Phosphorylation of Cardiac Troponin I at Protein Kinase C Site Threonine 144 Depresses Cooperative Activation of Thin Filaments
There is evidence for PKC-dependent multisite phosphorylation of cardiac troponin I (cTnI) at Ser-23 and Ser-24 (also PKA sites) in the cardiac-specific N-terminal extension and at Thr-144, a unique residue in the inhibitory region. The functional effect of these phosphorylations in combination is o...
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Veröffentlicht in: | The Journal of biological chemistry 2010-04, Vol.285 (16), p.11810-11817 |
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Zusammenfassung: | There is evidence for PKC-dependent multisite phosphorylation of cardiac troponin I (cTnI) at Ser-23 and Ser-24 (also PKA sites) in the cardiac-specific N-terminal extension and at Thr-144, a unique residue in the inhibitory region. The functional effect of these phosphorylations in combination is of interest in view of data indicating intramolecular interaction between the N-terminal extension and the inhibitory region of cTnI. To determine the role of PKC-dependent phosphorylation of cTnI on sarcomeric function, we measured contractile regulation at multiple levels of complexity. Ca2+ binding to thin filaments reconstituted with either cTnI(wild-type) or pseudo-phosphorylated cTnI(S23D/S24D), cTnI(T144E), and cTnI(S23D/S24D/T144E) was determined. Compared with controls regulated by cTnI(wild-type), thin filaments with cTnI(S23D/S24D) and cTnI(S23D/S24D/T144E) exhibited decreased Ca2+ sensitivity. In contrast, there was no significant difference between Ca2+ binding to thin filaments with cTnI(wild-type) and with cTnI(T144E). Studies of the pCa-force relations in skinned papillary fibers regulated by these forms of cTnI yielded similar results. However, in both the Ca2+ binding measurements and the skinned fiber tension measurements, the presence of cTnI(S23D/S24D/T144E) induced a much lower Hill coefficient than either wild type, S23D/S24D, or T144E. These data highlight the importance of thin filament-based cooperative mechanisms in cardiac regulation, with implications for mechanisms of control of function in normal and pathological hearts. |
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ISSN: | 0021-9258 1083-351X |
DOI: | 10.1074/jbc.M109.055657 |