betaPIX-activated Rac1 stimulates the activation of phospholipase D, which is associated with exocytosis in neuroendocrine cells.
Rho GTPases are crucial regulators of actin cytoskeletal rearrangements and play important roles in many cell functions linked to membrane trafficking processes. In neuroendocrine cells, we have previously demonstrated that RhoA and Cdc42 mediate part of the actin remodelling and vesicular trafficking events that are required for the release of hormones by exocytosis. Here, we investigate the functional importance of Rac1 for the exocytotic reaction and dissect the downstream and upstream molecular events that might integrate it to the exocytotic machinery. Using PC12 cells, we found that Rac1 is associated with the plasma membrane and is activated during exocytosis. Silencing of Rac1 by siRNA inhibits hormone release, prevents secretagogue (high K(+))-evoked phospholipase D1 (PLD1) activation and blocks the formation of phosphatidic acid at the plasma membrane. We identify betaPix as the guanine nucleotide-exchange factor integrating Rac1 activation to PLD1 and the exocytotic process. Finally, we show that the presence of the scaffolding protein Scrib at the plasma membrane is essential for betaPix/Rac1-mediated PLD1 activation and exocytosis. As PLD1 has recently emerged as a promoter of membrane fusion in various exocytotic events, our results define a novel molecular pathway linking a Rho GTPase, Rac1, to the final stages of Ca(2+)-regulated exocytosis in neuroendocrine cells.
Momboisse
Fanny
F
Département Neurotransmission et Sécrétion Neuroendocrine, Institut des Neurosciences Cellulaires et Intégratives (UPR 3212), Centre National de la Recherche Scientifique et Université de Strasbourg, 5 rue Blaise Pascal, 67084 Strasbourg, France.
Lonchamp
Etienne
E
Calco
Valerie
V
Ceridono
Mara
M
Vitale
Nicolas
N
Bader
Marie-France
MF
Gasman
Stéphane
S
D000818:Animals; D055162:Biocatalysis / Q000187:drug effects; D002118:Calcium / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D005089:Exocytosis / Q000187:drug effects; D013006:Growth Hormone / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D055099:Neuroendocrine Cells / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology*; D016716:PC12 Cells; D010712:Phosphatidic Acids / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D034622:RNA Interference / Q000187:drug effects; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D064067:Rho Guanine Nucleotide Exchange Factors; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*
D020662:Guanine Nucleotide Exchange Factors; D047908:Intracellular Signaling Peptides and Proteins; D010712:Phosphatidic Acids; D034741:RNA, Small Interfering; D064067:Rho Guanine Nucleotide Exchange Factors; D013006:Growth Hormone; D010739:Phospholipase D; C106801:phospholipase D1; D020830:rac1 GTP-Binding Protein; D002118:Calcium