DIFFERENTIAL TARGETING OF PROTEIN-KINASE-C AND CAM KINASE-II SIGNALINGS TO VIMENTIN

Hydrolysis of inositol phospholipids by receptor stimulation activates two separate signaling pathways, one leading to the activation of pro tein kinase C (C kinase) via formation of diacylglycerol. The other is the inositol trisphosphate (IP3)/Ca2+ pathway and a major downstream kinase which is activated is Ca2+/calmodulin-dependent protein kinase II (CaM kinase II). To examine signaling pathways of C kinase and CaM kinase II to the cytoskeletal protein vimentin, we prepared monoclonal antibodies YT33 and MO82 which recognize the phosphorylation state of vimentin by C kinase and by CaM kinase II, respectively. Ectopic expr ession of constitutively active C kinase or CaM kinase II in primary c ultured astrocytes by microinjection of the corresponding expression v ectors induced phosphorylation of vimentin at each specific phosphoryl ation site, followed by reorganization of vimentin filament networks. In contrast, simultaneous activation of C kinase and CaM kinase II by inositol. phospholipid hydrolysis with receptor stimulation led to an exclusive phosphorylation of vimentin at the CaM kinase II site, not a t the site of C kinase. These results indicate that the intracellular targeting of C kinase and CaM kinase II signalings to vimentin is regu lated separately, under physiological conditions.