through the repression of Eomes expression.
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http://dx.doi.org/10.4049/jimmunol.177.6.3721DOI Listing
September 2006

The IL-21 receptor augments Th2 effector function and alternative macrophage activation.

J Clin Invest 2006 Jul 15;116(7):2044-55. Epub 2006 Jun 15.

Immunopathogenesis Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases (NIAID), National Institutes of Health, Bethesda, Maryland 20892, USA.

The IL-21 receptor (IL-21R) shows significant homology with the IL-4R, and CD4+ Th2 cells are an important source of IL-21. Here we examined whether the IL-21R regulates the development of Th2 responses in vivo. To do this, we infected IL-21R-/- mice with the Th2-inducing pathogens Schistosoma mansoni and Nippostrongylus brasiliensis and examined the influence of IL-21R deficiency on the development of Th2-dependent pathology. We showed that granulomatous inflammation and liver fibrosis were significantly reduced in S. mansoni-infected IL-21R-/- mice and in IL-21R+/+ mice treated with soluble IL-21R-Fc (sIL-21R-Fc). The impaired granulomatous response was also associated with a marked reduction in Th2 cytokine expression and function, as evidenced by the attenuated IL-4, IL-13, AMCase, Ym1, and FIZZ1 (also referred to as RELMalpha) responses in the tissues. A similarly impaired Th2 response was observed following N. brasiliensis infection. In vitro, IL-21 significantly augmented IL-4Ralpha and IL-13Ralpha1 expression in macrophages, resulting in increased FIZZ1 mRNA and arginase-1 activity following stimulation with IL-4 and IL-13. As such, these data identify the IL-21R as an important amplifier of alternative macrophage activation. Collectively, these results illustrate an essential function for the IL-21R in the development of pathogen-induced Th2 responses, which may have relevance in therapies for both inflammatory and chronic fibrotic diseases.
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http://dx.doi.org/10.1172/JCI27727DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1479424PMC
July 2006

Induction of tolerance in arthritogenic B cells with receptors of differing affinity for self-antigen.

Proc Natl Acad Sci U S A 2006 Mar 27;103(10):3734-9. Epub 2006 Feb 27.

Section on Immunology and Immunogenetics, Joslin Diabetes Center, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02215, USA.

Multiple mechanisms of tolerance induction limit autoimmunity, but their relative contribution for lymphocytes recognizing self-antigens of differing availability is incompletely understood. The mechanisms applied to arthritogenic B cells expressing antigen-specific B cell receptors (BCRs) with different affinities for glucose-6-phosphate isomerase (GPI) were examined in the corresponding Ig gene knock-in mice. This ubiquitously expressed and blood-borne enzyme is the target autoantigen in the K/BxN model of inflammatory arthritis and perhaps in some humans with arthritis. Negative selection of B cells expressing high-affinity anti-GPI specificities, whose surface receptors were occupied by GPI, operated mainly at the transitional B cell stages in the spleen, preventing their final differentiation and entry into follicular areas. Receptor editing contributed to the purging of cells displaying anti-GPI BCRs, and significant numbers of autoreactive cells escaped through expression of an additional Ig light (L) chain, accumulating gradually in lymphoid organs. In contrast, low-affinity anti-GPI B cells, whose surface receptors did not carry GPI, matured normally. The "escaped" dual-L-chain cells and the "ignored" low-affinity cells are the likely precursors of cells that produce pathogenic autoantibodies once T cell help is provided. These studies portray, in a single system, the range of tolerance mechanisms applied to potentially pathogenic B cells, and serve as a base for dissecting where T cell help intervenes and where therapeutic agents impinge.
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http://dx.doi.org/10.1073/pnas.0600214103DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1450147PMC
March 2006

SLIM is a nuclear ubiquitin E3 ligase that negatively regulates STAT signaling.

Immunity 2005 Jun;22(6):729-36

Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, Massachusetts 02115, USA.

STAT proteins are a family of latent cytoplasmic transcription factors that are activated by tyrosine phosphorylation in response to a variety of cytokines, growth factors, and hormones. Once activated, STAT proteins translocate into the nucleus and help coordinate gene transcription. One striking feature of STAT signaling is its rapid and transient activation and deactivation cycle, although the molecular mechanisms responsible for this remain poorly understood. Here, we report on a nuclear protein that contains both PDZ and LIM domains and that interacts with activated STAT4 molecules. We show that SLIM is an ubiquitin E3 ligase that acts on STAT proteins to cause their proteosome-mediated degradation and enhance their dephosphorylation. Overexpression of SLIM leads to impaired STAT1 and STAT4 activity due to reduced STAT protein levels, while SLIM-deficiency results in increased STAT expression and thus enhanced IFNgamma production by Th1 cells. These studies suggest that SLIM is a novel ubiquitin E3 ligase whose targets include STAT proteins.
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http://dx.doi.org/10.1016/j.immuni.2005.04.008DOI Listing
June 2005

NFATc2 and T-bet contribute to T-helper-cell-subset-specific regulation of IL-21 expression.

