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Recruitment of histone deacetylase 3 to the interferon-A gene promoters attenuates interferon expression.

Génin P, Lin R, Hiscott J, Civas A - PLoS ONE (2012)

Bottom Line: Analysis of chromatin-protein association by Chip-QPCR demonstrated that recruitment of interferon regulatory factor (IRF)3 and IRF7, as well as TBP correlated with enhanced histone H3K9 and H3K14 acetylation, whereas recruitment of HDAC3 correlated with inhibition of histone H3K9/K14 acetylation, removal of IRF7 and TATA-binding protein (TBP) from IFN-A promoters and inhibition of virus-induced IFN-A gene transcription.Additionally, HDAC3 overexpression reduced, and HDAC3 depletion by siRNA enhanced IFN-A gene expression.Furthermore, activation of IRF7 enhanced histone H3K9/K14 acetylation and IFN-A gene expression, whereas activation of both IRF7 and IRF3 led to recruitment of HDAC3 to the IFN-A gene promoters, resulting in impaired histone H3K9 acetylation and attenuation of IFN-A gene transcription.

View Article: PubMed Central - PubMed

Affiliation: Centre National de la Recherche Scientifique-FRE3235, Paris Descartes University, Paris, France.

ABSTRACT

Background: Induction of Type I Interferon (IFN) genes constitutes an essential step leading to innate immune responses during virus infection. Sendai virus (SeV) infection of B lymphoid Namalwa cells transiently induces the transcriptional expression of multiple IFN-A genes. Although transcriptional activation of IFN-A genes has been extensively studied, the mechanism responsible for the attenuation of their expression remains to be determined.

Principal findings: In this study, we demonstrate that virus infection of Namalwa cells induces transient recruitment of HDAC3 (histone deacetylase 3) to IFN-A promoters. Analysis of chromatin-protein association by Chip-QPCR demonstrated that recruitment of interferon regulatory factor (IRF)3 and IRF7, as well as TBP correlated with enhanced histone H3K9 and H3K14 acetylation, whereas recruitment of HDAC3 correlated with inhibition of histone H3K9/K14 acetylation, removal of IRF7 and TATA-binding protein (TBP) from IFN-A promoters and inhibition of virus-induced IFN-A gene transcription. Additionally, HDAC3 overexpression reduced, and HDAC3 depletion by siRNA enhanced IFN-A gene expression. Furthermore, activation of IRF7 enhanced histone H3K9/K14 acetylation and IFN-A gene expression, whereas activation of both IRF7 and IRF3 led to recruitment of HDAC3 to the IFN-A gene promoters, resulting in impaired histone H3K9 acetylation and attenuation of IFN-A gene transcription.

Conclusion: Altogether these data indicate that reversal of histone H3K9/K14 acetylation by HDAC3 is required for attenuation of IFN-A gene transcription during viral infection.

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Recruitment of IRF3, IRF7 and TBP to IFN-A gene promoters.Recruitment to the IFN-A2, A14 and A1 gene promoters was determined by ChIP-QPCR assays in Namalwa B cells infected by Sendai virus. Cross-linked chromatin extracts were immunoprecipitated with antibodies specific for TBP, IRF7 or IRF3 and submitted to QPCR analysis after reversion of cross-linking, as described in Materials and Methods. Anti-rabbit IgG was used to determine background levels of non-specific binding. Data are expressed as % of the DNA input. Values are mean and standard deviation for two replicate samples derive from two representative experiments (n = 4). The error bars indicate the relative standard deviations.
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pone-0038336-g002: Recruitment of IRF3, IRF7 and TBP to IFN-A gene promoters.Recruitment to the IFN-A2, A14 and A1 gene promoters was determined by ChIP-QPCR assays in Namalwa B cells infected by Sendai virus. Cross-linked chromatin extracts were immunoprecipitated with antibodies specific for TBP, IRF7 or IRF3 and submitted to QPCR analysis after reversion of cross-linking, as described in Materials and Methods. Anti-rabbit IgG was used to determine background levels of non-specific binding. Data are expressed as % of the DNA input. Values are mean and standard deviation for two replicate samples derive from two representative experiments (n = 4). The error bars indicate the relative standard deviations.

