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Uncovering novel roles for splicing factor SF3B1 in transcription dynamics, R-loop metabolism, and chromatin structure

Award Number: F31GM136143
ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: FELLOWSHIP/SCHOLARSHIP/STUDENT LOANS

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Project Summary R-loops are non-B DNA structures that form during transcription when the nascent RNA strand anneals to the template DNA strand forming a RNA:DNA hybrid. The Chedin lab has demonstrated that R-loops are prevalent and conserved structures that form throughout the human genome. Understanding the function of R-loops under physiological and pathological conditions is an important goal in the field, because mis-regulation of R-loops has been implicated in a growing number of human disorders. A leading mechanism in the field is that splicing inhibition causes an increase in unspliced nascent transcripts that can then more readily invade the DNA behind the advancing RNA polymerase. To uncover the connections between splicing disruption and R-loops, I will focus on SF3B1, a subunit in the SF3b complex which plays a critical role in the early stages of spliceosome assembly. Importantly, Pladienolide B (PladB) is a natural product that directly inhibits splicing upon SF3b binding. Thus, PladB provides a tool for assessing dynamic changes in a temporal manner. In keeping with available literature, my initial hypothesis was that PladB treatment will lead to elevated R-loop formation over regions that accumulate unspliced transcripts. Preliminary data, however, is inconsistent with this idea and instead suggests that most R-loop changes that accompany SF3b inhibition are caused by perturbation of transcriptional dynamics. Early termination events cause directional R-loop losses through gene bodies. Lack of termination at gene ends, by contrast, cause “downstream of gene (DoG)” transcription and directional R-loop gains over DoG regions. Both events collectively affect over a thousand genes. DoG transcription has been observed in response to several environmental stresses. This raises the possibility that splicing inhibition is a shared molecular link that drives DoG transcription. DoG transcription upon viral infection has been further linked to large scale chromatin opening throughout the DoG region. This raises the possibility that R-loops, which include a rigid A-form-like RNA:DNA hybrid, cause chromatin decondensation by preventing nucleosome wrapping or deposition. Thus, my revised hypothesis is that acute splicing inhibition affects transcription elongation profiles and leads to shifts in the genomic patterns of co-transcriptional R-loops. Aim 1 will determine the global dynamic effects of acute splicing inhibition on splicing, R-loop and transcription patterns. I expect to clarify the temporal and positional relationships between splicing inhibition and R-loop formation at high-resolution and to identify a novel role for SF3b in regulating transcription dynamics. Aim 2 will determine if R-loops generated from DoG transcription drive changes in chromatin architecture under different cellular stresses. This project will provide key insights into the inter-relationship between co-transcriptional splicing and R-loop formation and their impact on transcriptional dynamics and chromatin architecture under stress conditions.


Issue Date FY	Funding FY	Legal Entity Name	Legal Entity Address	Legal Entity City	Legal Entity State	Legal Entity Zip Code	Legal Entity COUNTY	Legal Entity COUNTRY	Assistance Listing	Award Code	Budget Year	Action Date	Action Type	Action Amount

Issue Date FY: 2023 ( Subtotal = -$473 )
2023	2021	UNIVERSITY OF CALIFORNIA, DAVIS	1850 RESEARCH PARK DR, STE 300	DAVIS	CA	95618	YOLO	USA	Biomedical Research and Research Training	000	2	5/15/2023	NON-COMPETING CONTINUATION	-$473
														Subtotal = -$473

Issue Date FY: 2021 ( Subtotal = $38,422 )
2021	2021	UNIVERSITY OF CALIFORNIA, DAVIS	1850 RESEARCH PARK DR, STE 300	DAVIS	CA	95618	YOLO	USA	Biomedical Research and Research Training	000	2	3/8/2021	NON-COMPETING CONTINUATION	$38,422
														Subtotal = $38,422

Issue Date FY: 2020 ( Subtotal = $37,906 )
2020	2020	UNIVERSITY OF CALIFORNIA, DAVIS	1850 RESEARCH PARK DR, STE 300	DAVIS	CA	95618	YOLO	USA	Biomedical Research and Research Training	002	1	2/19/2020	NEW	$0
2020	2020	UNIVERSITY OF CALIFORNIA, DAVIS	1850 RESEARCH PARK DR, STE 300	DAVIS	CA	95618	YOLO	USA	Biomedical Research and Research Training	001	1	2/19/2020	NEW	$504
2020	2020	UNIVERSITY OF CALIFORNIA, DAVIS	1850 RESEARCH PARK DR, STE 300	DAVIS	CA	95618	YOLO	USA	Biomedical Research and Research Training	000	1	1/14/2020	NEW	$37,402
														Subtotal = $37,906

Grand Total All Awards = $75,855

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Uncovering novel roles for splicing factor SF3B1 in transcription dynamics, R-loop metabolism, and chromatin structure

Award Number: F31GM136143

ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

AWARD ACTIVITY TYPE: FELLOWSHIP/SCHOLARSHIP/STUDENT LOANS

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