Stimulation of calmodulin binding to skeletal muscle membrane proteins by 1,25-dihydroxy-vitamin D3.
Previous work has shown that 1,25-dihydroxy-vitamin D3 rapidly increases calmodulin levels of skeletal muscle membranes without altering the muscle cell calmodulin content. Therefore, the effects of the sterol on the binding of calmodulin to specific muscle membrane proteins were investigated. Soleus muscles from vitamin D-deficient chicks were treated in vitro for short intervals (5-15 min) with physiological concentrations of 1,25-dihydroxy-vitamin D3. Proteins of mitochondria and microsomes isolated by differential centrifugation were separated on sodium dodecyl sulfate polyacrylamide gels. Calmodulin-binding proteins were identified by a [125I]calmodulin gel overlay procedure followed by autoradiography. 1,25-Dihydroxy-vitamin D3 increased the binding of labelled calmodulin to a major, calcium-independent, calmodulin-binding protein of 28 Kda localized in microsomes, and to minor calmodulin-binding proteins of 78 and 130 Kda proteins localized in mitochondria. The binding of [125I]calmodulin to these proteins was abolished by flufenazine or excess non-radioactive calmodulin. 1,25-Dihydroxy-vitamin D3 rapidly increased muscle tissue Ca uptake and cyclic AMP levels and stimulated the phosphorylation of several membrane proteins including those whose calmodulin-binding capacity potentiates. Analogously to the sterol, forskolin increased membrane calmodulin content, calmodulin binding to the 28 Kda microsomal protein and 45Ca uptake by soleus muscle preparations. Forskolin also induced a similar profile of changes in muscle membrane protein phosphorylation as the hormone. These results suggest that 1,25-dihydroxy-vitamin D3 affects calmodulin distribution in muscle cells through cyclic AMP-dependent phosphorylation of membrane calmodulin-binding proteins. These changes may play a role in the stimulation of muscle Ca uptake by the sterol.
Massheimer
V
V
Departamento de Biología, Universidad Nacional del Sur, Bahía Blanca, Argentina.
Fernandez
L M
LM
de Boland
A R
AR
D000818:Animals; D002117:Calcitriol / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism*; D002148:Calmodulin-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D002645:Chickens; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D007700:Kinetics; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism*; D008970:Molecular Weight; D009132:Muscles / Q000187:drug effects / Q000378:metabolism*; D011485:Protein Binding; D013347:Subcellular Fractions / Q000378:metabolism; D014808:Vitamin D Deficiency / Q000378:metabolism
D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; D008565:Membrane Proteins; D005576:Colforsin; D000242:Cyclic AMP; D002117:Calcitriol; D002118:Calcium