publication

Enhancer RNA profiling predicts transcription factor activity.

Anonymous — Thu, 15 Feb 2018 16:29:07 +0000

Enhancer RNA profiling predicts transcription factor activity. Anonymous (not verified) Thu, 02/15/2018 - 09:29

Genome Res. 2018 Feb 15. doi: 10.1101/gr.225755.117.

Azofeifa JG^1,², Allen MA², Hendrix JR^1,³, Read T^2,⁴, Rubin JD⁴, Dowell RD^1,^2,3

Abstract

Transcription factors (TFs) exert their regulatory influence through the binding of enhancers, resulting in coordination of gene expression programs. Active enhancers are often characterized by the presence of short, unstable transcripts termed enhancer RNAs (eRNAs). While their function remains unclear, we demonstrate that eRNAs are a powerful readout of TF activity. We infer sites of eRNA origination across hundreds of publicly available nascent transcription data sets and show that eRNAs initiate from sites of TF binding. By quantifying the colocalization of TF binding motif instances and eRNA origins, we derive a simple statistic capable of inferring TF activity. In doing so, we uncover dozens of previously unexplored links between diverse stimuli and the TFs they affect.

PMID:29449408DOI:10.1101/gr.225755.117

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An Annotation Agnostic Algorithm for Detecting Nascent RNA Transcripts in GRO-Seq.

Anonymous — Fri, 01 Sep 2017 06:00:00 +0000

An Annotation Agnostic Algorithm for Detecting Nascent RNA Transcripts in GRO-Seq. Anonymous (not verified) Fri, 09/01/2017 - 00:00

IEEE/ACM Trans Comput Biol Bioinform. 2017 Sep-Oct;14(5):1070-1081. doi: 10.1109/TCBB.2016.2520919. Epub 2016 Jan 26.

An Annotation Agnostic Algorithm for Detecting Nascent RNA Transcripts in GRO-Seq.

Azofeifa JG, Allen MA, Lladser ME, Dowell RD.

Abstract

We present a fast and simple algorithm to detect nascent RNA transcription in global nuclear run-on sequencing (GRO-seq). GRO-seq is a relatively new protocol that captures nascent transcripts from actively engaged polymerase, providing a direct read-out on bona fide transcription. Most traditional assays, such as RNA-seq, measure steady state RNA levels which are affected by transcription, post-transcriptional processing, and RNA stability. GRO-seq data, however, presents unique analysis challenges that are only beginning to be addressed. Here, we describe a new algorithm, Fast Read Stitcher (FStitch), that takes advantage of two popular machine-learning techniques, hidden Markov models and logistic regression, to classify which regions of the genome are transcribed. Given a small user-defined training set, our algorithm is accurate, robust to varying read depth, annotation agnostic, and fast. Analysis of GRO-seq data without a priori need for annotation uncovers surprising new insights into several aspects of the transcription process.

PMID:26829802PMCID:PMC5667649[Available on 2018-09-01]DOI:10.1109/TCBB.2016.2520919

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Human TFIIH Kinase CDK7 Regulates Transcription-Associated Chromatin Modifications.

Anonymous — Tue, 01 Aug 2017 06:00:00 +0000

Human TFIIH Kinase CDK7 Regulates Transcription-Associated Chromatin Modifications. Anonymous (not verified) Tue, 08/01/2017 - 00:00

Cell Rep. 2017 Aug 1;20(5):1173-1186. doi: 10.1016/j.celrep.2017.07.021.

Human TFIIH Kinase CDK7 Regulates Transcription-Associated Chromatin Modifications.

Ebmeier CC¹, Erickson B², Allen BL³, Allen MA⁴, Kim H², Fong N², Jacobsen JR⁵, Liang K⁶, Shilatifard A⁶, Dowell RD⁷, Old WM⁸, Bentley DL⁹, Taatjes DJ¹⁰.

Author information

Abstract

CDK7 phosphorylates the RNA polymerase II (pol II) C-terminal domain CTD and activates the P-TEFb-associated kinase CDK9, but its regulatory roles remain obscure. Here, using human CDK7 analog-sensitive (CDK7as) cells, we observed reduced capping enzyme recruitment, increased pol II promoter-proximal pausing, and defective termination at gene 3' ends upon CDK7 inhibition. We also noted that CDK7 regulates chromatin modifications downstream of transcription start sites. H3K4me3 spreading was restricted at gene 5' ends and H3K36me3 was displaced toward gene 3' ends in CDK7as cells. Mass spectrometry identified factors that bound TFIIH-phosphorylated versus P-TEFb-phosphorylated CTD (versus unmodified); capping enzymes and H3K4 methyltransferase complexes, SETD1A/B, selectively bound phosphorylated CTD, and the H3K36 methyltransferase SETD2 specifically bound P-TEFb-phosphorylated CTD. Moreover, TFIIH-phosphorylated CTD stimulated SETD1A/B activity toward nucleosomes, revealing a mechanistic basis for CDK7 regulation of H3K4me3 spreading. Collectively, these results implicate a CDK7-dependent "CTD code" that regulates chromatin marks in addition to RNA processing and pol II pausing.

