Synaptotagmin I and IV define distinct populations of neuronal transport vesicles.
Mammalian synaptotagmins constitute a multigene family of at least 11 membrane proteins. We have characterized synaptotagmin IV using antibodies directed against the C2A domain of the protein. Antibodies reacted specifically with a protein band that migrated as a 41-44 kDa doublet. Synaptotagmin IV expression was regulated throughout development. A strong decrease in the amount detected by Western blotting occurred between postnatal day 5 and adulthood, in agreement with studies on the expression of synaptotagmin IV transcripts. In subcellular fractionation, synaptotagmin IV was not detected in the synaptic vesicle-enriched fraction. Immunofluorescence microscopy was concordant with this finding. In 6-day-old rat cerebellum and cultured hippocampal neurons the subcellular distribution of synaptotagmin IV was clearly different from that of synaptotagmin I. Synaptotagmin IV displayed a punctate non-polarized distribution on neuronal extensions, whereas synaptotagmin I staining was essentially synaptic. Synaptotagmin IV staining was also observed in the soma in strong perinuclear fluorescent puncta superimposed on that of Golgi/TGN markers. Furthermore, synaptotagmin IV was seen in the proximal part of the growth cone domain and not in the microfilament-rich region which includes filopodia. Co-localizations with the adhesion molecules vinculin and zyxin at the proximal part of growth cones were observed. Synaptotagmin IV may thus be involved in the regulation of specific membrane-trafficking pathways during brain development.
Berton
F
F
INSERM U 464, Institut Fédératif Jean Roche, Université de la Méditerranée, Faculté de Médecine, Marseille Cedex 20, France.
Cornet
V
V
Iborra
C
C
Garrido
J
J
Dargent
B
B
Fukuda
M
M
Seagar
M
M
Marquèze
B
B
D000818:Animals; D000906:Antibodies; D015153:Blotting, Western; D016466:CHO Cells; D002135:Calcium-Binding Proteins; D002451:Cell Compartmentation / Q000502:physiology; D006224:Cricetinae; D003593:Cytoplasm / Q000737:chemistry; D017930:Genes, Reporter; D006056:Golgi Apparatus / Q000737:chemistry / Q000378:metabolism; D049452:Green Fluorescent Proteins; D020439:Growth Cones / Q000737:chemistry / Q000378:metabolism; D006624:Hippocampus / Q000166:cytology; D007202:Indicators and Reagents / Q000378:metabolism; D007425:Intracellular Membranes / Q000737:chemistry* / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics; D008562:Membrane Glycoproteins / Q000032:analysis* / Q000276:immunology; D008667:Metalloproteins / Q000032:analysis / Q000276:immunology; D009419:Nerve Tissue Proteins / Q000032:analysis* / Q000276:immunology; D009474:Neurons / Q000737:chemistry* / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D013347:Subcellular Fractions / Q000737:chemistry; D013572:Synaptic Vesicles / Q000737:chemistry* / Q000378:metabolism; D050863:Synaptotagmin I; D050857:Synaptotagmins; D014162:Transfection; D016596:Vinculin / Q000032:analysis / Q000276:immunology
D000906:Antibodies; D002135:Calcium-Binding Proteins; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D008562:Membrane Glycoproteins; D008667:Metalloproteins; D009419:Nerve Tissue Proteins; D050863:Synaptotagmin I; C494312:Syt1 protein, rat; C494294:Syt4 protein, rat; D016596:Vinculin; D050857:Synaptotagmins; D049452:Green Fluorescent Proteins