Regulating type 1 IFN effects in CD8 T cells during viral infections: changing STAT4 and STAT1 expression for function
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE40666 | GEO | 2012/09/07
SECONDARY ACCESSION(S): PRJNA174526
REPOSITORIES: GEO
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Project description:Type 1 IFNs can conditionally activate all of the signal transducers and activators of transcription molecules (STATs), including STAT4. The best-characterized signaling pathways use STAT1, however, and type 1 IFN inhibition of cell proliferation is STAT1 dependent. We report that type 1 IFNs can basally stimulate STAT1- and STAT4- dependent effects in CD8 T cells, but that CD8 T cells responding to infections of mice with lymphocytic choriomenigitis virus have elevated STAT4 and lower STAT1 expression with significant consequences for modifying the effects of type 1 IFN exposure. The phenotype was associated with preferential type 1 IFN activation of STAT4 as compared to STAT1. Stimulation through the TCR induced elevated STAT4 expression, and STAT4 was required for peak expansion of antigen-specific CD8 T cells, low STAT1 levels, and resistance to type 1 IFN-mediated inhibition of proliferation. Thus, a mechanism is discovered for regulating the consequences of type 1 IFN exposure in CD8 T cells, with STAT4 acting as a key molecule in driving optimal antigen-specific responses and overcoming STAT1-dependent inhibition of proliferation. CD8 T cells were purified from uninfected WT, STAT1-deficient and STAT4-deficient mice or from D8 LCMV-infected WT mice and either control treated or treated with 1x104 U mouse IFNalpha for 90 minutes.
2012-09-07 | E-GEOD-40666 | biostudies-arrayexpress
Project description:Compared to primary human monocytes in whole blood cultures, few B cells activated STAT1 in response to stimulation of 2000 IU/ml IFN-beta for 45 minutes. Because activation of STAT1 leads to apoptosis induction, we tested the hypothesis that less pro-apoptotic genes are induced in B cells as compared to monocytes. Manuscript titled: Major differences in the responses of primary human leukocyte subsets to IFN-beta. Abstract: Treatment of cell lines with type I IFNs activates the formation of ISGF3 (STAT1/STAT2/ IRF9), which induces the expression of many genes. To study this response in primary cells, we treated fresh human blood with IFN-beta and used flow cytometry to analyze phosphorylated STATs1, 3 and 5 in CD4+ and CD8+ T cells, B cells, and monocytes. The activation of STAT1 was remarkably different among these leukocyte subsets. In contrast to monocytes, CD4+ and CD8+ T cells, few B cells activated STAT1 in response to IFN-beta, a finding that could not be explained by decreased levels of IFNAR2 or STAT1 or enhanced levels of SOCS1 or relevant protein tyrosine phosphatases in B cells. Micro-array and real-time PCR analyses revealed the induction of STAT1-dependent pro-apoptotic mRNAs in monocytes but not in B cells. These data show that ISGF3 or STAT1 homodimers are not the main activators of gene expression in primary B cells of healthy humans. Notably, in B cells and especially in CD4+ T cells, IFN-beta activated STAT5 in addition to STAT3, with biological effects often opposite from those driven by activated STAT1. These data help to explain why IFN-beta increases the survival of primary human B cells and CD4+ T cells, but enhances the apoptosis of monocytes, and also to understand how leukocyte subsets are differentially affected by endogenous type I IFNs during viral or bacterial infections, and by type I IFN treatment of patients with multiple sclerosis, hepatitis or cancer. Undiluted whole blood of 2 healthy individuals (HI) was used, and either stimulated with IFN-beta or left untreated (control) for 3 hrs. After 3 hrs, both B cells and monocytes were isolated from whole blood of the first healthy individual, and only B cells were isolated from whole blood of the second healthy individual.
2010-10-20 | E-GEOD-23307 | biostudies-arrayexpress
Project description:Compared to primary human monocytes in whole blood cultures, few B cells activated STAT1 in response to stimulation of 2000 IU/ml IFN-beta for 45 minutes. Because activation of STAT1 leads to apoptosis induction, we tested the hypothesis that less pro-apoptotic genes are induced in B cells as compared to monocytes. Manuscript titled: Major differences in the responses of primary human leukocyte subsets to IFN-beta. Abstract: Treatment of cell lines with type I IFNs activates the formation of ISGF3 (STAT1/STAT2/ IRF9), which induces the expression of many genes. To study this response in primary cells, we treated fresh human blood with IFN-beta and used flow cytometry to analyze phosphorylated STATs1, 3 and 5 in CD4+ and CD8+ T cells, B cells, and monocytes. The activation of STAT1 was remarkably different among these leukocyte subsets. In contrast to monocytes, CD4+ and CD8+ T cells, few B cells activated STAT1 in response to IFN-beta, a finding that could not be explained by decreased levels of IFNAR2 or STAT1 or enhanced levels of SOCS1 or relevant protein tyrosine phosphatases in B cells. Micro-array and real-time PCR analyses revealed the induction of STAT1-dependent pro-apoptotic mRNAs in monocytes but not in B cells. These data show that ISGF3 or STAT1 homodimers are not the main activators of gene expression in primary B cells of healthy humans. Notably, in B cells and especially in CD4+ T cells, IFN-beta activated STAT5 in addition to STAT3, with biological effects often opposite from those driven by activated STAT1. These data help to explain why IFN-beta increases the survival of primary human B cells and CD4+ T cells, but enhances the apoptosis of monocytes, and also to understand how leukocyte subsets are differentially affected by endogenous type I IFNs during viral or bacterial infections, and by type I IFN treatment of patients with multiple sclerosis, hepatitis or cancer.
