9402469

pubmed:Citation

3

Rabphilin 3a is a Rab 3-GTP binding protein concentrated on secretory vesicles of neurons and endocrine cells. There is evidence that rabphilin 3a undergoes cycles of association-dissociation with membranes and that recruitment of rabphilin 3a to secretory vesicles is mediated by Rab 3a, suggesting that rabphilin 3a is a downstream effector of this Rab. In this study we have investigated whether a membrane-anchored form of rabphilin 3a mimics the action of rabphilin 3a on secretion and bypasses the need for Rab 3 function. Overexpression of both wild-type rabphilin 3a and of a transmembrane anchored form of rabphilin 3a stimulated (about 2-fold) evoked secretion of coexpressed human proinsulin from clonal HIT-T15 cells. A similar transmembrane-anchored protein which lacked the Rab 3 binding region stimulated secretion even more effectively. Unexpectedly, a rabphilin 3a deletion mutant missing the Rab 3 binding domain was also stimulatory on secretion, although a further deletion of rabphilin to exclude the first of the two proline-rich regions abolished its stimulatory effect. The first of these two mutants was primarily particulate, while the second mutant was primarily soluble, suggesting that the first proline-rich region of rabphilin 3a plays a role in targeting rabphilin to its site of action. We conclude that the action of rabphilin 3a can be independent of Rab 3 if other mechanisms produce a sufficient concentration of the protein in proximity of exocytotic sites. These results provide new evidence for a fundamental similarity in the mechanisms by which Ras and Rab GTPase produce their distinct physiological effects.

eng

IM

MEDLINE

0171-9335

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NLM

209-16

pubmed-meshheading:9402469-Adaptor Proteins, Signal Transducing, pubmed-meshheading:9402469-Amino Acid Sequence, pubmed-meshheading:9402469-Animals, pubmed-meshheading:9402469-CHO Cells, pubmed-meshheading:9402469-Calcium-Binding Proteins, pubmed-meshheading:9402469-Cell Line, pubmed-meshheading:9402469-Cricetinae, pubmed-meshheading:9402469-Exocytosis, pubmed-meshheading:9402469-GTP-Binding Proteins, pubmed-meshheading:9402469-Humans, pubmed-meshheading:9402469-Membrane Glycoproteins, pubmed-meshheading:9402469-Molecular Sequence Data, pubmed-meshheading:9402469-Nerve Tissue Proteins, pubmed-meshheading:9402469-Proinsulin, pubmed-meshheading:9402469-Protein Precursors, pubmed-meshheading:9402469-Rats, pubmed-meshheading:9402469-Recombinant Fusion Proteins, pubmed-meshheading:9402469-Subcellular Fractions, pubmed-meshheading:9402469-Synaptotagmins, pubmed-meshheading:9402469-Vesicular Transport Proteins, pubmed-meshheading:9402469-rab GTP-Binding Proteins, pubmed-meshheading:9402469-rab3 GTP-Binding Proteins

The stimulatory effect of rabphilin 3a on regulated exocytosis from insulin-secreting cells does not require an association-dissociation cycle with membranes mediated by Rab 3.

Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't