KEYWORDS:

H3K36me3; H3K4me3; Mediator; P-TEFb; RNA-seq; TFIIH; THZ1; chromatin; epigenetic; proteomics

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RNA Pol II transcription model and interpretation of GRO-seq data.

Anonymous — Sun, 01 Jan 2017 07:00:00 +0000

RNA Pol II transcription model and interpretation of GRO-seq data. Anonymous (not verified) Sun, 01/01/2017 - 00:00

J Math Biol. 2017 Jan;74(1-2):77-97. doi: 10.1007/s00285-016-1014-4. Epub 2016 May 3.

RNA Pol II transcription model and interpretation of GRO-seq data.

Lladser ME¹, Azofeifa JG², Allen MA³, Dowell RD⁴.

Author information

Abstract

A mixture model and statistical method is proposed to interpret the distribution of reads from a nascent transcriptional assay, such as global run-on sequencing (GRO-seq) data. The model is annotation agnostic and leverages on current understanding of the behavior of RNA polymerase II. Briefly, it assumes that polymerase loads at key positions (transcription start sites) within the genome. Once loaded, polymerase either remains in the initiation form (with some probability) or transitions into an elongating form (with the remaining probability). The model can be fit genome-wide, allowing patterns of Pol II behavior to be assessed on each distinct transcript. Furthermore, it allows for the first time a principled approach to distinguishing the initiation signal from the elongation signal; in particular, it implies a data driven method for calculating the pausing index, a commonly used metric that informs on the behavior of RNA polymerase II. We demonstrate that this approach improves on existing analyses of GRO-seq data and uncovers a novel biological understanding of the impact of knocking down the Male Specific Lethal (MSL) complex in Drosophilia melanogaster.

KEYWORDS:

Double Geometric distribution; Elongation; GRO-seq; Gene; Initiation; Pausing index; RNA polymerase

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FStitch: A fast and simple algorithm for detecting nascent RNA transcripts.

Anonymous — Mon, 01 Sep 2014 06:00:00 +0000

FStitch: A fast and simple algorithm for detecting nascent RNA transcripts. Anonymous (not verified) Mon, 09/01/2014 - 00:00

FStitch: A fast and simple algorithm for detecting nascent RNA transcripts.

Azofiefa J, Allen M, Lladser M, Dowell R.

Proceedings of the 5th ACM Conference on Bioinformatics, Computational Biology, and Health Informatics (ACM-BCB ’14). 2014 September;

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Global analysis of p53-regulated transcription identifies its direct targets and unexpected regulatory mechanisms.

Anonymous — Tue, 27 May 2014 06:00:00 +0000

Global analysis of p53-regulated transcription identifies its direct targets and unexpected regulatory mechanisms. Anonymous (not verified) Tue, 05/27/2014 - 00:00

Elife. 2014 May 27;3:e02200. doi: 10.7554/eLife.02200.

Global analysis of p53-regulated transcription identifies its direct targets and unexpected regulatory mechanisms.

Allen MA¹, Andrysik Z², Dengler VL², Mellert HS², Guarnieri A², Freeman JA², Sullivan KD², Galbraith MD², Luo X³, Kraus WL³, Dowell RD⁴, Espinosa JM⁵.

Author information

Abstract

The p53 transcription factor is a potent suppressor of tumor growth. We report here an analysis of its direct transcriptional program using Global Run-On sequencing (GRO-seq). Shortly after MDM2 inhibition by Nutlin-3, low levels of p53 rapidly activate ∼200 genes, most of them not previously established as direct targets. This immediate response involves all canonical p53 effector pathways, including apoptosis. Comparative global analysis of RNA synthesis vs steady state levels revealed that microarray profiling fails to identify low abundance transcripts directly activated by p53. Interestingly, p53 represses a subset of its activation targets before MDM2 inhibition. GRO-seq uncovered a plethora of gene-specific regulatory features affecting key survival and apoptotic genes within the p53 network. p53 regulates hundreds of enhancer-derived RNAs. Strikingly, direct p53 targets harbor pre-activated enhancers highly transcribed in p53 null cells. Altogether, these results enable the study of many uncharacterized p53 target genes and unexpected regulatory mechanisms.DOI: http://dx.doi.org/10.7554/eLife.02200.001.

KEYWORDS:

PIG3; PUMA; eRNA; genomics; p21; tumor supressor

PMID:24867637PMCID:PMC4033189

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Retrospective reflections of a whistleblower: opinions on misconduct responses.

Anonymous — Thu, 12 Sep 2013 06:00:00 +0000

Retrospective reflections of a whistleblower: opinions on misconduct responses. Anonymous (not verified) Thu, 09/12/2013 - 00:00

Account Res. 2013;20(5-6):339-48. doi: 10.1080/08989621.2013.822249.

Retrospective reflections of a whistleblower: opinions on misconduct responses.

Allen M¹, Dowell R.

