Transmembrane domain length determines intracellular membrane compartment localization of syntaxins 3, 4, and 5.
Insulin recruits glucose transporter 4 (GLUT-4) vesicles from intracellular stores to the plasma membrane in muscle and adipose tissue by specific interactions between the vesicle membrane-soluble N-ethylmaleimide-sensitive factor attachment protein target receptor (SNARE) protein VAMP-2 and the target membrane SNARE protein syntaxin 4. Although GLUT-4 vesicle trafficking has been intensely studied, few have focused on the mechanism by which the SNAREs themselves localize to specific membrane compartments. We therefore set out to identify the molecular determinants for localizing several syntaxin isoforms, including syntaxins 3, 4, and 5, to their respective intracellular compartments (plasma membrane for syntaxins 3 and 4; cis-Golgi for syntaxin 5). Analysis of a series of deletion and chimeric syntaxin constructs revealed that the 17-amino acid transmembrane domain of syntaxin 5 was sufficient to direct the cis-Golgi localization of several heterologous reporter constructs. In contrast, the longer 25-amino acid transmembrane domain of syntaxin 3 was sufficient to localize reporter constructs to the plasma membrane. Furthermore, truncation of the syntaxin 3 transmembrane domain to 17 amino acids resulted in a complete conversion to cis-Golgi compartmentalization that was indistinguishable from syntaxin 5. These data support a model wherein short transmembrane domains (< or =17 amino acids) direct the cis-Golgi localization of syntaxins, whereas long transmembrane domains (> or =23 amino acids) direct plasma membrane localization.
Watson
R T
RT
Department of Physiology and Biophysics, The University of Iowa, Iowa City, Iowa 52242, USA.
Pessin
J E
JE
D016475:3T3 Cells; D017667:Adipocytes; D000595:Amino Acid Sequence; D000818:Animals; D020126:Brefeldin A / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism; D002451:Cell Compartmentation; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D017930:Genes, Reporter / Q000235:genetics; D006056:Golgi Apparatus / Q000378:metabolism; D000077614:Golgi Matrix Proteins; D006360:Heat-Shock Proteins; D007425:Intracellular Membranes / Q000737:chemistry / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008856:Microscopy, Fluorescence; D018832:Molecular Chaperones / Q000378:metabolism; D008969:Molecular Sequence Data; D020033:Protein Isoforms / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D050765:Qa-SNARE Proteins; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D050600:SNARE Proteins; D016415:Sequence Alignment; D033921:Vesicular Transport Proteins
D002352:Carrier Proteins; D000091342:Endoplasmic Reticulum Chaperone BiP; D000077614:Golgi Matrix Proteins; D006360:Heat-Shock Proteins; D008565:Membrane Proteins; D018832:Molecular Chaperones; D020033:Protein Isoforms; D011500:Protein Synthesis Inhibitors; D050765:Qa-SNARE Proteins; D011993:Recombinant Fusion Proteins; D050600:SNARE Proteins; D033921:Vesicular Transport Proteins; C087574:macrogolgin; D020126:Brefeldin A