In situ replacement of slow skeletal myosin binding protein-c

Abstract

Myosin binding protein (MyBP-C) is a modular protein that interacts with both thin and thick filaments and regulates striated muscle contraction. The functional characteristics of MyBP-C have yet to be fully uncovered in all 3 isoforms (cardiac, fast skeletal, and slow skeletal). To manipulate cardiac (c) MyBP-C at its precise position in the sarcomere in situ, we created gene-edited ‘SpyC’ mice that allow us to use a novel ‘cut and paste’ method for cMyBP-C recombination in cardio-myocytes. This ‘cut and paste’ method allows for the in situ removal of endogenous N’-terminal domains from MyBP-C via TEV proteolysis, followed by its replacement with any desired recombinant N’-terminal MyBP-C.