title: Specific degradation of troponin T and I by mu-calpain and its modulation by substrate phosphorylation.
abstract: The degradation of troponin (Tn) subunits by calpain was studied by incubating either isolated cardiac Tns or myocardial cryosections with two different calpain isoenzymes isolated from rat skeletal muscle. Western-blot analysis with monoclonal antibodies against TnI and TnT showed that mu-calpain was at least ten times more active than m-calpain in degrading TnI and TnT both in vitro and in situ. TnC was completely resistant to both proteinase forms. Phosphorylation by cyclic AMP-dependent protein kinase (PKA) isolated from rat skeletal muscle reduced the sensitivity of TnI to degradation. This effect in combination with an increased efficiency of the endogenous inhibitor [Salamino, De Tullio, Michetti, Mengotti, Melloni and Pontremoli (1994) Biochem. Biophys. Res. Commun. 199, 1326-1332] probably reduces the proteolytic activity of calpain in cells on PKA stimulation. Conversely, phosphorylation by protein kinase C (PKC) resulted in a twofold increase in the degradation of TnI. Degradation by m-calpain was not modified by Tn phosphorylation. The different sensitivity to mu-calpain might be related to changes in TnI oligomeric structure. Indeed, on PKC phosphorylation, the apparent molecular mass of TnI calculated from the distribution coefficient of Tn complex in Sephadex G-100 matrix was reduced from 90 to 30 kDa suggesting dissociation of the Tn complex.
authors.affiliation: Dipartimento di Chimica Biologica, Università di Padova, Italy.
authors.forename: F
authors.initials: F
authors.lastname: Di Lisa
authors.forename: R
authors.initials: R
authors.lastname: De Tullio
authors.forename: F
authors.initials: F
authors.lastname: Salamino
authors.forename: R
authors.initials: R
authors.lastname: Barbato
authors.forename: E
authors.initials: E
authors.lastname: Melloni
authors.forename: N
authors.initials: N
authors.lastname: Siliprandi
authors.forename: S
authors.initials: S
authors.lastname: Schiaffino
authors.forename: S
authors.initials: S
authors.lastname: Pontremoli
mesh_terms: D000818:Animals; D002154:Calpain / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000378:metabolism; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology; D009206:Myocardium / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D013329:Structure-Activity Relationship; D014336:Troponin / Q000378:metabolism*; D019210:Troponin I; D020107:Troponin T
chemical_list: D007527:Isoenzymes; D010750:Phosphoproteins; D014336:Troponin; D019210:Troponin I; D020107:Troponin T; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D002154:Calpain