The TFIID components TAF7 and TAF1 regulate eukaryotic transcription initiation. TAF7 regulates transcription initiation of TAF1-dependent genes by binding to the acetyltransferase (AT) domain of TAF1 and inhibiting the enzymatic activity that is essential for transcription. TAF7 is released from the TAF1/TFIID complex upon completion of preinitiation complex assembly, allowing transcription to initiate. However, not all transcription is TAF1-dependent and the role of TAF7 in regulating TAF1-independent transcription has not been defined. The IFN-gamma--induced transcription factor CIITA is a coactivator and general transcription factor that regulates both MHC Class I and Class II gene transcription, and is thus critical to activated immune response. Known as the master regulator of Class II expression, CIITA is required for both Class II transcription and activated Class I expression. Its deficiency is linked to bare lymphocyte syndrome. Comparison of eukaryotic basal class I transcription and interferon-activated transcription initiation reveals similar preinitiation complex recruitment mechanisms, and striking parallels between their respective critical components, the TATA-binding protein (TBP)-associated factor 1 (TAF1) and CIITA. Both TAF1 and CIITA possess instrinsic acetyltransferase (AT) activity required to activate MHC transcription, which is regulated by TAF7. Moreover, CIITA can bypass the requirement for TAF1 to activate both the MHC class I and II promoters. TAF1 has two distinct kinase activities, located with its amino-terminal and carboxy-terminal domains. However, despite the striking functional parallels between CIITA and TAF1, no similar kinase activity has been reported for CIITA thus far. We have now identified the transcriptional coactivator CIITA as a novel atypical serine-threonine kinase whose substrates include various general transcription factors. We have characterized the kinase activity of the protein and mapped the putative kinase domains. We have proposed a model in which the kinase activity of CIITA serves a function similar to that of TAF1, in which it regulates TAF7 binding and release, and thus MHC transcription initiation. This may elucidate a novel role for CIITA in the regulation of activated transcription initiation and stimulated immune response during pathogenic infection. To further characterize regulation of class I gene expression, we have undertaken to characterize the role of core promoter elements in hormonal responses in vivo. The minimal class I core promoter has been localized to a segment between -65 bp and +14 bp. Contained within this segment are a canonical CCAAT box, a TATAA-like element, an Sp1 binding site (Sp1BS) and an Initiator (Inr). In past studies, we have reported that no single element is necessary for expression of a reporter in transient transfection assays. However, we have now examined the dependence of transcription on each of the core promoter elements in the context of the native gene in transgenic mice. Remarkably, all of the mutant promoters supported transcription. Each element contributed uniquely to tissue-specificity, hormonal responses or both. Whereas the WT, TATAA-like and CCAAT mutant promoters were activated by gamma-interferon, the Inr and Sp1 mutants were repressed, implicating these elements in regulation of hormonal responses. Transcription initiation is not a continuous process, but rather occurs in bursts which differ in amplitude and frequency. Although the roles of enhancers on the bursting characteristics of promoters in yeast and cultured mammalian cells have been described, the contributions of individual core promoter elements to bursting amplitude and frequency have not been assessed. Having demonstrated that the core promoter elements that constitute the MHC class I promoter all contribute to transcription initiation, we have investigated whether interferon-activated transcription affects burst amplitude, frequency orboth and the role of the core promoter elements in determining these bursting characteristics in primary splenic B cells by single molecule RNA FISH. We have demonstrated that each core promoter element modulates different components of transcriptional bursting. Remarkably, the activities of the core promoter elements differ during basal or activated transcription. Thus, in primary B cells, SP1BS governs transcriptional burst size under basal, constitutive conditions but burst frequency in response to gamma-interferon induction. Conversely, while the Inr regulates burst frequency in the absence of gamma-interferon, it contributes to increased burst size in response to gamma-interferon induction. The TATAA-like element regulates burst size and frequency under both conditions. Paradoxically, in the presence of gamma-interferon, the burst frequency of the mutated TATAA-like element is reduced while size is increased, such that the overall effect was a modest increase in mean mRNA/cell, but an increase in noise. Finally, the CCAAT element does not contribute to bursting during constitutive transcription. However, in response to gamma-interferon activation, the CCAAT box modulates both burst size and frequency. The complex alterations in bursting of the core promoter element mutations in both constitutive and activated transcription indicate that core promoter architecture determines the specific bursting properties and enforces their roles in mediating proper response to signaling events.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
1ZIABC009285-31
Application #
9556230
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
31
Fiscal Year
2017
Total Cost
Indirect Cost
Name
Basic Sciences
Department
Type
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State
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Devaiah, Ballachanda N; Case-Borden, Chanelle; Gegonne, Anne et al. (2016) BRD4 is a histone acetyltransferase that evicts nucleosomes from chromatin. Nat Struct Mol Biol 23:540-8
Roy, Ananda L; Singer, Dinah S (2015) Core promoters in transcription: old problem, new insights. Trends Biochem Sci 40:165-71
Soe, Katherine C; Devaiah, Ballachanda N; Singer, Dinah S (2013) Transcriptional coactivator CIITA, a functional homolog of TAF1, has kinase activity. Biochim Biophys Acta 1829:1184-90
Devaiah, Ballachanda N; Singer, Dinah S (2013) CIITA and Its Dual Roles in MHC Gene Transcription. Front Immunol 4:476
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