2010-10-20 | GSE23307 | GEO
Project description:Type-I (e.g. IFN-alpha, IFN-beta) and type-II IFNs (IFN-gamma) have antiviral, antiproliferative, and immunomodulatory properties. Both types of IFN signal through the Jak/STAT pathway to elicit antiviral activity, yet IFN-gamma is thought to do so only through STAT1 homodimers while type-I IFNs activate both STAT1- and STAT2-containing complexes such as ISGF3. Here we show that ISGF3II - composed of phosphorylated STAT1, unphosphorylated STAT2, and IRF9 - also plays a role in IFN-gamma-mediated antiviral activity in humans. Using phosphorylated STAT1 as a marker for IFN signaling, western blot analysis of IFN-alpha2a treated human A549 cells revealed that pSTAT1 (Y701) levels peaked at 1h, decreased by 6h, and remained at low levels for up to 48h. Cells treated with IFN-gamma showed a biphasic pSTAT1 response with an early peak at 1-2h and a second peak at 15-24h. Gene expression microarray following IFN-gamma treatment for 24h indicated an induction of antiviral genes that are induced by ISGF3 and associated with a type-1 IFN response. Induction of these genes by autocrine type-I and type-III IFN signaling was ruled out using neutralizing antibodies to these IFNs in biological assays and by qRT-PCR. Despite the absence of autocrine IFNs, IFN-gamma treatment induced formation of ISGF3II. This novel transcription factor complex binds to ISRE promoter sequences, as shown by ChIP analysis of the PKR promoter. STAT2 and IRF9 knockdown in A549 cells reversed IFN-gamma-mediated ISRE induction and antiviral activity - implicating ISGF3II formation as a significant component of the cellular response and biological activity of IFN-gamma. Two treatments using three biological replicates each were performed using three million A549 cells. Each was seeded overnight in 10mL complete RPMI and treated. Three were treated with alpha-IFN and three treated with gamma-IFN for 24h. Control samples were left untreated.
2011-01-19 | E-GEOD-25113 | biostudies-arrayexpress
Project description:Type I interferons (IFNs) play a key role in linking the innate and adaptive immune responses. IFNs can have both positive and negative effects on the development of memory CD8+ T cells, but their contribution to the priming of naïve T cells is unclear. Using OT-I TCR transgenic T cells, we show strong effects on TCR-induced functional response when sensitized by IFN-β prior to their stimulation with cognate antigen. These effects are only seen in the absence of co-stimulation and at suboptimal TCR stimulation conditions. Pre-stimulation of naïve T cells inhibits certain TCR-mediated functions like cell proliferation and the induction of genes such as IFN-γ and IL-2. However, type I IFN does not influence the induction of other TCR-induced functions like the expression of the T cell activation marker CD25 and a number of other genes. IFN-mediated impairment of proliferation is due to the lack of IL-2 induction since it can be compensated by exogenous IL-2. Further analysis reveals an IFN-mediated inhibition of Ca2+ flux and ERK phosphorylation. In vivo studies confirm that pre-stimulation with IFNs inhibits early T cell responses, whereas long-term responses are positively affected. Our results reveal that IFN exerts negative effects on naïve T cells in a timely defined range under sub-optimal TCR stimulation conditions. These results highlight the importance of timing of immune-regulatory events and reveal a novel role for type I IFNs by shaping the immune response, such that T cells are not able to react until optimal stimulation is provided. 8 Arrays, untreated cells as references, IFNb treatment and/or TCR stimulation at various lengths of time
2015-12-31 | E-GEOD-41381 | biostudies-arrayexpress
Project description:Type-I (e.g. IFN-alpha, IFN-beta) and type-II IFNs (IFN-gamma) have antiviral, antiproliferative, and immunomodulatory properties. Both types of IFN signal through the Jak/STAT pathway to elicit antiviral activity, yet IFN-gamma is thought to do so only through STAT1 homodimers while type-I IFNs activate both STAT1- and STAT2-containing complexes such as ISGF3. Here we show that ISGF3II - composed of phosphorylated STAT1, unphosphorylated STAT2, and IRF9 - also plays a role in IFN-gamma-mediated antiviral activity in humans. Using phosphorylated STAT1 as a marker for IFN signaling, western blot analysis of IFN-alpha2a treated human A549 cells revealed that pSTAT1 (Y701) levels peaked at 1h, decreased by 6h, and remained at low levels for up to 48h. Cells treated with IFN-gamma showed a biphasic pSTAT1 response with an early peak at 1-2h and a second peak at 15-24h. Gene expression microarray following IFN-gamma treatment for 24h indicated an induction of antiviral genes that are induced by ISGF3 and associated with a type-1 IFN response. Induction of these genes by autocrine type-I and type-III IFN signaling was ruled out using neutralizing antibodies to these IFNs in biological assays and by qRT-PCR. Despite the absence of autocrine IFNs, IFN-gamma treatment induced formation of ISGF3II. This novel transcription factor complex binds to ISRE promoter sequences, as shown by ChIP analysis of the PKR promoter. STAT2 and IRF9 knockdown in A549 cells reversed IFN-gamma-mediated ISRE induction and antiviral activity - implicating ISGF3II formation as a significant component of the cellular response and biological activity of IFN-gamma.
