Saturday, September 7, 2024 9/7/2024

Leveraging genetic variation to dissect gene regulatory networks of reprogramming to pluripotency

Award Number: U01HG012079
ORGANIZATION: NATIONAL HUMAN GENOME RESEARCH INSTITUTE
OPDIV: NIH
AWARD CLASS: COOPERATIVE AGREEMENT
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

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PROJECT SUMMARY The reprogramming of somatic human cells to induced pluripotent stem cells (iPSCs) by only four transcription factors (TFs) Oct4, Sox2, Klf4, and cMyc (OSKM) is one of the most striking remodelings of gene regulatory networks. The remarkable ability of OSKM to reprogram diverse somatic cell types into iPSCs that are functionally indistinguishable from embryonic stem cells indicates that OSKM leverages a fundamental mechanism for network remodeling that may be generally applicable to all cell fate transitions. Previous studies of reprogramming have identified the crucial role of cooperative TF binding in repressing somatic programs and activating pluripotent ones. However, associating TF binding dynamics and epigenomic remodeling with key bifurcation events during reprogramming is confounded by the highly heterogeneous nature of the reprogramming process and the lack of knowledge regarding how the transition from somatic to pluripotent regulatory programs occurs in individual cells. In this project, we aim to model the regulatory network underlying the cell fate change of reprogramming using three types of single-cell multi-omic profiles generated from critical time points during reprogramming. We will interrogate the network leveraging natural perturbation of reprogramming and pluripotency by genetic variants. Genetic variation is well known to modulate the regulatory network of pluripotency and contributes to the variability of cellular phenotypes and differentiation capacity of iPSC lines. We will generate population-scale single-cell joint profiling of RNA and DNA methylation (snmCT- seq), joint profiling of RNA and chromatin accessibility (scRNA + ATAC-seq) and single-nucleus joint profiling of chromatin conformation and DNA methylation (sn-m3C-seq), allowing the cell-type-specific determination of transcriptome, chromatin accessibility and methylation states at regulatory elements, as well as enhancer-gene looping to connect non-coding variants to their regulatory target. To integrate OSKM binding with the single-cell transcriptomic and epigenomic dynamics, we will determine the allele-specific binding of TFs and histone modifications using a pooled-alleles ChIP-seq strategy. We will use Dynamic Regulatory Events Miner (DREM) to construct predictive models by integrating transcription factor-gene interaction information with time- and pseudotime-series genomics data. To determine the genetic regulation of the reprogramming network, we will apply the novel statistical method FastGxE to distinguish cell-type-specific from the shared genetic component of gene expression regulation, to enhance the sensitivity for identifying cell-type-specific quantitative trait loci (QTLs). To test the regulatory network, we will experimentally determine the function of network hub genes and non-coding variants using high-throughput CRISPR interference and precise variant replacement experiments. Our proposed project integrates diverse approaches including single-cell multi-omics, computational modeling, and genetic engineering, and will likely provide new insights into the mechanism by which TFs remodel regulatory networks of cell type identity and serve as a model for similar analyses in other systems.


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: 2024 ( Subtotal = $931,002 )
2024	2024	UNIVERSITY OF CALIFORNIA, LOS ANGELES	10889 WILSHIRE BLVD STE 700	LOS ANGELES	CA	90024	LOS ANGELES	USA	Human Genome Research	000	4	7/10/2024	NON-COMPETING CONTINUATION	$931,002
														Subtotal = $931,002

Issue Date FY: 2023 ( Subtotal = $1,351,290 )
2023	2023	UNIVERSITY OF CALIFORNIA, LOS ANGELES	10889 WILSHIRE BLVD STE 700	LOS ANGELES	CA	90024	LOS ANGELES	USA	Human Genome Research	001	3	9/22/2023	NON-COMPETING CONTINUATION	$401,290
2023	2023	UNIVERSITY OF CALIFORNIA, LOS ANGELES	10889 WILSHIRE BLVD STE 700	LOS ANGELES	CA	90024	LOS ANGELES	USA	Human Genome Research	000	3	6/23/2023	NON-COMPETING CONTINUATION	$950,000
														Subtotal = $1,351,290

Issue Date FY: 2022 ( Subtotal = $1,351,290 )
2022	2022	UNIVERSITY OF CALIFORNIA, LOS ANGELES	10889 WILSHIRE BLVD STE 700	LOS ANGELES	CA	90024	LOS ANGELES	USA	Human Genome Research	001	2	9/21/2022	NON-COMPETING CONTINUATION	$401,290
2022	2022	UNIVERSITY OF CALIFORNIA, LOS ANGELES	10889 WILSHIRE BLVD STE 700	LOS ANGELES	CA	90024	LOS ANGELES	USA	Human Genome Research	000	2	7/5/2022	NON-COMPETING CONTINUATION	$950,000
														Subtotal = $1,351,290

Issue Date FY: 2021 ( Subtotal = $1,351,290 )
2021	2021	UNIVERSITY OF CALIFORNIA, LOS ANGELES	10889 WILSHIRE BLVD STE 700	LOS ANGELES	CA	90024	LOS ANGELES	USA	Human Genome Research	001	1	9/21/2021	NEW	$401,290
2021	2021	UNIVERSITY OF CALIFORNIA, LOS ANGELES	10889 WILSHIRE BLVD STE 700	LOS ANGELES	CA	90024	LOS ANGELES	USA	Human Genome Research	000	1	8/23/2021	NEW	$950,000
														Subtotal = $1,351,290

Grand Total All Awards = $4,984,872

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Leveraging genetic variation to dissect gene regulatory networks of reprogramming to pluripotency

Award Number: U01HG012079

ORGANIZATION: NATIONAL HUMAN GENOME RESEARCH INSTITUTE

OPDIV: NIH

AWARD CLASS: COOPERATIVE AGREEMENT

AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

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