Functional characterization of the human α-cardiac actin mutations Y166C and M305L involved in hypertrophic cardiomyopathy

Cell Mol Life Sci. 2012 Oct;69(20):3457-79. doi: 10.1007/s00018-012-1030-5. Epub 2012 May 29.


Inherited cardiomyopathies are caused by point mutations in sarcomeric gene products, including α-cardiac muscle actin (ACTC1). We examined the biochemical and cell biological properties of the α-cardiac actin mutations Y166C and M305L identified in hypertrophic cardiomyopathy (HCM). Untagged wild-type (WT) cardiac actin, and the Y166C and M305L mutants were expressed by the baculovirus/Sf9-cell system and affinity purified by immobilized gelsolin G4-6. Their correct folding was verified by a number of assays. The mutant actins also displayed a disturbed intrinsic ATPase activity and an altered polymerization behavior in the presence of tropomyosin, gelsolin, and Arp2/3 complex. Both mutants stimulated the cardiac β-myosin ATPase to only 50 % of WT cardiac F-actin. Copolymers of WT and increasing amounts of the mutant actins led to a reduced stimulation of the myosin ATPase. Transfection of established cell lines revealed incorporation of EGFP- and hemagglutinin (HA)-tagged WT and both mutant actins into cytoplasmic stress fibers. Adenoviral vectors of HA-tagged WT and Y166C actin were successfully used to infect adult and neonatal rat cardiomyocytes (NRCs). The expressed HA-tagged actins were incorporated into the minus-ends of NRC thin filaments, demonstrating the ability to form hybrid thin filaments with endogenous actin. In NRCs, the Y166C mutant led after 72 h to a shortening of the sarcomere length when compared to NRCs infected with WT actin. Thus our data demonstrate that a mutant actin can be integrated into cardiomyocyte thin filaments and by its reduced mode of myosin interaction might be the basis for the initiation of HCM.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actin Cytoskeleton / metabolism*
  • Actins / genetics*
  • Actins / metabolism*
  • Adenoviridae / genetics
  • Animals
  • Animals, Newborn
  • Baculoviridae / genetics
  • Binding Sites
  • Cardiomyopathy, Hypertrophic / genetics
  • Cardiomyopathy, Hypertrophic / metabolism*
  • Cells, Cultured
  • Humans
  • Immunoblotting
  • Mutation / genetics*
  • Myocytes, Cardiac / cytology
  • Myocytes, Cardiac / metabolism
  • Myosins / metabolism
  • Rats
  • Sarcomeres / physiology


  • ACTC1 protein, human
  • Actins
  • Myosins