rab3A attachment to the synaptic vesicle membrane mediated by a conserved polyisoprenylated carboxy-terminal sequence.
rab3A is a small neuronal GTP-binding protein specifically localized to synaptic vesicles. Membrane-bound rab3A behaves like an intrinsic membrane protein in vitro, but reversibly dissociates from synaptic vesicles after exocytosis in vivo. Here we demonstrate that rab3A is attached to synaptic vesicle membranes by a carboxy-terminal Cys-X-Cys sequence that is posttranslationally modified. This modification is inhibited by compactin in a mevalonate-dependent manner, suggesting that the Cys-X-Cys sequence represents a novel polyisoprenylation sequence. Isolation of a rab3 homolog from D. melanogaster reveals high evolutionary conservation of rab3A, including its carboxy-terminal Cys-X-Cys sequence. The posttranslational modifications of soluble and membrane-bound rab3A are biochemically different, but both require the carboxy-terminal Cys-X-Cys sequence and are faithfully reproduced in nonneuronal cells. Our results suggest that the carboxy-terminal Cys-X-Cys sequence of rab3A is polyisoprenylated and is used as its regulatable membrane anchor. Furthermore, the hydrophobic modification of rab3A and its correct intracellular targeting to synaptic vesicles are independent, presumably consecutive events.
Johnston
P A
PA
Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas 75235.
Archer
B T
BT
Robinson
K
K
Mignery
G A
GA
Jahn
R
R
Südhof
T C
TC
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D004331:Drosophila melanogaster / Q000235:genetics; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D008148:Lovastatin / Q000031:analogs & derivatives / Q000494:pharmacology; D008798:Mevalonic Acid / Q000494:pharmacology; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014176:Protein Biosynthesis; D012689:Sequence Homology, Nucleic Acid; D013572:Synaptic Vesicles / Q000378:metabolism*; D020692:rab3 GTP-Binding Proteins
D009419:Nerve Tissue Proteins; C012258:mevastatin; D008148:Lovastatin; D019204:GTP-Binding Proteins; D020692:rab3 GTP-Binding Proteins; D008798:Mevalonic Acid