Protein kinase C phosphomimetics alter thin filament Ca2+ binding properties.
Adrenergic stimulation modulates cardiac function by altering the phosphorylation status of several cardiac proteins. The Troponin complex, which is the Ca(2+) sensor for cardiac contraction, is a hot spot for adrenergic phosphorylation. While the effect of β-adrenergic related PKA phosphorylation of troponin I at Ser23/24 is well established, the effects of α-adrenergic induced PKC phosphorylation on multiple sites of TnI (Ser43/45, Thr144) and TnT (Thr194, Ser198, Thr203 and Thr284) are much less clear. By utilizing an IAANS labeled fluorescent troponin C, TnC(IAANS)(T53C), we systematically examined the site specific effects of PKC phosphomimetic mutants of TnI and TnT on TnC's Ca(2+) binding properties in the Tn complex and reconstituted thin filament. The majority of the phosphomemetics had little effect on the Ca(2+) binding properties of the isolated Tn complex. However, when incorporated into the thin filament, the phosphomimetics typically altered thin filament Ca(2+) sensitivity in a way consistent with their respective effects on Ca(2+) sensitivity of skinned muscle preparations. The altered Ca(2+) sensitivity could be generally explained by a change in Ca(2+) dissociation rates. Within TnI, phosphomimetic Asp and Glu did not always behave similar, nor were Ala mutations (used to mimic non-phosphorylatable states) benign to Ca(2+) binding. Our results suggest that Troponin may act as a hub on the thin filament, sensing physiological stimuli to modulate the contractile performance of the heart.
Department of Physiology and Cell Biology, The Ohio State University, Columbus, Ohio, United States of America.
Bin
B
Liu
Department of Physiology and Cell Biology, The Ohio State University, Columbus, Ohio, United States of America.
Joseph J
JJ
Lopez
Department of Physiology and Cell Biology, The Ohio State University, Columbus, Ohio, United States of America.
Brandon J
BJ
Biesiadecki
Department of Physiology and Cell Biology, The Ohio State University, Columbus, Ohio, United States of America.
Jonathan P
JP
Davis
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002417:Cattle; D006801:Humans; D007700:Kinetics; D046912:Multiprotein Complexes / Q000378:metabolism; D009154:Mutation; D010766:Phosphorylation; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism*; D011817:Rabbits; D014336:Troponin / Q000378:metabolism; D019209:Troponin C / Q000235:genetics / Q000378:metabolism*
D046912:Multiprotein Complexes; D014336:Troponin; D019209:Troponin C; D011493:Protein Kinase C; D002118:Calcium