A role for protein kinase C in the regulation of membrane fluidity and Ca²(+) flux at the endoplasmic reticulum and plasma membranes of HEK293 and Jurkat cells.
Protein kinase C (PKC) plays a prominent role in the regulation of a variety of cellular functions, including Ca²(+) signalling. In HEK293 and Jurkat cells, the Ca²(+) release and Ca²(+) uptake stimulated by several different activators were attenuated by activation of PKC with phorbol myristate acetate (PMA) or 1-oleoyl-2-acetyl-sn-glycerol (OAG) and potentiated by PKC inhibition with Gö6983 or knockdown of PKCα or PKCβ using shRNA. Immunostaining and Western blotting analyses revealed that PKCα and PKCβII accumulated at the plasma membrane (PM) and that these isoforms, along with PKCβI, also translocated to the endoplasmic reticulum (ER) upon activation with PMA. Measurements of membrane fluidity showed that, like the cell membrane stabilizers bovine serum albumin (BSA) and ursodeoxycholate (UDCA), PMA and OAG significantly reduced the fluidity of both the PM and ER membranes; these effects were blocked in PKC-knockdown cells. Interestingly, both BSA and UDCA inhibited the Ca²(+) responses to agonists to the same extent as PMA, whereas Tween 20, which increases membrane fluidity, raised the internal Ca²(+) concentration. Thus, activation of PKC induces both translocation of PKC to the PM and ER membranes and downregulation of membrane fluidity, thereby negatively modulating Ca²(+) flux.
Department of Pharmacology, Capital Medical University, Beijing 100069, China.
Lihong
L
Chen
Qingli
Q
Meng
Xian
X
Jing
Pingxiang
P
Xu
Dali
D
Luo
D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling; D002462:Cell Membrane / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D019169:Jurkat Cells; D008560:Membrane Fluidity; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D021381:Protein Transport; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D015398:Signal Transduction
D043562:Receptors, G-Protein-Coupled; D011493:Protein Kinase C; D002118:Calcium