BRD4 regulates genes involved in cell cycle progression from G0, G1 through S, thus impacting on cell growth. Recently, it has been shown that BRD4 is required for induction of inflammatory genes in macrophages, suggesting its role in innate immunity. We asked whether BRD4 plays a role in transcription of interferon (IFN) stimulated genes (ISGs). Type I IFN activates a large number of genes through the JAK/STAT pathway and establishes natural resistance against viruses and other pathogens. We found that IFN stimulation triggers recruitment of BRD4 to the transcription start site (TSS) of a number of ISGs. Recruitment of BRD4 to ISGs coincided with increased histone acetylation at the TSS and induction of ISG mRNAs. Further, BRD4 recruitment coincided with the binding of the elongation factor, P-TEFb, which is known to interact with BRD4. We also show that the pausing complex, NELF and DSIF are recruited to the ISGs after IFN stimulation. IFN-induced NELF/DSIF recruitment was unexpected, since the pausing complex was reported to be present prior to stimulation and dissociate after activation as studed for some inducible genes. Furthermore, another elongation factor, SPT6 was recruited to ISGs after IFN stimulation, a factor known to function as a histone H3 chaperon. These results gave a picture where a number of elongation factors are induced to assemble on the ISGs. To delineate a hierarchical order of the assembly of these factors, we tested the effect of small molecule inhibitor that inhibits binding of acetylated histones to bromodomains. Results showed that this inhibitor not only inhibits BRD4 recruitment, but recruitment of P-TEFb, NELF/DSIF and SPT6. In addition, the inhibitor markedly reduced ISG transcription. Similarly shRNA-based BRD4 knockdown led to reduced recruitment all of above factors. These data support a view that BRD4 recruitment is the primary event that initiates a cascade of factor binding that shapes overall ISG transcription. Additional knockdown studies showed that NELF and SPT6 have opposing activities in elongation, the former repressing ISG transcription, while the latter promoting transcription. Together, our results indicate that BRD4 plays a central role in SG transcription for its ability to sequentially recruit various elongation factors. These studies are extended to analysis of Brd4f/f mice expressing cell type specific Cre.

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8
Fiscal Year
2014
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U.S. National Inst/Child Hlth/Human Dev
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Kamada, Rui; Yang, Wenjing; Zhang, Yubo et al. (2018) Interferon stimulation creates chromatin marks and establishes transcriptional memory. Proc Natl Acad Sci U S A 115:E9162-E9171
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Huang, X F; Nandakumar, V; Tumurkhuu, G et al. (2016) Mysm1 is required for interferon regulatory factor expression in maintaining HSC quiescence and thymocyte development. Cell Death Dis 7:e2260

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