Syntaxin 7 complexes with mouse Vps10p tail interactor 1b, Syntaxin 6, vesicle-associated membrane protein (VAMP)8, and VAMP7 in B16 melanoma cells

Syntaxin 7 is a mammalian target soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) involved in membrane transport between late endosomes and lysosomes. The aim of the present study was to use immun oaffinity techniques to identify proteins that interact with Syntaxin 7. We reasoned that this would be facilitated by the use of cells producing high levels of Syntaxin 7, Screening of a large number of tissues and cell line s revealed that Syntaxin 7 is expressed at very high levels in B16 melanoma cells. Moreover, the expression of Syntaxin 7 increased in these cells as they underwent melanogenesis. From a large scale Syntaxin 7 immunoprecipita tion, we have identified six polypeptides using a combination of electrospr ay mass spectrometry and immunoblotting. These polypeptides corresponded to Syntaxin 7, Syntaxin 6, mouse Vps10p tail interactor 1b (mVti1b), alpha -s ynaptosome-associated protein (SNAP), vesicle-associated membrane protein ( VAMP)8, VAMP7, and the protein phosphatase 1M regulatory subunit. We also o bserved partial colocalization between Syntaxin 6 and Syntaxin 7, between S yntaxin 6 and mVti1b, but not between Syntaxin 6 and the early endosomal t- SNARE Syntaxin 13. Based on these and data reported previously, we propose that Syntaxin 7/mVti1b/Syntaxin 6 may form discrete SNARE complexes with ei ther VAMP7 or VAMPS to regulate fusion events within the late endosomal pat hway and that these events may play a critical role in melanogenesis.