Changes in end-to-end interactions of tropomyosin affect mouse cardiac muscle dynamics

1 Cardiovascular Research Institute and Department of Systems Biology and Translational Medicine, College of Medicine, Texas A&M University System Health Science Center; and 2 Department of Biophysics and Biochemistry, Texas A&M University, College Station, Texas; and 3 Division of Molecular Cardiovascular Biology, Cincinnati Children's Hospital, Cincinnati, Ohio Submitted 23 June 2005 ; accepted in final form 21 February 2006 The ends of striated muscle tropomyosin (TM) are integral for thin filament cooperativity, determining the cooperative unit size and regulating the affinity of TM for actin. We hypothesized that altering the -TM carboxy terminal overlap end to the -TM counterpart would affect the amino-terminal association, which would alter the end-to-end interactions of TM molecules in the thin filament regulatory strand and affect the mechanisms of cardiac muscle contraction. To test this hypothesis, we generated transgenic (TG) mouse lines that express a mutant form of -TM in which the first 275 residues are from -TM and the last nine amino acids are from -TM ( -TM9aa ). Molecular analyses show that endogenous -TM mRNA and protein are nearly completely replaced with -TM9aa . Working heart preparations data show that the rates of contraction and relaxation are reduced in -TM9aa hearts. Left ventricular pressure and time to peak pressure are also reduced (–12% and –13%, respectively). The ratio of maximum to minimum first derivatives of change in left ventricular systolic pressure with respect to time (ratio of +dP/d t to –dP/d t , respectively) is increased, but is not changed significantly. Force-intracellular calcium concentration ([Ca 2+ ] i ) measurements from intact papillary fibers demonstrate that -TM9aa TG fibers produce less force per given [Ca 2+ ] i compared with nontransgenic fibers. Taken together, the data demonstrate that the rate of contraction is primarily affected in TM TG hearts. Protein docking studies show that in the mutant molecule, the overall carbon backbone is perturbed about 1.5 Å, indicating that end-to-end interactions are altered. These results demonstrate that the localized flexibility present in the coiled-coil structures of TM isoforms is different, and that plays an important role in interacting with neighboring thin filament regulatory proteins and with differentially modulating the myofilament activation processes. force-calcium; thin filament; force-frequency; myofilament activation Address for reprint requests