SNIP, a novel SNAP-25-interacting protein implicated in regulated exocytosis

Synaptosome-associated protein of 25 kDa (SNAP-25) is a presynaptic membran e protein that has been clearly implicated in membrane fusion in both devel oping and mature neurons, although its mechanisms of action are unclear. We have now identified a novel SNAP-BB-interacting protein named SNIP, SNIP i s a hydrophilic, 145-kDa protein that comprises two predicted coiled-coil d omains, two highly charged regions, and two proline-rich domains with multi ple PPXY and PXXP motifs, SNIP is selectively expressed in brain where it c odistributes with SNAP-25 in most brain regions. Biochemical studies have r evealed that SNIP is tightly associated with the brain cytoskeleton. Subcel lular fractionation and immunofluorescence localization studies have demons trated that SNIP co-localizes with SNAP-25 as well as the cortical actin cy toskeleton, suggesting that SMP serves as a linker protein connecting SNAP- 25 to the submembranous cytoskeleton. By using deletion analysis, we have m apped the binding domains of SNIP and SNAP-25, and we have demonstrated tha t the SNIP-SNAP-25 association is mediated via coiled-coil interactions. Mo reover, we have shown that overexpression of SNIP or its SNAP-25-interactin g domain inhibits Ca2+-dependent exocytosis from PC12 cells. These results indicate that SNIP is involved in regulation of neurosecretion, perhaps via its interaction with SNAP-25 and the cytoskeleton.