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Subcellular distribution and function of Rab3A, B, C, and D isoforms in insulin-secreting cells

Escher, G
Baldini, G
Darchen, F
Published in Molecular Endocrinology. 1999, vol. 13, no. 2, p. 202-12
Abstract Insulin-secreting cells express four GTPases of the Rab3 family. After separation of extracts of INS-1 cells on a sucrose density gradient, the bulk of the A, B, and C isoforms was recovered in the fractions enriched in insulin-containing secretory granules. Rab3D was also mainly associated with secretory granules, but a fraction of this isoform was localized on lighter organelles. Analyses by confocal microscopy of immunostained HIT-T15 cells transfected with epitope-tagged constructs confirmed the distribution of the Rab3 isoforms. Transfection of HIT-T15 cells with GTPase-deficient mutants of the Rab3 isoforms decreased nutrient-induced insulin release to different degrees (D>B>A>C), while overexpression of Rab3 wild types had minor or no effects. Expression of the same Rab3 mutants in PC12 cells provoked an inhibition of K+-stimulated secretion of dense core vesicles, indicating that, in beta-cells and neuroendocrine cells, the four Rab3 isoforms play a similar role in exocytosis. A Rab3A/C chimera in which the carboxyterminal domain of A was replaced with the corresponding region of C inhibited insulin secretion as Rab3A. In contrast, a Rab3C/A chimera containing the amino-terminal domain of C was less potent and reduced exocytosis as Rab3C. This suggests that the degree of inhibition obtained after transfection of the Rab3 isoforms is determined by differences in the variable amino-terminal region.
Keywords AnimalsBlotting, WesternCells, CulturedCentrifugation, Density GradientCytoplasmic Granules/secretionDensitometryEnzyme-Linked Immunosorbent AssayExocytosisGTP-Binding Proteins/metabolismHumansImage Processing, Computer-AssistedInsulin/secretionIslets of Langerhans/cytology/secretionMicroscopy, FluorescenceMutagenesis, Site-DirectedPC12 CellsRatsRecombinant Fusion Proteins/metabolismTransfectionRab3 GTP-Binding Proteins
PMID: 9973251
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IEZZI-BAKHTIARI, Mariella et al. Subcellular distribution and function of Rab3A, B, C, and D isoforms in insulin-secreting cells. In: Molecular Endocrinology, 1999, vol. 13, n° 2, p. 202-12. doi: 10.1210/mend.13.2.0228 https://archive-ouverte.unige.ch/unige:35090

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Deposited on : 2014-03-28

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