Synaptobrevin 2 is palmitoylated in synaptic vesicles prepared from adult, but not from embryonic brain.
Neuronal SNARE-proteins such as synaptobrevin, SNAP 25, and synaptotagmin are key players during neurosecretion. So far palmitoylation of SNAP-25 and synaptotagmin 1 have been described in vivo. Here we have analyzed palmitoylation of the SNARE-proteins synaptobrevin 2 and synaptotagmin in vitro using synaptosomal and synaptic vesicle preparations from rat brain. Labeling of synaptic vesicles prepared from adult brain with [3H]palmitate revealed synaptobrevin 2 besides synaptotagmin 1 as major palmitoylated proteins. [3H]Palmitoylation of synaptobrevin 2 was resistant to chloroform/methanol extraction, but sensitive to reducing agents indicating a covalent fatty acid bond to cysteine residues. Palmitoylation of synaptobrevin 2 was also confirmed using endogenous synaptobrevin 2 present in PC-12 cells and synaptobrevin 2 expressed with a vacciniavirus system in Cos cells. In contrast to the situation seen with membrane preparations obtained from adult brain, synaptic vesicles prepared from embryonic rat brain did not support [3H]palmitoylation of synaptobrevin and synaptotagmin. These results suggest, that both synaptobrevin 2 and synaptotagmin were efficiently palmitoylated from mature synaptic vesicles. However, at least one component of the palmitoylation machinery is developmentally upregulated.
Veit
M
M
Department of Immunology and Molecular Biology, Free University, Berlin, Germany. mveit@city.vetmed.fu-berlin.de
Becher
A
A
Ahnert-Hilger
G
G
D000367:Age Factors; D000818:Animals; D001921:Brain / Q000166:cytology / Q000196:embryology* / Q000378:metabolism*; D019556:COS Cells; D002135:Calcium-Binding Proteins; D005333:Fetus / Q000166:cytology; D008562:Membrane Glycoproteins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D016716:PC12 Cells; D019308:Palmitic Acid / Q000378:metabolism*; D050683:R-SNARE Proteins; D051381:Rats; D050600:SNARE Proteins; D013572:Synaptic Vesicles / Q000378:metabolism*; D013574:Synaptosomes / Q000378:metabolism; D050863:Synaptotagmin I; D050857:Synaptotagmins; D033921:Vesicular Transport Proteins
D002135:Calcium-Binding Proteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D050683:R-SNARE Proteins; D050600:SNARE Proteins; D050863:Synaptotagmin I; C494312:Syt1 protein, rat; D033921:Vesicular Transport Proteins; D050857:Synaptotagmins; D019308:Palmitic Acid