Gene Regulatory Sequences And Protein Binding in Genome Sequences

National Library Of Medicine

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Abstract

We worked on 3 projects in the past 12 months. The first one is about the spatial organization of gene transcription regulatory elements. Using transcription-factor binding enrichment as an indicator of enhancer strength, we identified a portion of H3K27ac peaks as potentially strong enhancers and discovered a universal pattern of promoter and enhancer distribution: at actively transcribed regions of length of 200300 kb, the numbers of active promoters and enhancers are inversely related. Enhancer clusters are associated with isolated active promoters, regardless of the gene's cell-type specificity. As the number of nearby active promoters increases, the number of enhancers decreases. At regions where multiple active genes are closely located, there are few distant enhancers. Using Hi-C analysis, we demonstrate that the interactions among the regulatory elements (active promoters and enhancers) occur predominantly in clusters and multiway among linearly close elements and the distance between adjacent elements shows a preference of 30 kb. We propose a simple rule of spatial organization of active promoters and enhancers: gene transcription and regulation mainly occurs at local active transcription hubs contributed to dynamically by multiple elements from linearly close enhancers and/or active promoters. The hub model can be represented by a flower-shaped structure and implies an enhancer-like role of active promoters. Another project is a collaboration with Dr. Randall Morses group from Wadsworth Center of New York. The project concerns Mediator occupancy and dynamic at promoters of gene cohorts induced by stress or alternative growth conditions. By performing ChIP-seq analysis in yeast strains with different genetic background, we investigated Mediator occupancy following heat shock or CdCl2 induction, with or without depletion of Kin28. We find that Pol II occupancy exhibits similar dependence on Mediator under normal and heat shock conditions, and that Mediator occupancy at promoters increases upon Kin28 depletion after heat shock as it does under normal growth conditions, indicating similar transient occupancy. Unexpectedly, Mediator occupancy persists at genes repressed by heat shock or CdCl2 induction and, is shifted upstream from the proximal promoter region under conditions of Kin28 depletion, suggesting that Mediator is recruited by activators but is unable to engage PIC components at these repressed targets. Finally, we show a reduced dependence on PIC components for Mediator occupancy at promoters after heat shock, suggesting an altered dynamics or stronger engagement with activators under these conditions. This work was submitted to Genome Research and is under revision. The third project is about the role of HMGN proteins on 3D chromatin structure. HMGNs are architectural proteins that bind to nucleosomes and modulate the structure and function of chromatin. In order to know whether they affect 3D chromatin structure, we did Hi-C sequencing and promoter capture Hi-C sequence in wild type (WT) and HMGN1/HMGN2 double knock (DKO) cells. We found that HMGN proteins are mostly located within compartment A chromatin and they show remarkable increase crossing boundaries from compartment B to A. The overall 3D chromatin structures are very similar between WT and DKO cells, which indicates that HMGN proteins do not play critical roles in maintaining 3D chromatin structure.