http://www.people.vcu.edu/~mreimers/SysBio/Li%20-%20Role%20of%20chromatin%20in%20transcription.pdf

The Role of Chromatin during Transcription

Rapid improvements in sequencing and array-based platforms are resulting in a flood of diverse genome-wide data, including data from exome and whole-genome sequencing, epigenetic surveys, expression profiling of coding and noncoding RNAs, single nucleotide polymorphism (SNP) and copy number profiling, and functional assays. Analysis of these large, diverse data sets holds the promise of a more comprehensive understanding of the genome and its relation to human disease. Experienced and knowledgeable human review is an essential component of this process, complementing computational approaches. This calls for efficient and intuitive visualization tools able to scale to very large data sets and to flexibly integrate multiple data types, including clinical data. However, the sheer volume and scope of data pose a significant challenge to the development of such tools.

Integrative Genomics Viewer

Organisms require an appropriate balance of stability and reversibility in gene expression programmes to maintain cell identity or to enable responses to stimuli; epigenetic regulation is integral to this dynamic control. Post-translational modification of histones by methylation is an important and widespread type of chromatin modification that is known to influence biological processes in the context of development and cellular responses. To evaluate how histone methylation contributes to stable or reversible control, we provide a broad overview of how histone methylation is regulated and leads to biological outcomes. The importance of appropriately maintaining or reprogramming histone methylation is illustrated by its links to disease and ageing and possibly to transmission of traits across generations.

/pdf/histone-methylation-a-dynamic-mark-in-health-disease-and-53e9mgdnmh.pdf

Histone methylation: a dynamic mark in health, disease and inheritance

Circadian clocks are endogenous oscillators that control 24-hour physiological and behavioural processes in organisms. These cell-autonomous clocks are composed of a transcription-translation-based autoregulatory feedback loop. With the development of next-generation sequencing approaches, biochemical and genomic insights into circadian function have recently come into focus. Genome-wide analyses of the clock transcriptional feedback loop have revealed a global circadian regulation of processes such as transcription factor occupancy, RNA polymerase II recruitment and initiation, nascent transcription, and chromatin remodelling. The genomic targets of circadian clocks are pervasive and are intimately linked to the regulation of metabolism, cell growth and physiology.

/pdf/transcriptional-architecture-of-the-mammalian-circadian-1a5gaw4u7a.pdf

Transcriptional architecture of the mammalian circadian clock

https://rnajc.ucsf.edu/sites/rnajc.ucsf.edu/files/cmyc.pdf

c-Myc regulates transcriptional pause release.

In the eukaryotic genome, the thousands of genes that encode messenger RNA are transcribed by a molecular machine called RNA polymerase II. Analysing the distribution and status of RNA polymerase II across a genome has provided crucial insights into the long-standing mysteries of transcription and its regulation. These studies identify points in the transcription cycle where RNA polymerase II accumulates after encountering a rate-limiting step. When coupled with genome-wide mapping of transcription factors, these approaches identify key regulatory steps and factors and, importantly, provide an understanding of the mechanistic generalities, as well as the rich diversities, of gene regulation.

/pdf/defining-mechanisms-that-regulate-rna-polymerase-ii-4m3xaapttp.pdf

Defining mechanisms that regulate RNA polymerase II transcription in vivo

Histone H3 lysine 4 trimethylation (H3K4me3) is a major hallmark of promoter-proximal histones at transcribed genes. Here, we report that a previously uncharacterized Drosophila H3K4 methyltransferase, dSet1, and not the other putative histone H3K4 methyltransferases (Trithorax; Trithorax-related protein), is predominantly responsible for histone H3K4 trimethylation. Functional and proteomics studies reveal that dSet1 is a component of a conserved H3K4 trimethyltransferase complex and polytene staining and live cell imaging assays show widespread association of dSet1 with transcriptionally active genes. dSet1 is present at the promoter region of all tested genes, including activated Hsp70 and Hsp26 heat shock genes and is required for optimal mRNA accumulation from the tested genes. In the case of Hsp70, the mRNA production defect in dSet1 RNAi-treated cells is accompanied by retention of Pol II at promoters. Our data suggest that dSet1-dependent H3K4me3 is responsible for the generation of a chromatin structure at active promoters that ensures optimal Pol II release into productive elongation.

/pdf/drosophila-set1-is-the-major-histone-h3-lysine-4-qq7jiu3t2e.pdf

Drosophila Set1 is the major histone H3 lysine 4 trimethyltransferase with role in transcription

Several eukaryotic transcription factors have been shown to modulate the elongation rate of RNA polymerase II (Pol II) on naked or chromatin-reconstituted templates in vitro. However, none of the tested factors have been shown to directly affect the elongation rate of Pol II in vivo. We performed a directed RNAi knock-down (KD) screen targeting 141 candidate transcription factors and identified multiple factors, including Spt6, that alter the induced Hsp70 transcript levels in Drosophila S2 cells. Spt6 is known to interact with both nucleosome structure and Pol II, and it has properties consistent with having a role in elongation. Here, ChIP assays of the first wave of Pol II after heat shock in S2 cells show that KD of Spt6 reduces the rate of Pol II elongation. Also, fluorescence recovery after photobleaching assays of GFP-Pol II in salivary gland cells show that this Spt6-dependent effect on elongation rate persists during steady-state-induced transcription, reducing the elongation rate from ∼1100 to 500 bp/min. Furthermore, RNAi depletion of Spt6 reveals its broad requirement during different stages of development.

https://www.embopress.org/doi/pdf/10.1038/emboj.2009.56

Spt6 enhances the elongation rate of RNA polymerase II in vivo

The Paf1 complex in yeast has been reported to influence a multitude of steps in gene expression through interactions with RNA polymerase II (Pol II) and chromatin-modifying complexes; however, it is unclear which of these many activities are primary functions of Paf1 and are conserved in metazoans. We have identified and characterized the Drosophila homologs of three subunits of the yeast Paf1 complex and found striking differences between the yeast and Drosophila Paf1 complexes. We demonstrate that although Drosophila Paf1, Rtf1, and Cdc73 colocalize broadly with actively transcribing, phosphorylated Pol II, and all are recruited to activated heat shock genes with similar kinetics; Rtf1 does not appear to be a stable part of the Drosophila Paf1 complex. RNA interference (RNAi)-mediated depletion of Paf1 or Rtf1 leads to defects in induction of Hsp70 RNA, but tandem RNAi-chromatin immunoprecipitation assays show that loss of neither Paf1 nor Rtf1 alters the density or distribution of phosphorylated Pol II on the active Hsp70 gene. However, depletion of Paf1 reduces trimethylation of histone H3 at lysine 4 in the Hsp70 promoter region and significantly decreases the recruitment of chromatin-associated factors Spt6 and FACT, suggesting that Paf1 may manifest its effects on transcription through modulating chromatin structure.

Drosophila Paf1 modulates chromatin structure at actively transcribed genes.

A relatively simple but powerful method to measure RNA polymerase II transcription elongation as well as co-transcriptional RNA splicing rates at many genes in vivo is described in this issue. The results demonstrate a rather uniform, and high, elongation rate at large human genes and co-transcriptional pre-mRNA splicing of both U2- and U12-dependent primary transcripts.

M. Behfar Ardehali

Papers

Defining mechanisms that regulate RNA polymerase II transcription in vivo

Drosophila Set1 is the major histone H3 lysine 4 trimethyltransferase with role in transcription

Spt6 enhances the elongation rate of RNA polymerase II in vivo

Drosophila Paf1 modulates chromatin structure at actively transcribed genes.

Tracking rates of transcription and splicing in vivo