Partial replacement of cardiac troponin I with a non-phosphorylatable mutant at serines 43/45 attenuates the contractile dysfunction associated with PKCε phosphorylation

We have previously reported a transgenic mouse that over-expresses constitutively active PKC epsilon in the myocardium and exhibits a steady progression to heart failure. Associated with the decline in function was an increased phosphorylation of sarcomeric proteins including cardiac troponin I (cTnI). To determine whether PKC epsilon phosphorylation of cTnI is sufficient to induce cardiac maladaptation, we have generated a double transgenic mouse (DbTG) that expresses constitutively active PKC epsilon. and cTnI harboring non-phosphorylatable mutations in the putative PKC phosphorylation sites (S43A, S45A). We compared the hemodynamic and biochemical properties of the hearts from the DbTG mice to the nontransgenic and single transgenic lines at both 3 and 12 months of age. While no significant differences in LV function were noted in 3-month groups, the depression of function in the PKU mice was attenuated in the double transgenic mice at 12 months. The improvement in cardiac function was correlated with decreased beta-myosin heavy chain and ANF mRNA expression in the 12m DbTG mice. The extent of cTnl phosphorylation was determined using a novel one-dimensional, non-equilibrium isoelectric focusing technique. At 3 months the migration of cTnI phospho-species was different in the PKC epsilon mice and to a lesser degree in the DbTG compared to all other groups. At 12 months additional phospho-species were observed in both the PKC epsilon and DbTG samples, along with all overall shift in the distribution of phospho-species in all groups due to age. These results Suggest that phosphorylation of cTnI by PKC epsilon is associated with contractile dysfunction and partial replacement of serines 43/45 improves cardiac performance. Therefore, We Conclude that phosphorylation of cTnI at Ser 43 and 45 may contribute to the progression of failure. (c) 2006 Elsevier Ltd. All rights reserved.