Mechanism of diacylglycerol-induced membrane targeting and activation of protein kinase Ctheta.
Protein kinase C (PKC) is a novel PKC that plays a key role in T lymphocyte activation. PKC has been shown to be specifically recruited to the immunological synapse in response to T cell receptor activation. To understand the basis of its unique subcellular localization properties, we investigated the mechanism of in vitro and cellular sn-1,2-diacylglycerol (DAG)-mediated membrane binding of PKC. PKC showed phosphatidylserine selectivity in membrane binding and kinase action, which contributes to its translocation to the phosphatidylserine-rich plasma membrane in HEK293 cells. Unlike any other PKCs characterized so far, the isolated C1B domain of PKC had much higher affinity for DAG-containing membranes than the C1A domain. Also, the mutational analysis indicates that the C1B domain plays a predominant role in the DAG-induced membrane binding and activation of PKC. Furthermore, the Ca(2+)-independent C2 domain of PKC has significant affinity for anionic membranes, and the truncation of the C2 domain greatly enhanced the membrane affinity and enzyme activity of PKC. In addition, membrane binding properties of Y90E and Y90F mutants indicate that phosphorylation of Tyr(90) of the C2 domain enhances the affinity of PKC for model and cell membranes. Collectively, these results show that PKC has a unique membrane binding and activation mechanism that may account for its subcellular targeting properties.
Melowic
Heather R
HR
Department of Chemistry, University of Illinois, Chicago, Illinois 60607, USA.
Stahelin
Robert V
RV
Blatner
Nichole R
NR
Tian
Wen
W
Hayashi
Keitaro
K
Altman
Amnon
A
Cho
Wonhwa
W
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D004075:Diglycerides / Q000737:chemistry*; D004789:Enzyme Activation; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D007313:Insecta; D007527:Isoenzymes / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D018613:Microscopy, Confocal; D010718:Phosphatidylserines / Q000737:chemistry; D011493:Protein Kinase C / Q000737:chemistry* / Q000378:metabolism*; D000074121:Protein Kinase C-theta; D021381:Protein Transport; D020349:Surface Plasmon Resonance; D013601:T-Lymphocytes / Q000378:metabolism
D004075:Diglycerides; D007527:Isoenzymes; D010718:Phosphatidylserines; C076233:PRKCQ protein, human; D011493:Protein Kinase C; D000074121:Protein Kinase C-theta