A PHOSPHATIDYLINOSITOL 4,5-BISPHOSPHATE-SENSITIVE CASEIN KINASE I-ALPHA ASSOCIATES WITH SYNAPTIC VESICLES AND PHOSPHORYLATES A SUBSET OF VESICLE PROTEINS

In interphase cells, alpha-casein kinase I (alpha-CKI) is found associ ated with cytosolic vesicular structures, the centrosome, and within t he nucleus. To identify the specific vesicular structures with which a lpha-CKI is associated, established cell lines and primary rat neurons were immunofluorescently labeled with an antibody raised to alpha-CKI . In nonneuronal cells, alpha-CKI colocalizes with vesicular structure s which align with microtubules and are partially coincident with both Golgi and endoplasmic reticulum markers. In neurons, alpha-CKI coloca lizes with synaptic vesicle markers. When synaptic vesicles were purif ied from rat brain, they were highly enriched in a CKI, based on activ ity and immunoreactivity. The synaptic vesicle-associated CKI is an ex trinsic kinase and was eluted from synaptic vesicles and purified. Thi s purified CKI has properties most similar to alpha-CKI. When the acti vities of casein kinase I or II were specifically inhibited on isolate d synaptic vesicles, CKI was shown to phosphorylate a specific subset of vesicle proteins, one of which was identified as the synaptic vesic le-specific protein SV2. As with alpha-CKI, the synaptic vesicle CKI i s inhibited by phosphatidylinositol 4,5-bisphosphate (PIP2). However, synthesis of PIP2 was detected only in plasma membrane-containing frac tions. Therefore, PIP2 may spatially regulate CKI. Since PIP2 synthesi s is required for secretion, this inhibition of CKI may be important f or the regulation of secretion.