Abstract 4366180: Sorbs2 Deficiency-induced Cardiac MyBP-C Downregulation Underlies Atrial Fibrotic Cardiomyopathy in Mice

Background: Cardiac myosin-binding protein C (cMyBP-C, encoded by MYBPC3 ) and Sorbin and SH3 domain-containing protein 2 (Sorbs2, encoded by SORBS2 ) are critical sarcomeric components that maintain cardiac structure and function. MYBPC3 and SORBS2 are linked to hypertrophic and dilated cardiomyopathies. But the relationship between Sorbs2 and cMyBP-C regulation, and their roles in the pathogenesis of atrial fibrotic cardiomyopathy (AFCM), remains unclear. Hypothesis: Sorbs2 acts as a cytoskeletal and RNA-binding protein that regulates cMyBP-C expression in atrial myocardium. Methods: We created the cardiac-specific Sorbs2 knockout ( c-Sorbs2 KO mice. We used next-generation RNA sequencing, molecular biology techniques, and in silico analysis to study how Sorbs2 regulates cMyBP-C expression in the atria of c-Sorbs2 KO mice. Results: Compared to age-matched wild-type (WT) mice at 8-9 months of age, the c-Sorbs2 KO male exhibited significant left atrial (LA) enlargement (diameter: 2.41±0.11 mm vs 1.82 ± 0.05 mm in WT; n = 8; p < 0.001) and severe atrial fibrosis (fibrotic area: 12.29±1.73 vs 2.83 ± 0.41% in WT; p < 0.001), accompanied by marked reductions in both mRNA and protein levels of cMyBP-C in LA. In vitro knock-down of Sorbs2 by 60% via shRNA led to an 86.3% decrease in cMyBP-C protein levels, while a 3-fold Sorbs2 overexpression via adenoviral transduction resulted in a 1.4-fold increase in cMyBP-C protein levels in HL-1 cells . cMyBP-C mRNAs were identified in the ribonucleoprotein complexes immunoprecipitated with anti-Sorbs2 antibodies from cardiomyocyte lysates. Loss of Sorbs2 significantly reduced the half-life of cMyBP-C mRNA (from 2.1 hrs in WT to 1.2 hrs in KO). Co-immunoprecipitation with truncated Sorbs2 constructs showed that the C-terminal SH3 domains of Sorbs2, but not the N-terminal SoHo domain, interacted with cMyBP-C protein. Molecular docking and modeling showed hydrophobic interactions and hydrogen bonds between residues H906 and G907 in the SH3 domain of Sorbs2 and residue A954, P955 and T957 in the SH3-binding motif (949-MA P GA P VT-957) of cMyBP-C with a weighted score of -1036 using ClusPro online service. Conclusions: Sorbs2 regulates cMyBP-C expression through direct interaction with both cMyBP-C mRNA and protein. Loss of Sorbs2 results in reduced cMyBP-C expression, atrial fibrosis, and chamber enlargement, revealing a novel Sorbs2/cMyBP-C axis underlying AFCM pathogenesis.