Mentions: ChIP-QPCR experiments were next performed to establish whether transient IFN-A gene expression correlated with the recruitment pattern of TATA-binding protein (TBP) to different IFN-A gene promoters following virus infection. The high sequence homology within IFN-A gene promoter regions, extending 400 bp upstream from the transcription initiation site [53], [54], prompted us to select IFN-A1, A2, A7 and A14 genes for analysis. Recruitment of TBP to the IFN-A promoters was detected at 6–12 h p.i (an average of 2% of the input, with p<0.01), with 2- to 4-fold enrichment calculated for IFN-A2, A14 and A1 (Fig. 2). TBP recruitment correlated with maximal IFN-A gene expression, and decreased at 14 h p.i (% of the input values comparable to those determined in uninduced cells). These results indicated that the removal of TBP from IFN-A gene promoters was concomitant with the inhibition of gene expression, occurring rapidly after gene induction. We also determined the dynamics of IRF3 and IRF7 recruitment to the IFN-A gene promoters in the course of virus infection. Both factors were recruited to the IFN-A1, A2 and IFN-A14 gene promoters following virus infection (Fig. 2); occupancy by IRF7 and IRF3 reached maximal values at 8–10 h; IRF7 enrichment of the promoters then decreased at 14 h p.i, concomitant with transcriptional inhibition, while significant recruitment of IRF3 was observed up to 16 h following infection. These data indicated that virus-induced IFN-A gene expression correlated with the transient recruitment of IRF7 and TBP to the IFN-A gene promoters. They also indicated that removal of both IRF7 and TBP from the IFN-A gene promoters was associated with considerably reduced levels of IFN-A gene expression. Higher levels of IRF7 recruitment to IFN-A14 gene promoter (in comparison to IFN-A1 or A2) were in agreement with previous results indicating the high responsiveness of this promoter to IRF7-mediated transcription [22]. Similarly, high levels of IRF3 recruitment might be related to the presence of IRF sites exhibiting preferential binding for IRF3 in the IFN-A1 and A2 gene promoters [22]. Sustained recruitment of IRF3 from 8 to 16 h p.i to IFN-A gene promoters during virus-induced activation and inhibition of gene expression suggested that this factor might be involved both in the initiation and post-inductional repression of IFN-A gene transcription.


Recruitment of histone deacetylase 3 to the interferon-A gene promoters attenuates interferon expression.

Génin P, Lin R, Hiscott J, Civas A - PLoS ONE (2012)

Recruitment of IRF3, IRF7 and TBP to IFN-A gene promoters.Recruitment to the IFN-A2, A14 and A1 gene promoters was determined by ChIP-QPCR assays in Namalwa B cells infected by Sendai virus. Cross-linked chromatin extracts were immunoprecipitated with antibodies specific for TBP, IRF7 or IRF3 and submitted to QPCR analysis after reversion of cross-linking, as described in Materials and Methods. Anti-rabbit IgG was used to determine background levels of non-specific binding. Data are expressed as % of the DNA input. Values are mean and standard deviation for two replicate samples derive from two representative experiments (n = 4). The error bars indicate the relative standard deviations.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3369917&req=5

