A 29-KILODALTON GOLGI SOLUBLE N-ETHYLMALEIMIDE-SENSITIVE FACTOR ATTACHMENT PROTEIN-RECEPTOR (VTI1-RP2) IMPLICATED IN PROTEIN TRAFFICKING INTHE SECRETORY PATHWAY

Expressed sequence tags coding for a potential SNARE (soluble N-ethylm aleimide-sensitive factor attachment protein receptor) were revealed d uring data base searches. The deduced amino acid sequence of the compl ete coding region predicts a 217-residue protein with a COOH-terminal hydrophobic membrane anchor. Affinity-purified antibodies raised again st the cytoplasmic region of this protein specifically detect a 29-kil o-dalton integral membrane protein enriched in the Gels membrane. Indi rect immunofluorescence microscopy reveals that this protein is mainly associated with the Golgi apparatus. When detergent extracts of the G olgi membrane are incubated with immobilized glutathione S-transferase a soluble N-ethylmaleimide-sensitive factor attachment protein (GST-a lpha-SNAP), this protein was specifically retained. This protein has b een independently identified and termed Vtil-rp2, and it is homologous to Vti1p, a yeast Gels SNARE. We further show that Vti1-rp2 can be qu alitatively coimmunoprecipitated with Gels syntaxin 5 and syntaxin 6, suggesting that Vti1-rp2 exists in at least two distinct Gels SNARE co mplexes. In cells microinjected with antibodies against Vti1-rp2, tran sport of the envelope protein (G-protein) of vesicular stomatitis viru s from the endoplasmic reticulum to the plasma membrane was specifical ly arrested at the Golgi apparatus, providing further evidence for fun ctional importance of Vtil-rpa in protein trafficking in the secretory pathway.