Wednesday, May 8, 2024 5/8/2024

Studies to Explore DNA Replication Proteins in Functional Assemblies through Intrinsically Disordered Domains

Award Number: R01GM141045
ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

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PROJECT SUMMARY Our long-term goal is to define the molecular mechanisms by which DNA replication is initiated and regulated in metazoans. Cells rely on two AAA+ ATPases, ORC and Cdc6, along with a third factor, Cdt1, to load a latent helicase (the Mcm2-7 complex) as a double hexamer onto replication origins. Upon entering S-phase, Mcm2-7 is activated by the GINS and Cdc45 accessory factors, melting the duplex origin. The resultant CMG (Cdc45/Mcm2-7/GINS) assembly unwinds parental DNA strands and coordinates DNA synthesis by the replisome. Recently, we discovered that metazoan replication initiation factors – specifically the Orc1 subunit of ORC, as well as Cdc6 and Cdt1 – use long, intrinsically disordered regions (IDRs) to bind DNA and partition into liquid phase condensates (LPCs). This and other observations led us to a new functional model for replication, whereby initiator IDRs and LPC propensity controls not only chromatin association, but also Mcm2-7 loading, partner selection, and heterochromatin status. In Aim 1, we will resolve the molecular determinants by which initiator IDRs facilitate condensation. In Aim 2, we will define how initiator IDRs control partner-protein interactions. In Aim 3, we will establish how Orc1 uses its IDR to interface with pericentric heterochromatin through interactions with other LPC-forming proteins such as Hp1. Significant outcomes expected to result from the proposed work include: 1) defining how initiator IDRs – which we have shown to be a novel class of condensate-promoting element – interface with DNA and each other, 2) uncovering new proteins capable of associating with initiation factors, and 3) explaining how ORC connects to the formation and maintenance of genome organization and expression. IDRs have been predominantly thought to serve either as flexible linkers that allow mobility between ordered domains, or as segments that undergo an induced-fit transition into folded structures through protein-protein interactions. Recent work shows that IDRs can fulfill another role in specifying partner-protein interactions and driving the formation of membraneless compartments through liquid phase separation. Our proposal will establish how IDRs can lead to specificity for co-association and potential compartmentalization with origins and other factors. Our efforts will inform areas of molecular biology where IDRs are used to manifest phase-separated compartments or protein/nucleic-acid clustering for functional purposes. As ~25% of proteomes are predicted to be unstructured, the utility of such insights will be broadly significant.


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 = $484,052 )
2024	2024	REGENTS OF THE UNIVERSITY OF CALIFORNIA, THE	1608 4TH ST STE 201	BERKELEY	CA	94710	ALAMEDA	USA	Biomedical Research and Research Training	000	4	2/7/2024	NON-COMPETING CONTINUATION	$484,052
														Subtotal = $484,052

Issue Date FY: 2023 ( Subtotal = $537,835 )
2023	2023	REGENTS OF THE UNIVERSITY OF CALIFORNIA, THE	1608 4TH ST STE 201	BERKELEY	CA	94710	ALAMEDA	USA	Biomedical Research and Research Training	000	3	1/25/2023	NON-COMPETING CONTINUATION	$484,052
2023	2023	REGENTS OF THE UNIVERSITY OF CALIFORNIA, THE	1608 4TH ST STE 201	BERKELEY	CA	94710	ALAMEDA	USA	Biomedical Research and Research Training	001	3	3/28/2023	NON-COMPETING CONTINUATION	$53,783
														Subtotal = $537,835

Issue Date FY: 2022 ( Subtotal = $593,935 )
2022	2022	REGENTS OF THE UNIVERSITY OF CALIFORNIA, THE	1608 4TH ST STE 201	BERKELEY	CA	94710	ALAMEDA	USA	Biomedical Research and Research Training	002	2	9/20/2022	SUPPLEMENT FOR EXPANSION	$56,101
2022	2022	REGENTS OF THE UNIVERSITY OF CALIFORNIA, THE	1608 4TH ST STE 201	BERKELEY	CA	94710	ALAMEDA	USA	Biomedical Research and Research Training	001	2	8/3/2022	NON-COMPETING CONTINUATION	$53,783
2022	2022	REGENTS OF THE UNIVERSITY OF CALIFORNIA, THE	1608 4TH ST STE 201	BERKELEY	CA	94710	ALAMEDA	USA	Biomedical Research and Research Training	000	2	2/17/2022	NON-COMPETING CONTINUATION	$484,051
														Subtotal = $593,935

Issue Date FY: 2021 ( Subtotal = $552,529 )
2021	2021	REGENTS OF THE UNIVERSITY OF CALIFORNIA, THE	1608 4TH ST STE 220	BERKELEY	CA	94710	ALAMEDA	USA	Biomedical Research and Research Training	000	1	4/30/2021	NEW	$552,529
														Subtotal = $552,529

Grand Total All Awards = $2,168,351

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Studies to Explore DNA Replication Proteins in Functional Assemblies through Intrinsically Disordered Domains

Award Number: R01GM141045

ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

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

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