Complex regulation of store-operated Ca2+ entry pathway by PKC-epsilon in vascular SMCs.
The role of PKC in the regulation of store-operated Ca2+ entry (SOCE) is rather controversial. Here, we used Ca2+-imaging, biochemical, pharmacological, and molecular techniques to test if Ca2+-independent PLA2beta (iPLA2beta), one of the transducers of the signal from depleted stores to plasma membrane channels, may be a target for the complex regulation of SOCE by PKC and diacylglycerol (DAG) in rabbit aortic smooth muscle cells (SMCs). We found that the inhibition of PKC with chelerythrine resulted in significant inhibition of thapsigargin (TG)-induced SOCE in proliferating SMCs. Activation of PKC by the diacylglycerol analog 1-oleoyl-2-acetyl-sn-glycerol (OAG) caused a significant depletion of intracellular Ca2+ stores and triggered Ca2+ influx that was similar to TG-induced SOCE. OAG and TG both produced a PKC-dependent activation of iPLA2beta and Ca2+ entry that were absent in SMCs in which iPLA2beta was inhibited by a specific chiral enantiomer of bromoenol lactone (S-BEL). Moreover, we found that PKC regulates TG- and OAG-induced Ca2+ entry only in proliferating SMCs, which correlates with the expression of the specific PKC-epsilon isoform. Molecular downregulation of PKC-epsilon impaired TG- and OAG-induced Ca2+ influx in proliferating SMCs but had no effect in confluent SMCs. Our results demonstrate that DAG (or OAG) can affect SOCE via multiple mechanisms, which may involve the depletion of Ca2+ stores as well as direct PKC-epsilon-dependent activation of iPLA2beta, resulting in a complex regulation of SOCE in proliferating and confluent SMCs.
Smani
Tarik
T
Ion Channel and Calcium Signaling Unit, Boston University School of Medicine, Boston, Massachusetts, USA. tasmani@us.es
Patel
Tina
T
Bolotina
Victoria M
VM
D000818:Animals; D053119:Benzophenanthridines / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D049109:Cell Proliferation; D002478:Cells, Cultured; D004075:Diglycerides / Q000378:metabolism; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D054522:Group VI Phospholipases A2 / Q000037:antagonists & inhibitors / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D009281:Naphthalenes / Q000494:pharmacology; D051744:Protein Kinase C-epsilon / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011753:Pyrones / Q000494:pharmacology; D011817:Rabbits; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism; D019284:Thapsigargin / Q000494:pharmacology; D013997:Time Factors; D014162:Transfection
C520323:1-oleoyl-2-acetoyl-sn-glycerol; D053119:Benzophenanthridines; D004075:Diglycerides; D004791:Enzyme Inhibitors; D009281:Naphthalenes; D011753:Pyrones; D019284:Thapsigargin; C068314:6-(bromomethylene)tetrahydro-3-(1-naphthaleneyl)-2H-pyran-2-one; C016299:chelerythrine; D051744:Protein Kinase C-epsilon; D054522:Group VI Phospholipases A2; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D002118:Calcium