Author information

Abstract

Almost 10 years ago, when I was in my fourth year of graduate school, my fellow graduate students discovered that our thesis advisor had engaged in misconduct by falsifying and fabricating data in two grant applications. We informed the university and my advisor resigned. This event was a turning point in my life. Years later, I have gathered my thoughts and reflections on the experience. I believe we must first prevent what misconduct we can. But unfortunately some misconduct will still occur and in those circumstances we must respond to protect those affected by the misconduct and to progress beyond the event. In so doing, we get the most value out of scientific research.

PMID:24028481DOI:10.1080/08989621.2013.822249

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HIF1A employs CDK8-mediator to stimulate RNAPII elongation in response to hypoxia.

Anonymous — Thu, 06 Jun 2013 06:00:00 +0000

HIF1A employs CDK8-mediator to stimulate RNAPII elongation in response to hypoxia. Anonymous (not verified) Thu, 06/06/2013 - 00:00

Cell. 2013 Jun 6;153(6):1327-39. doi: 10.1016/j.cell.2013.04.048.

HIF1A employs CDK8-mediator to stimulate RNAPII elongation in response to hypoxia.

Galbraith MD¹, Allen MA, Bensard CL, Wang X, Schwinn MK, Qin B, Long HW, Daniels DL, Hahn WC, Dowell RD, Espinosa JM.

Author information

Abstract

The transcription factor HIF1A is a key mediator of the cellular response to hypoxia. Despite the importance of HIF1A in homeostasis and various pathologies, little is known about how it regulates RNA polymerase II (RNAPII). We report here that HIF1A employs a specific variant of the Mediator complex to stimulate RNAPII elongation. The Mediator-associated kinase CDK8, but not the paralog CDK19, is required for induction of many HIF1A target genes. HIF1A induces binding of CDK8-Mediator and the super elongation complex (SEC), containing AFF4 and CDK9, to alleviate RNAPII pausing. CDK8 is dispensable for HIF1A chromatin binding and histone acetylation, but it is essential for binding of SEC and RNAPII elongation. Global analysis of active RNAPII reveals that hypoxia-inducible genes are paused and active prior to their induction. Our results provide a mechanistic link between HIF1A and CDK8, two potent oncogenes, in the cellular response to hypoxia.

PMID:23746844PMCID:PMC3681429DOI:10.1016/j.cell.2013.04.048

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Basal cell carcinomas in mice arise from hair follicle stem cells and multiple epithelial progenitor populations.

Anonymous — Sun, 01 May 2011 06:00:00 +0000

Basal cell carcinomas in mice arise from hair follicle stem cells and multiple epithelial progenitor populations. Anonymous (not verified) Sun, 05/01/2011 - 00:00

J Clin Invest. 2011 May;121(5):1768-81. doi: 10.1172/JCI46307. Epub 2011 Apr 25.

Basal cell carcinomas in mice arise from hair follicle stem cells and multiple epithelial progenitor populations.

Grachtchouk M¹, Pero J, Yang SH, Ermilov AN, Michael LE, Wang A, Wilbert D, Patel RM, Ferris J, Diener J, Allen M, Lim S, Syu LJ, Verhaegen M, Dlugosz AA.

Author information

Abstract

Uncontrolled Hedgehog (Hh) signaling leads to the development of basal cell carcinoma (BCC), the most common human cancer, but the cell of origin for BCC is unclear. While Hh pathway dysregulation is common to essentially all BCCs, there exist multiple histological subtypes, including superficial and nodular variants, raising the possibility that morphologically distinct BCCs may arise from different cellular compartments in skin. Here we have shown that induction of a major mediator of Hh signaling, GLI2 activator (GLI2ΔN), selectively in stem cells of resting hair follicles in mice, induced nodular BCC development from a small subset of cells in the lower bulge and secondary hair germ compartments. Tumorigenesis was markedly accelerated when GLI2ΔN was induced in growing hair follicles. In contrast, induction of GLI2ΔN in epidermis led to the formation of superficial BCCs. Expression of GLI2ΔN at reduced levels in mice yielded lesions resembling basaloid follicular hamartomas, which have previously been linked to low-level Hh signaling in both mice and humans. Our data show that the cell of origin, tissue context (quiescent versus growing hair follicles), and level of oncogenic signaling can determine the phenotype of Hh/Gli-driven skin tumors, with high-level signaling required for development of superficial BCC-like tumors from interfollicular epidermis and nodular BCC-like tumors from hair follicle stem cells.

Comment in

Mommy - where do tumors come from? [J Clin Invest. 2011]

PMID:21519145PMCID:PMC3083781DOI:10.1172/JCI46307

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A global analysis of C. elegans trans-splicing.

Anonymous — Mon, 21 Feb 2011 07:00:00 +0000

A global analysis of C. elegans trans-splicing. Anonymous (not verified) Mon, 02/21/2011 - 00:00

Allen MA, Hillier LW, Waterston RH, Blumenthal T.

Genome research. 2011; 21(2):255-64.

PMID:21177958PMCID:PMC3032929

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