2011-01-19 | GSE25113 | GEO
Project description:CD4+ T follicular helper cells (TFH) are critical for the formation and function of B cell responses to infection or immunization, but also play an important role in autoimmunity. The factors that contribute to the differentiation of this helper cell subset are incompletely understood, although several cytokines including IL-6, IL-21 and IL-12 can promote TFH cell formation. Yet, none of these factors, nor their downstream cognate STATs, have emerged as non-redundant, essential drivers of TFH cells. This suggests a model in which multiple factors can contribute to the phenotypic characteristics of TFH cells. As type I interferons (IFNs) are often generated in immune responses, we set out to investigate if these factors are relevant to TFH cell differentiation. Type I IFNs promote Th1 responses, thus one possibility was these factors antagonized TFH-expressed genes. However, we show that type I IFNs (IFN-α/β) induced Bcl6 expression, the master regulator transcription factor for TFH cells, and CXCR5 and PD-1 (encoded by Pdcd1), key surface molecules expressed by TFH cells. In contrast, type I IFNs failed to induce IL-21, the signature cytokine for TFH cells. The induction of Bcl6 was regulated directly by STAT1, which bound to the Bcl6, Cxcr5 and Pdcd1 loci. These data suggest that type I IFNs (IFN-α/β) and STAT1 can contribute to some features of TFH cells but are inadequate in inducing complete programming of this subset. The role of STAT1 in type I interferon treated CD4+ T cells was investigated by Chip-seq of STAT1.
2014-01-22 | E-GEOD-51531 | biostudies-arrayexpress
Project description:Interferons (IFNs) are pleotropic cytokines secreted upon encounter of pathogens and tumors. Applying their antipathogenic, antiproliferative and immune stimulatory capacities, recombinant IFNs are frequently prescribed as drugs to treat different diseases. IFNs act by changing the transcription of cells. Due to characteristics like rapid induction and signaling, IFNs represent prototypic model systems for various aspects of biomedical research. In respect to the signaling and activated promoters, IFNs can be subdivided in two groups. Here, alterations of the cellular proteome of human cells treated with IFN and IFN were elucidated in a time-dependent manner by mass spectrometry-based quantitative proteome analysis. The majority of protein regulations were strongly IFN type- and time-dependent. In addition to the expected upregulation of IFN-induced proteins, an astonishing number of proteins become profoundly repressed especially by IFN. Taken together, our comprehensive analysis deciphers the human interferome and its kinetics of protein induction and repression.
2018-05-25 | PXD006442 | Pride
Project description:A number of IFN-induced proteins are believed to be responsible for the antiviral state induced by IFNs. Our microarray analysis of IFN-regulated genes in MEFs from wild-type mice identified 124 probe sets that were differentially regulated upon IFN treatment. This group consisted of many known ISGs such as Ifit1, Gbp2, mx1, Isg15, Stat1, nmi, mx2, If204, Adar, Irf1, and protein kinase R. Experiment Overall Design: 3 control and 3 IFNb-treated biological replicates were analyzed.
2007-10-12 | E-GEOD-3400 | biostudies-arrayexpress
Project description:Type I interferons (IFNs) play a key role in linking the innate and adaptive immune responses. IFNs can have both positive and negative effects on the development of memory CD8+ T cells, but their contribution to the priming of naïve T cells is unclear. Using OT-I TCR transgenic T cells, we show strong effects on TCR-induced functional response when sensitized by IFN-β prior to their stimulation with cognate antigen. These effects are only seen in the absence of co-stimulation and at suboptimal TCR stimulation conditions. Pre-stimulation of naïve T cells inhibits certain TCR-mediated functions like cell proliferation and the induction of genes such as IFN-γ and IL-2. However, type I IFN does not influence the induction of other TCR-induced functions like the expression of the T cell activation marker CD25 and a number of other genes. IFN-mediated impairment of proliferation is due to the lack of IL-2 induction since it can be compensated by exogenous IL-2. Further analysis reveals an IFN-mediated inhibition of Ca2+ flux and ERK phosphorylation. In vivo studies confirm that pre-stimulation with IFNs inhibits early T cell responses, whereas long-term responses are positively affected. Our results reveal that IFN exerts negative effects on naïve T cells in a timely defined range under sub-optimal TCR stimulation conditions. These results highlight the importance of timing of immune-regulatory events and reveal a novel role for type I IFNs by shaping the immune response, such that T cells are not able to react until optimal stimulation is provided.
2015-12-31 | GSE41381 | GEO