Protein kinase C alpha modulates the Ca2+ influx phase of the Ca2+ response to 1alpha,25-dihydroxy-vitamin-D3 in skeletal muscle cells.
Treatment of chick skeletal muscle cells with 1alpha,25-dihydroxy-vitamin D3 [1alpha,25(OH)2D3] triggers a rapid and sustained increase in cytosolic Ca2+ ([Ca2+]i), which depends on Ca2+ mobilization from inner stores and extracellular Ca2+ entry. Fluorimetric analysis of changes in [Ca2+]i in Fura-2-loaded cells revealed that the hormone significantly stimulates the Ca2+ influx phase within the concentration range of 10(-12)-10(-6) M, with maximal effects (3.5-fold increase) at 10(-9) M 1alpha,25(OH)2D3. The effects of the sterol on the Ca2+ entry pathway were abolished by the PKC inhibitors bisindolylmaleimide and calphostin. We have recently shown that, in these cells, 1alpha,25(OH)2D3 activates and translocates PKC alpha to the membrane, suggesting that this isozyme accounts for PKC-dependent 1alpha,25(OH)2D3 modulation of Ca2+ entry. The role of PKC alpha was specifically addressed here using antisense technology. When the expression of PKC alpha was selectively knocked out by intranuclear microinjection of an antisense oligonucleotide against PKC alpha mRNA, the Ca2+ influx component of the response to 1alpha,25(OH)2D3 was markedly reduced (-60%). These results demonstrate that 1alpha,25(OH)2D3-induced activation of PKC alpha enhances extracellular Ca2+ entry partially contributing to maintainance of the sustained phase of the Ca2+ response to the sterol.
Capiati
D A
DA
Departamento de Biología, Bioquímica y Farmacia, Universidad Nacional del Sur, San Juan Bahia Blanca, Argentina.
Vazquez
G
G
Boland
R L
RL
D000818:Animals; D002117:Calcitriol / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D002120:Calcium Channel Agonists / Q000494:pharmacology*; D015220:Calcium Channels / Q000378:metabolism; D002478:Cells, Cultured; D002642:Chick Embryo; D002645:Chickens; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D007527:Isoenzymes / Q000235:genetics* / Q000378:metabolism*; D018485:Muscle Fibers, Skeletal / Q000166:cytology / Q000201:enzymology*; D018482:Muscle, Skeletal / Q000166:cytology; D016376:Oligonucleotides, Antisense; D011493:Protein Kinase C / Q000235:genetics* / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D012333:RNA, Messenger / Q000032:analysis
D002120:Calcium Channel Agonists; D015220:Calcium Channels; D007527:Isoenzymes; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D002117:Calcitriol; D002118:Calcium