Analysis of myosin heavy chain functionality in the heart

Abstract

Comparison of mammalian cardiac α- and β-myosin heavy chain isoforms reveals 93% identity. To date, genetic methodologies have effected only minor switches in the mammalian cardiac myosin isoforms. Using cardiac-specific transgenesis, we have now obtained major myosin isoform shifts and/or replacements. Clusters of non-identical amino acids are found in functionally important regions, i.e. the surface loops 1 and 2, suggesting that these structures may regulate isoform-specific characteristics. Loop 1 alters filament sliding velocity, whereas Loop 2 modulates actin-activated ATPase rate in Dictyostelium myosin, but this remains untested in mammalian cardiac myosins. α → β isoform switches were engineered into mouse hearts via transgenesis. To assess the structural basis of isoform diversity, chimeric myosins in which the sequences of either Loop 1 + Loop 2 or Loop 2 of α-myosin were exchanged for those of β-myosin were expressed in vivo. 2-fold differences in filament sliding velocity and ATPase activity were found between the two isoforms. Filament sliding velocity of the Loop 1 + Loop 2 chimera and the ATPase activities of both loop chimeras were not significantly different compared with α-myosin. In mouse cardiac isoforms, myosin functionality does not depend on Loop 1 or Loop 2 sequences and must lie partially in other non-homologous residues.