pone-0038336-g002: Recruitment of IRF3, IRF7 and TBP to IFN-A gene promoters.Recruitment to the IFN-A2, A14 and A1 gene promoters was determined by ChIP-QPCR assays in Namalwa B cells infected by Sendai virus. Cross-linked chromatin extracts were immunoprecipitated with antibodies specific for TBP, IRF7 or IRF3 and submitted to QPCR analysis after reversion of cross-linking, as described in Materials and Methods. Anti-rabbit IgG was used to determine background levels of non-specific binding. Data are expressed as % of the DNA input. Values are mean and standard deviation for two replicate samples derive from two representative experiments (n = 4). The error bars indicate the relative standard deviations.
Mentions: ChIP-QPCR experiments were next performed to establish whether transient IFN-A gene expression correlated with the recruitment pattern of TATA-binding protein (TBP) to different IFN-A gene promoters following virus infection. The high sequence homology within IFN-A gene promoter regions, extending 400 bp upstream from the transcription initiation site [53], [54], prompted us to select IFN-A1, A2, A7 and A14 genes for analysis. Recruitment of TBP to the IFN-A promoters was detected at 6–12 h p.i (an average of 2% of the input, with p<0.01), with 2- to 4-fold enrichment calculated for IFN-A2, A14 and A1 (Fig. 2). TBP recruitment correlated with maximal IFN-A gene expression, and decreased at 14 h p.i (% of the input values comparable to those determined in uninduced cells). These results indicated that the removal of TBP from IFN-A gene promoters was concomitant with the inhibition of gene expression, occurring rapidly after gene induction. We also determined the dynamics of IRF3 and IRF7 recruitment to the IFN-A gene promoters in the course of virus infection. Both factors were recruited to the IFN-A1, A2 and IFN-A14 gene promoters following virus infection (Fig. 2); occupancy by IRF7 and IRF3 reached maximal values at 8–10 h; IRF7 enrichment of the promoters then decreased at 14 h p.i, concomitant with transcriptional inhibition, while significant recruitment of IRF3 was observed up to 16 h following infection. These data indicated that virus-induced IFN-A gene expression correlated with the transient recruitment of IRF7 and TBP to the IFN-A gene promoters. They also indicated that removal of both IRF7 and TBP from the IFN-A gene promoters was associated with considerably reduced levels of IFN-A gene expression. Higher levels of IRF7 recruitment to IFN-A14 gene promoter (in comparison to IFN-A1 or A2) were in agreement with previous results indicating the high responsiveness of this promoter to IRF7-mediated transcription [22]. Similarly, high levels of IRF3 recruitment might be related to the presence of IRF sites exhibiting preferential binding for IRF3 in the IFN-A1 and A2 gene promoters [22]. Sustained recruitment of IRF3 from 8 to 16 h p.i to IFN-A gene promoters during virus-induced activation and inhibition of gene expression suggested that this factor might be involved both in the initiation and post-inductional repression of IFN-A gene transcription.

Bottom Line: Analysis of chromatin-protein association by Chip-QPCR demonstrated that recruitment of interferon regulatory factor (IRF)3 and IRF7, as well as TBP correlated with enhanced histone H3K9 and H3K14 acetylation, whereas recruitment of HDAC3 correlated with inhibition of histone H3K9/K14 acetylation, removal of IRF7 and TATA-binding protein (TBP) from IFN-A promoters and inhibition of virus-induced IFN-A gene transcription.Additionally, HDAC3 overexpression reduced, and HDAC3 depletion by siRNA enhanced IFN-A gene expression.Furthermore, activation of IRF7 enhanced histone H3K9/K14 acetylation and IFN-A gene expression, whereas activation of both IRF7 and IRF3 led to recruitment of HDAC3 to the IFN-A gene promoters, resulting in impaired histone H3K9 acetylation and attenuation of IFN-A gene transcription.

View Article: PubMed Central - PubMed

Affiliation: Centre National de la Recherche Scientifique-FRE3235, Paris Descartes University, Paris, France.

ABSTRACT

Background: Induction of Type I Interferon (IFN) genes constitutes an essential step leading to innate immune responses during virus infection. Sendai virus (SeV) infection of B lymphoid Namalwa cells transiently induces the transcriptional expression of multiple IFN-A genes. Although transcriptional activation of IFN-A genes has been extensively studied, the mechanism responsible for the attenuation of their expression remains to be determined.

Principal findings: In this study, we demonstrate that virus infection of Namalwa cells induces transient recruitment of HDAC3 (histone deacetylase 3) to IFN-A promoters. Analysis of chromatin-protein association by Chip-QPCR demonstrated that recruitment of interferon regulatory factor (IRF)3 and IRF7, as well as TBP correlated with enhanced histone H3K9 and H3K14 acetylation, whereas recruitment of HDAC3 correlated with inhibition of histone H3K9/K14 acetylation, removal of IRF7 and TATA-binding protein (TBP) from IFN-A promoters and inhibition of virus-induced IFN-A gene transcription. Additionally, HDAC3 overexpression reduced, and HDAC3 depletion by siRNA enhanced IFN-A gene expression. Furthermore, activation of IRF7 enhanced histone H3K9/K14 acetylation and IFN-A gene expression, whereas activation of both IRF7 and IRF3 led to recruitment of HDAC3 to the IFN-A gene promoters, resulting in impaired histone H3K9 acetylation and attenuation of IFN-A gene transcription.

Conclusion: Altogether these data indicate that reversal of histone H3K9/K14 acetylation by HDAC3 is required for attenuation of IFN-A gene transcription during viral infection.

Show MeSH
Related in: MedlinePlus