Abstract 191: Functional Dissection of Myosin Binding Protein-C Phosphorylation
Rationale: Phosphorylation of cardiac myosin binding protein C (cMyBP-C) plays an important role in cardiac function and is actively regulated both in the normal heart and during the development of cardiac disease. Hence it is critical to understand the mechanistic and functional consequences of cMyBP-C phosphorylation
Objective: To examine the non-equivalency of the group of three phosphorylatable sites (Ser-273, Ser-282, and Ser-302) whose phosphorylation states have been defined as being essential when ablated in concert in a transgenic mouse model. However, the distinct role, if any, of each individual phosphorylation site is not established.
Methods and Results: Three transgenic lines were generated: DAA that expressed cMyBP-C containing Asp273-Ala282-Ala302, AAD, in which a charged amino acid was placed at residue 302 (Ala273-Ala282-Asp302) and SDS Ala273-Asp 282-Ala302). The effects of these replacements were determined at the whole animal and organ levels. The DAA and AAD mice showed pathology that was more severe than either the cMyBP-C null animals or animals that expressed cMyBP-C that was unable to be phosphorylated at any of the three sites. Both the DAA and AAD hearts exhibited atrial enlargement, left ventricular chamber dilation, extensive interstitial fibrosis, irregular cardiac rhythm and sudden cardiac death. Ultrastructural studies of DAA mice confirmed myocyte disarray. Yeast two hybrid analyses were used to determine the effects of altering the charge at these sites with respect to the ability of cMyBP-C to bind to the S2 domain of myosin or cardiac actin. While phosphorylation at either Ser-273 or Ser302 did not affect myosin S2 interaction, substitution of Ser 282 with a charged residue prevented interaction of cMyBP-C with both S2 myosin and actin.
Conclusion: Our results confirm the non-equivalency of these three sites in terms of the effects of their post-translational modifications on cMyBP-C functionality in the heart. Ser-282 appears to be the key residue in controlling S2 interaction with the protein. Chronic phosphorylation at one of these sites in the absence of the other, adjacent sites' ability to be phosphorylated leads to severe cardiac remodeling and sudden death.
- © 2012 by American Heart Association, Inc.