Tuesday, July 1, 2025 7/1/2025

Skeletal Myosin-Binding Protein C Regulation and Structural Dynamics

Award Number: R01AR079435
ORGANIZATION: NATIONAL INSTITUTE OF ARTHRITIS & MUSCULOSKELETAL & SKIN DISEASES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)
PERIOD OF PERFORMANCE START DATE: 07/15/2022
PERIOD OF PERFORMANCE END DATE: 05/31/2027

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Skeletal Myosin-Binding Protein C Regulation and Structural Dynamics - PROJECT SUMMARY Arthrogryposis is present in 1 in 3,000 live births causing joint contractures in both upper and lower limbs. There is no cure making it an unmet medical need. Mutations in the MYBPC1 gene encoding slow skeletal myosin-binding protein C (sMyBP-C), expressed in both slow and fast muscle types are associated with distal arthrogryposis (DA). MYBPC2 encodes for fast skeletal MyBP-C (fMyBP-C) and is found only in fast- twitch muscle. As a myosin-anchored protein of muscle, MyBP-C extends toward actin, positioned centrally in the sarcomere to regulate actomyosin interactions in force development. MyBP-C in skeletal muscle has three major regulators: isoform (slow vs. fast), splice variant (long vs. short sMyBP-C), and posttranslational modification (phosphorylation). sMyBP-C is phosphorylated by protein kinase A (PKA) at its N terminus. The role(s) of sMyBP-C, its phosphorylation and DA mutations in skeletal muscle are not known. Our preliminary studies of sMyBP-C show that binding to actomyosin is dependent on phosphorylation and DA mutations. We have developed innovative biophysical tools that enable evaluation of skeletal MyBP-C structural dynamics, actomyosin interactions in muscle, and effects of phosphorylation and mutations. Our new preliminary studies demonstrate that we have successfully developed fluorescent sensors in N terminal sMyBP-C whose structure and dynamics are sensitive to PKA-mediated phosphorylation and binding to actin. We have also developed inter-molecular fluorescence assays that resolve actin binding between fMyBP-C, long sMyBP-C, and short MyBP-C due to phosphorylation and the presence of tropomyosin on actin. These preliminary results suggest key physiological mechanisms of regulation for the different skeletal MyBP-C and provides additional scientific premise and feasibility for pursuing the proposed studies. Aim 1 will evaluate effects of sMyBP-C binding and DA mutations on interactions with actomyosin, capturing structure and proximities of N terminal sMyBP-C, actin and myosin. Spectroscopic probes will be placed in these proteins and approaches will be employed to detect key conformations in vitro and in situ with wild type and DA mutant sMyBP-C. For fiber experiments, muscle will be isolated from novel sMyBP-C knockout (KO) mice and permeabilized with recombinant sMyBP- C, DA mutants, and muscle protein probes. Samples will be assessed for binding and contractile function. Aim 2 will determine how PKA-mediated phosphorylation of sMyBP-C affects the parameters evaluated in Aim 1. Aim 3 will determine how fMyBP-C affects the parameters evaluated in Aim 1 except using fMyBP-C KO and sMyBP-C/fMyBP-C double-KO mice for fibers experiments. The proposed studies capture structural dynamics and interactions in real time and myofilament space using novel high-resolution approaches. These aims outline a stepwise plan for studying normal and mutant skeletal MyBP-C during the contractile cycle. By monitoring distances between points on proteins and the order (or disorder) of those distances under physiological conditions, mutants can be separated into bins to facilitate targeted mechanistic-based therapies.


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: 2025 ( Subtotal = $425,739 )
2025	2025	UNIVERSITY OF ARIZONA	845 N PARK AVE RM 538	TUCSON	AZ	85721	PIMA	USA	Arthritis, Musculoskeletal and Skin Diseases Research	001	4	6/4/2025	NON-COMPETING CONTINUATION	$425,739
2025	2024	UNIVERSITY OF ARIZONA	845 N PARK AVE RM 538	TUCSON	AZ	85721	PIMA	USA	Arthritis, Musculoskeletal and Skin Diseases Research	000	3	4/30/2025	SUPPLEMENT FOR EXPANSION	$0
														Subtotal = $425,739

Issue Date FY: 2024 ( Subtotal = $496,910 )
2024	2024	UNIVERSITY OF ARIZONA	845 N PARK AVE RM 538	TUCSON	AZ	85721	PIMA	USA	Arthritis, Musculoskeletal and Skin Diseases Research	001	3	6/6/2024	SUPPLEMENT FOR EXPANSION	$69,778
2024	2024	UNIVERSITY OF ARIZONA	845 N PARK AVE RM 538	TUCSON	AZ	85721	PIMA	USA	Arthritis, Musculoskeletal and Skin Diseases Research	000	3	6/5/2024	NON-COMPETING CONTINUATION	$427,132
														Subtotal = $496,910

Issue Date FY: 2023 ( Subtotal = $494,596 )
2023	2023	UNIVERSITY OF ARIZONA	845 N PARK AVE RM 538	TUCSON	AZ	85721	PIMA	USA	Arthritis, Musculoskeletal and Skin Diseases Research	001	2	9/6/2023	SUPPLEMENT FOR EXPANSION	$52,180
2023	2023	UNIVERSITY OF ARIZONA	845 N PARK AVE RM 538	TUCSON	AZ	85721	PIMA	USA	Arthritis, Musculoskeletal and Skin Diseases Research	000	2	6/16/2023	NON-COMPETING CONTINUATION	$442,416
														Subtotal = $494,596

Issue Date FY: 2022 ( Subtotal = $456,984 )
2022	2022	UNIVERSITY OF ARIZONA	888 N EUCLID AVE RM 510	TUCSON	AZ	85719	PIMA	USA	Arthritis, Musculoskeletal and Skin Diseases Research	000	1	7/15/2022	NEW	$456,984
														Subtotal = $456,984

Grand Total All Awards = $1,874,229

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Skeletal Myosin-Binding Protein C Regulation and Structural Dynamics

Award Number: R01AR079435

ORGANIZATION: NATIONAL INSTITUTE OF ARTHRITIS & MUSCULOSKELETAL & SKIN DISEASES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

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

PERIOD OF PERFORMANCE START DATE: 07/15/2022

PERIOD OF PERFORMANCE END DATE: 05/31/2027

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