Differential regulation of cardiac actomyosin S-1 MgATPase by protein kinase C isozyme-specific phosphorylation of specific sites in cardiac troponin I and its phosphorylation site mutants

被引:70
作者
Noland, TA
Raynor, RL
Jideama, NM
Guo, XD
Kazanietz, MG
Blumberg, PM
Solaro, RJ
Kuo, JF
机构
[1] UNIV ILLINOIS,COLL MED,DEPT PHYSIOL & BIOPHYS,CHICAGO,IL 60612
[2] NCI,CELLULAR CARCINOGENESIS & TUMOR PROMOT LAB,MOL MECHANISMS TUMOR PROMOT SECT,BETHESDA,MD 20892
关键词
D O I
10.1021/bi9616357
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The significance of site-specific phosphorylation by protein kinase C (PKC) isozymes alpha and delta and protein kinase A (PKA) of troponin I (TnI) and its phosphorylation site mutants in the regulation of Ca2+-stimulated MgATPase activity of reconstituted actomyosin S-1 was investigated. The genetically defined TnI mutants used were T144A, S43A/S45A, S43A/S45A/T144A (in which the PKC phosphorylation sites Thr-144 and Ser-43/Ser-45 were respectively substituted by Ala) and N32 (in which the first 32 amino acids in the NH2-terminal sequence containing Ser-23/Ser-24 were deleted). Although the PKC isozymes displayed different substrate phosphorylation kinetics, PKC-alpha phosphorylated equally well TnI wild type and all mutants, whereas N32 was a much poorer substrate for PKC-delta. Furthermore, the two PKC isozymes exhibited discrete specificities in phosphorylating distinct sites in TnI and its mutants, either as individual subunits or as components of the reconstituted troponin complex. Unlike PKC-alpha, PKC-delta favorably phosphorylated the PKA-preferred site Ser-23/Ser-24 and hence, like PKA, reduced the Ca2+ sensitivity of the reconstituted actomyosin S-1 MgATPase. In contrast, PKC-alpha preferred to phosphorylate Ser-43/Ser-45 (common sites for all isozymes) and thus reduced the maximal Ca2+-stimulated activity of the MgATPase. In this respect, PKC-delta, by cross-phosphorylating the PKA sites, functioned as a hybrid of PKC-alpha and PKA. The site specificities and hence functional differences between PKC-a and -delta were most evident at low phosphorylation (1 mol of phosphate/mol) of TnI wild type and were magnified when S43A/S45A and N32 were used as substrates. The present study has demonstrated, for the first time, that distinct functional consequences could arise from the site-selective preferences of PKC-alpha and -delta for phosphorylating a single substrate in the myocardium, i.e., TnI.
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页码:14923 / 14931
页数:9
相关论文
共 53 条
[1]   CHARACTERIZATION OF PROTEIN-KINASE-C ISOTYPE EXPRESSION IN ADULT-RAT HEART - PROTEIN-KINASE C-EPSILON IS A MAJOR ISOTYPE PRESENT, AND IT IS ACTIVATED BY PHORBOL ESTERS, EPINEPHRINE, AND ENDOTHELIN [J].
BOGOYEVITCH, MA ;
PARKER, PJ ;
SUGDEN, PH .
CIRCULATION RESEARCH, 1993, 72 (04) :757-767
[2]   ALPHA-1-ADRENERGIC AND MUSCARINIC CHOLINERGIC STIMULATION OF PHOSPHOINOSITIDE HYDROLYSIS IN ADULT-RAT CARDIOMYOCYTES [J].
BROWN, JH ;
BUXTON, IL ;
BRUNTON, LL .
CIRCULATION RESEARCH, 1985, 57 (04) :532-537
[3]   PHORBOL ESTER AND DIOCTANOYLGLYCEROL STIMULATE MEMBRANE ASSOCIATION OF PROTEIN KINASE-C AND HAVE A NEGATIVE INOTROPIC EFFECT MEDIATED BY CHANGES IN CYTOSOLIC CA-2+ IN ADULT-RAT CARDIAC MYOCYTES [J].
CAPOGROSSI, MC ;
KAKU, T ;
FILBURN, CR ;
PELTO, DJ ;
HANSFORD, RG ;
SPURGEON, HA ;
LAKATTA, EG .
CIRCULATION RESEARCH, 1990, 66 (04) :1143-1155
[4]   EXPRESSION OF PROTEIN-KINASE-C ISOFORMS DURING CARDIAC VENTRICULAR DEVELOPMENT [J].
CLERK, A ;
BOGOYEVITCH, MA ;
FULLER, SJ ;
LAZOU, A ;
PARKER, PJ ;
SUGDEN, PH .
AMERICAN JOURNAL OF PHYSIOLOGY-HEART AND CIRCULATORY PHYSIOLOGY, 1995, 269 (03) :H1087-H1097
[5]   ARACHIDONIC ACID-DEPENDENT PHOSPHORYLATION OF TROPONIN-I AND MYOSIN LIGHT-CHAIN-2 IN CARDIAC MYOCYTES [J].
DAMRON, DS ;
DARVISH, A ;
MURPHY, L ;
SWEET, W ;
MORAVEC, CS ;
BOND, M .
CIRCULATION RESEARCH, 1995, 76 (06) :1011-1019
[6]   PROTEIN-KINASE-C - A QUESTION OF SPECIFICITY [J].
DEKKER, LV ;
PARKER, PJ .
TRENDS IN BIOCHEMICAL SCIENCES, 1994, 19 (02) :73-77
[7]   LOCALIZATION OF PROTEIN-KINASE-C ISOZYMES IN CARDIAC MYOCYTES [J].
DISATNIK, MH ;
BURAGGI, G ;
MOCHLYROSEN, D .
EXPERIMENTAL CELL RESEARCH, 1994, 210 (02) :287-297
[8]   PHORBOL ESTER INCREASES CALCIUM CURRENT AND SIMULATES THE EFFECTS OF ANGIOTENSIN-II ON CULTURED NEONATAL RAT-HEART MYOCYTES [J].
DOSEMECI, A ;
DHALLAN, RS ;
COHEN, NM ;
LEDERER, WJ ;
ROGERS, TB .
CIRCULATION RESEARCH, 1988, 62 (02) :347-357
[9]   STUDIES ON PHOSPHORYLATION OF INHIBITORY SUBUNIT OF TROPONIN DURING MODIFICATION OF CONTRACTION IN PERFUSED RAT-HEART [J].
ENGLAND, PJ .
BIOCHEMICAL JOURNAL, 1976, 160 (02) :295-304
[10]  
FABIATO A, 1979, J PHYSIOL-PARIS, V75, P463