Regulation and recruitment of phosphatidylinositol 4-kinase on immature secretory granules is independent of ADP-ribosylation factor 1.
Heterotrimeric G-proteins, as well as small GTPases of the Rho and ADP-ribosylation factor (ARF) family, are implicated in the regulation of lipid kinases, including PtdIns 4-kinases and PtdIns(4)P 5-kinases. Here, we describe a PtdIns 4-kinase activity on immature secretory granules (ISGs), regulated secretory organelles formed from the trans-Golgi network (TGN), and investigate the regulation of PtdIns4P levels on these membranes. Over 50% of the PtdIns 4-kinase activity on ISGs is inhibited by both a low concentration of adenosine and the monoclonal antibody 4C5G, a specific inhibitor of the type II PtdIns 4-kinase. Treatment of ISGs with mastoparan 7 (M7) stimulates the type II PtdIns 4-kinase via pertussis-toxin-sensitive G(i)/G(0) proteins, which, in contrast with previous results obtained with chromaffin granules [Gasman, Chasserot-Golaz, Hubert, Aunis and Bader (1998) J. Biol. Chem. 273, 16913-16920], does not require Rho A, B or C. M7 treatment also leads to an inhibition in the recruitment of ARF to ISG membranes: this inhibition is not dependent on G(i)/G(0) activation, and is not linked to the stimulation of PtdIns 4-kinase observed with M7. PtdIns 4-kinase activity on ISGs is not regulated by myristoylated ARF1-GTP, in contrast with results obtained with Golgi membranes [Godi, Pertile, Meyers, Marra, Di Tullio, Iurisci, Luini, Corda and De Matteis (1999) Nat. Cell Biol. 1, 280-287; Jones, Morris, Morgan, Kondo, Irvine and Cockcroft (2000) J. Biol. Chem. 275, 13962-13170], whereas ARF1-GTP does regulate the production of PtdIns(4,5)P(2). Our results suggest that the regulation of PtdIns 4-kinase on the ISGs differs in comparison with that on the TGN, and might be related to a specific requirement of ISG maturation.
Panaretou
Christina
C
The Secretory Pathways Laboratory, Cancer Research UK London Institute, 44 Lincoln's Inn Fields, London WC2A 3PX, U.K.
Tooze
Sharon A
SA
D019870:1-Phosphatidylinositol 4-Kinase / Q000378:metabolism*; D020823:ADP-Ribosylation Factor 1 / Q000378:metabolism*; D000818:Animals; D019204:GTP-Binding Proteins / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins; D016716:PC12 Cells; D010455:Peptides; D037342:Pertussis Toxin; D051381:Rats; D022142:Secretory Vesicles / Q000187:drug effects / Q000201:enzymology*; D018808:Transcription Factor AP-1 / Q000378:metabolism; D010566:Virulence Factors, Bordetella / Q000494:pharmacology; D014862:Wasp Venoms / Q000494:pharmacology; D021601:trans-Golgi Network / Q000201:enzymology
D036341:Intercellular Signaling Peptides and Proteins; D010455:Peptides; D018808:Transcription Factor AP-1; D010566:Virulence Factors, Bordetella; D014862:Wasp Venoms; C037500:mastoparan; D037342:Pertussis Toxin; D019870:1-Phosphatidylinositol 4-Kinase; D019204:GTP-Binding Proteins; D020823:ADP-Ribosylation Factor 1