Proc Natl Acad Sci U S A 2005 Feb 31;102(6):2016-21. Epub 2005 Jan 31.

Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, MA 02115, USA.

T helper (Th) 2 cells selectively express IL-21 in addition to the classic Th2 cytokines IL-4, IL-5, and IL-13. In contrast to these clustered Th2 cell cytokine genes, the IL-21 gene resides on a different chromosome and is not coordinately regulated by the same locus control region that directs the expression of other Th2 cytokines. We demonstrate that the proximal promoter of IL-21 controls its Th-cell-subset-specific expression through the action of NFATc2 and T-bet. Whereas NFATc2 directly binds to and activates transcription of the IL-21 promoter in Th2 cells, T-bet represses IL-21 transcription by inhibiting the binding of NFATc2 to the promoter in Th1 cells. These data suggest that there are multiple mechanisms by which Th-cell-subset-specific cytokine genes are regulated.
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http://dx.doi.org/10.1073/pnas.0409512102DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC548571PMC
February 2005

Biology of IL-21 and the IL-21 receptor.

Immunol Rev 2004 Dec;202:84-95

Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, MA 02115, USA.

Interleukin-21 (IL-21) is the newest member of the common gamma-chain family of cytokines, which includes IL-2, IL-4, IL-7, IL-9, IL-13, and IL-15. Its private receptor, IL-21R, has been shown to activate the Janus kinase/signal transducers and activators of transcription signaling pathway upon ligand binding. Initial studies have demonstrated that IL-21 has pleiotropic effects on the proliferation, differentiation, and effector functions of B, T, natural killer, and dendritic cells. More recently, the potential therapeutic capacity of IL-21 in the treatment of cancers has been widely investigated. The biological role of IL-21 in the immune system is complex, as IL-21 has been shown to have the ability to both promote and inhibit immune responses. Overall, the current data point to IL-21 being a novel immunomodulatory cytokine, whose regulation of any given immune response is highly dependent on the surrounding environmental context.
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http://dx.doi.org/10.1111/j.0105-2896.2004.00201.xDOI Listing
December 2004

Simultaneous deficiency in CD28 and STAT6 results in chronic ectoparasite-induced inflammatory skin disease.

Infect Immun 2004 Jul;72(7):3706-15

Department of Microbiology and Immunology, Uniformed Services University of the Health Sciences, 4301 Jones Bridge Rd., Bethesda, MD 20814, USA.

A mouse lacking CD28, a T-cell costimulatory molecule, and STAT6, a transcription factor that mediates interleukin-4 (IL-4) signaling, was developed from parental CD28- and STAT6-deficient mice. STAT6/CD28(-/-) BALB/c mice that were 8 weeks old had a normal phenotype, and IL-4 production was induced following infection with nematode parasites. Unexpectedly, when they were between 4 and 8 months old, all mice examined spontaneously developed severe chronic dermatitis associated with pronounced numbers of Demodex ectoparasites. In addition, pronounced CD4 and CD8 T-cell infiltrates in the dermis and subcutaneous fat, increased serum immunoglobulin G2a levels, and lymphadenopathy associated with increased gamma interferon and IL-12 expression were observed. Single-knockout siblings lacking either CD28 or STAT6 had a phenotype similar to that of BALB/c wild-type controls. To distinguish whether the ectoparasite Demodex or the Th1 immunity was the proximal cause of the inflammatory skin disease, STAT6/CD28(-/-) mice were treated with a miticide that eliminated the ectoparasites. This treatment markedly reduced the severity of the dermatitis and the associated lymphoid infiltrates. These findings suggest that ubiquitous ectoparasites, which are generally considered to be commensal, may contribute to disease when specific molecules required for an effective Th2 response are blocked.
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http://dx.doi.org/10.1128/IAI.72.7.3706-3715.2004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC427407PMC
July 2004

Defining Th1 and Th2 immune responses in a reciprocal cytokine environment in vivo.

J Immunol 2004 Apr;172(7):4260-5

Center for Neurologic Diseases, Laboratory of Immunogenetics and Transplantation, Brigham and Women's Hospital, Harvard School of Public Health, Harvard Medical School, Boston, MA 02115, USA.

The ability of committed Th1 and Th2 cells to function in altered cytokine environments is a central issue in autoimmune and immune-mediated diseases. Therefore, it is of interest to study the ability of Th1 or Th2 cells to expand and produce cytokine reciprocal environments in vivo. Using STAT4- and STAT6-deficient mice, we studied the expansion and cytokine production of Ag-specific Th1 or Th2 cells after transfer into Th1, Th2, or wild-type recipients. Our data show that these Th1 or Th2 cells proliferated and clonally expanded normally, regardless of the in vivo cytokine environment. These data have implications for the treatment of immune-mediated diseases by immunomodulatory agents that alter the cytokine milieu in vivo.
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http://dx.doi.org/10.4049/jimmunol.172.7.4260DOI Listing
April 2004