Transcription from non-protein-coding DNA (ncDNA), including intergenic DNA, is widespread among eukaryotes. However, the consequences of this transcription are only beginning to be investigated. The long-term objective for this project is to understand how transcription of ncDNA regulates gene expression. This grant focuses on our recent discovery of a previously uncharacterized mechanism of gene regulation: repression by intergenic transcription. In Saccharomyces cerevisiae, we showed that expression of an intergenic transcript, SRG1, represses the transcription of the adjacent SER3 gene. However, we found that it is the act of transcription, not the SRG1 ncRNA product, which mediates repression by a transcription interference mechanism. In this proposal, we plan to extend these studies by using a combination of genetic, molecular, genomic and biochemical approaches. First, we will identify and characterize factors involved in both the activation and repression of SER3. The results of these studies will provide important insights into the mechanism of repression by intergenic transcription and establish tools to characterize the function of new cases of intergenic transcription. Second, we will begin to dissect the molecular details of this repression mechanism. Using both gene expression analysis and chromatin immunoprecipitation methods, we will determine the relationship between the rate of intergenic transcription and repression of the adjacent gene and test the hypothesis that intergenic transcription modifies chromatin structure to repress the adjacent gene. Third, we will identify and characterize new cases of repression by intergenic transcription. This will allow us to extend our analysis to more genes in order to understand the general features of this repression mechanism. We will focus our analysis on a subset of intergenic DNAs that represent strong candidates for repression by intergenic transcription based on results from our genome- wide RNA Pol II ChIP studies and more recent yeast whole-genome expression studies. It is becoming increasingly clear that transcription of ncDNA plays an important role in regulating normal cell growth and development in humans and thus is likely to be involved in the prevention of human diseases, including cancer. Given that factors and mechanisms of transcription regulation are largely conserved among eukaryotes, our analysis of repression by intergenic transcription in yeast may provide new insight into the roles of transcription of ncDNA in humans.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Molecular Genetics C Study Section (MGC)
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Tompkins, Laurie
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University of Pittsburgh
Schools of Arts and Sciences
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Hainer, Sarah J; Martens, Joseph A (2016) Regulation of chaperone binding and nucleosome dynamics by key residues within the globular domain of histone H3. Epigenetics Chromatin 9:17
Raupach, Elizabeth A; Martens, Joseph A; Arndt, Karen M (2016) Evidence for Regulation of ECM3 Expression by Methylation of Histone H3 Lysine 4 and Intergenic Transcription in Saccharomyces cerevisiae. G3 (Bethesda) 6:2971-81
Hainer, Sarah J; Charsar, Brittany A; Cohen, Shayna B et al. (2012) Identification of Mutant Versions of the Spt16 Histone Chaperone That Are Defective for Transcription-Coupled Nucleosome Occupancy in Saccharomyces cerevisiae. G3 (Bethesda) 2:555-67
Pruneski, Justin A; Hainer, Sarah J; Petrov, Kostadin O et al. (2011) The Paf1 complex represses SER3 transcription in Saccharomyces cerevisiae by facilitating intergenic transcription-dependent nucleosome occupancy of the SER3 promoter. Eukaryot Cell 10:1283-94
Hainer, Sarah J; Martens, Joseph A (2011) Identification of histone mutants that are defective for transcription-coupled nucleosome occupancy. Mol Cell Biol 31:3557-68
Thebault, Philippe; Boutin, Genevieve; Bhat, Wajid et al. (2011) Transcription regulation by the noncoding RNA SRG1 requires Spt2-dependent chromatin deposition in the wake of RNA polymerase II. Mol Cell Biol 31:1288-300
Pruneski, Justin A; Martens, Joseph A (2011) Transcription of intergenic DNA deposits nucleosomes on promoter to silence gene expression. Cell Cycle 10:1021-2
Hainer, Sarah J; Pruneski, Justin A; Mitchell, Rachel D et al. (2011) Intergenic transcription causes repression by directing nucleosome assembly. Genes Dev 25:29-40
Hainer, Sarah J; Martens, Joseph A (2011) Transcription of ncDNA: Many roads lead to local gene regulation. Transcription 2:120-123
Hartzog, Grant A; Martens, Joseph A (2009) ncRNA transcription makes its mark. EMBO J 28:1679-80