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  • 2-NBDG Interestingly the C domain is also

    2019-07-24

    Interestingly, the C1 domain is also known to harbor a large number of pathogenic HCM-causing mutations [35], and most mutations implicated in HCM in the C1 domain are clustered towards the C-terminal end [31]. One such single amino 2-NBDG mutation located in the C terminal end of the C1 domain is a tyrosine to serine mutation at residue 237 in humans (residue 235 in mouse sequence) first identified in 2003 (mutations are abbreviated with one-letter amino acid code and its residue number, e.g. Y235S) [36]. This missense mutation is caused by an A to C nucleotide transversion in exon 6 of MYBPC3 [14], showing mild familial and strong disease association in humans [36]. The male human proband carrying the Y237S mutation was diagnosed at 37 years old and displayed cardiac hypertrophy, fairly reduced fractional shortening (FS), and left ventricular outflow tract (LVOT) obstruction [36]. A recent study identified an additional mutation of tyrosine 237 to a cysteine (Y237C), where in this case, the human proband was a 33 year old male with a family history of HCM presenting with an aortic murmur and severe hypertrophy [37]. Thus, Y237 mutations in humans cause clinically significant disease phenotypes. The tyrosine residue 237 is located inside the hydrophobic core of C1 domain, near the C-terminal end, where myosin and/or actin binding occurs. The molecular mechanism of how the Y237S mutation causes disease is unknown; however, based on the disease phenotypes of the Y237S mutation and the position of the tyrosine residue, we can infer that Y237 will have an important role in the development of HCM. Since Y237 participates in a web of hydrogen bonding, hydrophobic and ionic networks, a mutation at the site is expected to destabilize the local structure and possibly cause changes to be propagated across the whole C1 domain [38], thereby affecting the overall cMyBPC function and leading to organ level phenotypic changes described above. Additionally, Y237 is highly conserved across >25 species in the animal kingdom (Fig. 1) and across all MyBPC isoforms suggesting it is has the potential to be functionally important [39].
    Materials and methods
    Results
    Discussion Although cMyBPC mutations are a leading cause of HCM, the molecular mechanisms of its mutations are still unclear. Available data show that around 50% of MYBPC3 mutations result in truncated protein products [96,97], while the rest result in insertion/deletions, frameshifts, or single amino acid missense mutations [35]. It has been suggested that some truncated proteins may act as dominant negative poison poly-peptides [17], while other studies have suggested that haploinsufficiency is the main mechanism of disease [98]. In a clinical study of missense mutations of MYBPC3, the total protein expression was reduced, which suggested that some missense variants may increase the susceptibility to degradation through nonsense mediated mRNA decay, ubiquitin mediated proteolysis, and other pathways [99,100]. More recent studies have shown that MYBPC3 missense mutations produced full length mutant proteins; some seem to lead to haploinsufficiency [98,101,102], while others did not [103]. There is evidence that truncated proteins may cause disease directly through haploinsufficiency or indirectly by inducing cytotoxicity or disrupting myofibrillar architecture [98]. Therefore, the pathologic mechanism of missense mutations is thought to be heterogeneous based on the degree of structural stability and is unique to alterations specific to the mutation in the protein. In this study, we characterized the mechanism by which the pathogenic MYBPC3 Y235S missense mutation caused contractile dysfunction by performing mechanical experiments on skinned myocardium isolated from cMyBPC null mouse hearts following in vivo viral transfection of wild-type or Y235S MYBPC3. To probe the molecular mechanisms underlying mechanical dysfunction due to mutant Y235S cMyBPC expression, we performed in silico computational studies and MDS to determine its impact on C1 domain structure.