title: Tyrosine phosphorylation of a SNARE protein, syntaxin 17: implications for membrane trafficking in the early secretory pathway.
abstract: The T-cell protein tyrosine phosphatase is expressed as two splice variants - TC45, a nuclear protein, and TC48, which is localized predominantly in the ER (endoplasmic reticulum). Yeast two-hybrid screening revealed direct interaction of TC48 with Syntaxin17, a SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) protein localized predominantly in the ER and to some extent in the ER-Golgi intermediate compartment. Syntaxin 17 did not interact with TC45. C-terminal 40 amino acids of TC48 were sufficient for interaction with syntaxin 17. Overexpressed syntaxin 17 was phosphorylated at tyrosine upon pervanadate treatment (a tyrosine phosphatase inhibitor/tyrosine kinase activator) of COS-1 cells. Mutational analysis identified Tyr156 in the cytoplasmic domain as the major site of phosphorylation. Endogenous syntaxin 17 was phosphorylated by pervanadate treatment in CHO and MIN6 cells but was not phosphorylated in a variety of other cell lines tested. c-Abl was identified as one of the kinases, which phosphorylates syntaxin 17 in MIN6 cells. Phosphorylation of endogenous and overexpressed syntaxin 17 was reduced in the presence of IGF receptor and EGF receptor kinase inhibitors. Serum depletion reduced pervanadate-induced phosphorylation of endogenous syntaxin 17. TC48 coexpression reduced phosphorylation of syntaxin 17 by pervanadate and purified TC48 directly dephosphorylated syntaxin 17. β-COP dispersal by overexpressed syntaxin 17 was reduced after pervanadate-induced phosphorylation. A phospho-mimicking mutant (Y156E) of syntaxin 17 showed reduced interaction with COPI vesicles. These results suggest that tyrosine phosphorylation of syntaxin 17 is likely to have a role in regulating syntaxin 17 dependent membrane trafficking in the early secretory pathway.
authors.affiliation: Centre for Cellular and Molecular Biology, Council of Scientific and Industrial Research, Uppal Road, Hyderabad 500 007, India.
authors.forename: Madhavi
authors.initials: M
authors.lastname: Muppirala
authors.forename: Vijay
authors.initials: V
authors.lastname: Gupta
authors.forename: Ghanshyam
authors.initials: G
authors.lastname: Swarup
mesh_terms: D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism*; D002522:Chlorocebus aethiops; D016313:Genes, abl / Q000502:physiology*; D006801:Humans; D047468:Immunoprecipitation; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation; D021381:Protein Transport; D054578:Protein Tyrosine Phosphatase, Non-Receptor Type 2 / Q000235:genetics / Q000378:metabolism*; D050765:Qa-SNARE Proteins / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D055571:Secretory Pathway / Q000502:physiology*; D020798:Two-Hybrid System Techniques; D014443:Tyrosine / Q000378:metabolism*
chemical_list: D050765:Qa-SNARE Proteins; D014443:Tyrosine; D054578:Protein Tyrosine Phosphatase, Non-Receptor Type 2