Regulation of Ca2+ uptake in skeletal muscle by 1,25-dihydroxyvitamin D3: role of phosphorylation and calmodulin.
Experiments were carried out to obtain information about the mechanism underlying the fast action of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) in skeletal muscle. N-2'-o-dibutyryladenosine-3',5'-cyclic monophosphate (dbcAMP), similarly as 1,25(OH)2D3 (5 x 10(-10) M), rapidly increased 45Ca uptake by soleus muscle from vitamin D-deficient chicks (+25% and +98% at 3 min and 10 min, respectively) in a dose-dependent manner. The effects of the cAMP analog (10 microM) and 1,25(OH)2D3 could be abolished by the Ca(2+)-channel blocker nifedipine and the calmodulin antagonist flufenazine. Calmodulin binding by two muscle microsomal proteins of 28 kDa and 30 kDa was stimulated within 1 min of exposure of the tissue to 1,25(OH)2D3. Direct effects of the sterol on membrane calmodulin binding were shown with isolated microsomes. The 1,25(OH)2D3-mediated rise of [125I]calmodulin binding to microsomal membranes was dependent on the presence of medium ATP. Forskolin (10 microM) and cAMP (10 microM) also increased [125I]calmodulin binding (+75% and +64%, respectively, with respect to controls). Pretreatment of microsomal membranes with cAMP-dependent protein kinase inhibitor (1 microgram/ml) or addition of alkaline phosphates (1 U/ml) after hormonal treatment caused complete inhibition of 1,25(OH)2D3-induced [125I]calmodulin binding to microsomal membrane proteins. These results imply modifications of membrane protein phosphorylation through the cAMP signal pathway and in turn of calmodulin binding in the mechanism by which 1,25(OH)2D3 rapidly stimulates skeletal muscle Ca2+ uptake.
Massheimer
V
V
Departamento de Biologia, Universidad Nacional del Sur, Bahia Blanca, Argentina.
Fernandez
L M
LM
Boland
R
R
de Boland
A R
AR
D000255:Adenosine Triphosphate / Q000502:physiology; D000469:Alkaline Phosphatase / Q000494:pharmacology; D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D002117:Calcitriol / Q000494:pharmacology*; D002118:Calcium / Q000493:pharmacokinetics*; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology; D002147:Calmodulin / Q000187:drug effects / Q000502:physiology*; D002645:Chickens; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D066298:In Vitro Techniques; D008861:Microsomes / Q000187:drug effects / Q000378:metabolism; D009132:Muscles / Q000378:metabolism*; D009543:Nifedipine / Q000494:pharmacology; D010766:Phosphorylation; D011494:Protein Kinases / Q000494:pharmacology; D013997:Time Factors
D015220:Calcium Channels; D002147:Calmodulin; D005576:Colforsin; D003994:Bucladesine; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D011494:Protein Kinases; D000469:Alkaline Phosphatase; D002117:Calcitriol; D009543:Nifedipine; D002118:Calcium