Indication for differential sorting of the rat v-SNARE splice isoforms VAMP-1a and -1b.
Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins are essential constituents of the intracellular trafficking machinery. The variable C-terminus in the 2 rat VAMP-1 splice isoforms VAMP-1a and -1b potentially acts as a sorting signal, because similar changes at the C-terminal end of a human VAMP-1 splice isoform resulted in its sorting to mitochondria. To evaluate the differences in the subcellular localization of these two v-SNARE proteins, VAMP-1a and -1b proteins tagged with green fluorescent protein (GFP) and red fluorescent protein (RFP) were expressed in HeLa, COS-7, and MDCK cells and evaluated by conventional confocal as well as total internal reflection fluorescence microscopy. Regions consistent with the endoplasmic reticulum and Golgi apparatus demonstrated a major overlap of both signals. In the periphery, vesicular structures were observed that mainly expressed one of the 2 isoforms. Within our experimental settings, we could not observe sorting of any of the 2 isoforms to mitochondria or peroxisomes, whereas both isoforms were found expressed in a minor subset of singular vesicles, which sporadically appeared to co-localize with the exocyst marker EXOC3/Sec6. Because vesicular structures were seen that expressed only one of the two splice variants, it is possible that VAMP-1a and VAMP-1b are sorted to distinct cellular compartments that require further characterization.
a Department of Otorhinolaryngology, Head and Neck Surgery, University Hospital Giessen and Marburg, Campus Marburg, 3.BA, Room +3/08070, Baldingerstrasse, D-35033 Marburg, Germany.
Fiona R
FR
Rodepeter
a Department of Otorhinolaryngology, Head and Neck Surgery, University Hospital Giessen and Marburg, Campus Marburg, 3.BA, Room +3/08070, Baldingerstrasse, D-35033 Marburg, Germany.
Susanne
S
Wiegand
b Department of Cell Biology, Institute of Anatomy and Cell Biology, Philipps-Universität, Marburg, Germany.
Hans-Georg
HG
Lüers
c Department of Medical Cell Biology, Philipps-Universität, Marburg, Germany.
Gabriel A
GA
Bonaterra
d Department of Medicine, Stanford University, Stanford, CA, USA.
Anson W
AW
Lowe
e Department of Molecular Neuroscience, Institute of Anatomy and Cell Biology, Philipps-Universität, Marburg, Germany.
Michael
M
Bette
f Institute of Cell Biology and Cell Pathology, Philipps-Universität, Marburg, Germany.
Ralf
R
Jacob
a Department of Otorhinolaryngology, Head and Neck Surgery, University Hospital Giessen and Marburg, Campus Marburg, 3.BA, Room +3/08070, Baldingerstrasse, D-35033 Marburg, Germany.
Robert
R
Mandic
D000818:Animals; D006801:Humans; D008856:Microscopy, Fluorescence; D020033:Protein Isoforms / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D051381:Rats; D014407:Tumor Cells, Cultured; D050987:Vesicle-Associated Membrane Protein 1 / Q000032:analysis / Q000235:genetics* / Q000378:metabolism*
D020033:Protein Isoforms; D050987:Vesicle-Associated Membrane Protein 1
SNARE; VAMP; compartiment subcellulaire; intracellular trafficking; sorting; subcellular compartment; trafic intracellulaire; triage