Identification of triadin and of histidine-rich Ca(2+)-binding protein as substrates of 60 kDa calmodulin-dependent protein kinase in junctional terminal cisternae of sarcoplasmic reticulum of rabbit fast muscle.
The endogenous calmodulin-protein kinase system of sarcoplasmic reticulum terminal cisternae of rabbit fast-twitch muscle was studied. Investigation of a single Ca(2+)-channel in terminal cisternae fused to planar lipid bilayers demonstrated that the endogenous kinase inhibits the channel, although it remained unclear whether the phosphorylation sites are on the channel protein or on other junctional sarcoplasmic reticulum specific proteins [Hain et al., (1994) Biophys. J. 67, 1823-1833]. Our results, which show that two junctional sarcoplasmic reticulum specific proteins, i.e., triadin and histidine-rich Ca(2+)-binding protein, but not the ryanodine receptor/Ca(2+)-channel protein, are phosphorylated by membrane-bound 60 kDa protein kinase, seem to be able to resolve this ambiguity. Furthermore, such a probably specific protein isoform of calmodulin-protein kinase, by its substrate specificity and exposure to the cytoplasmic side of terminal cisternae at the junctional membrane domain and based on protease sensitivity, also seems to possess some of the potential requirements for a regulatory role in the functional state of the Ca(2+)-channel.
Damiani
E
E
Dipartimento di Scienze biomediche sperimentali, Universitá di Padova, Italy.
Picello
E
E
Saggin
L
L
Margreth
A
A
D000818:Animals; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism; D002352:Carrier Proteins; D002451:Cell Compartmentation; D008970:Molecular Weight; D009124:Muscle Proteins / Q000378:metabolism*; D009132:Muscles / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000378:metabolism* / Q000648:ultrastructure
D002135:Calcium-Binding Proteins; D002147:Calmodulin; D002352:Carrier Proteins; C489419:HRC protein, Oryctolagus cuniculus; D009124:Muscle Proteins; D010750:Phosphoproteins; C070048:triadin; D017871:Calcium-Calmodulin-Dependent Protein Kinases