PHOSPHORYLATION OF VAMP SYNAPTOBREVIN IN SYNAPTIC VESICLES BY ENDOGENOUS PROTEIN-KINASES

VAMP/synaptobrevin (SYB), an integral membrane protein of small synapt ic vesicles, is specifically cleaved by tetanus neurotoxin and botulin um neurotoxins B, D, F, and G and is thought to play an important role in the docking and/or fusion of synaptic vesicles with the presynapti c membrane. Potential phosphorylation sites for various kinases are pr esent in SYB sequence, We have studied whether SYB is a substrate for protein kinases that are present in nerve terminals and known to modul ate neurotransmitter release. SYB can be phosphorylated within the sam e Vesicle by endogenous Ca2+/calmodulin-dependent protein kinase II (C aMKII) associated with synaptic vesicles. This phosphorylation reactio n occurs rapidly and involves serine and threonine residues in the cyt oplasmic region of SYB. Similarly to CaMKII, a casein kinase II (CasKI I) activity copurifying with synaptic vesicles is able to phosphorylat e SYB selectively on serine residues of the cytoplasmic region. This p hosphorylation reaction is markedly stimulated by sphingosine, a sphin golipid known to activate CasKII and to inhibit CaMKII and protein kin ase C. The results show that SYB is a potential substrate for protein kinases involved in the regulation of neurotransmitter release and ope n the possibility that phosphorylation of SYB plays a role in modulati ng the molecular interactions between synaptic vesicles and the presyn aptic membrane.