Nonclassical PITPs activate PLD via the Stt4p PtdIns-4-kinase and modulate function of late stages of exocytosis in vegetative yeast.
Phospholipase D (PLD) is a PtdCho-hydrolyzing enzyme that plays central signaling functions in eukaryotic cells. We previously demonstrated that action of a set of four nonclassical and membrane-associated Sec14p-like phosphatidylinositol transfer proteins (PITPs) is required for optimal activation of yeast PLD in vegetative cells. Herein, we focus on mechanisms of Sfh2p and Sfh5p function in this regulatory circuit. We describe several independent lines of in vivo evidence to indicate these SFH PITPs regulate PLD by stimulating PtdIns-4,5-P2 synthesis and that this stimulated PtdIns-4,5-P2 synthesis couples to action of the Stt4p PtdIns 4-kinase. Furthermore, we provide genetic evidence to suggest that specific subunits of the yeast exocyst complex (i.e. a component of the plasma membrane vesicle docking machinery) and the Sec9p plasma membrane t-SNARE are regulated by PtdIns(4,5)P2 and that Sfh5p helps regulate this interface in vivo. The collective in vivo and biochemical data suggest SFH-mediated stimulation of Stt4p activity is indirect, most likely via a substrate delivery mechanism.
Routt
Sheri M
SM
Department of Cell and Developmental Biology, Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7090, USA.
Ryan
Margaret M
MM
Tyeryar
Kimberly
K
Rizzieri
Kellie E
KE
Mousley
Carl
C
Roumanie
Olivier
O
Brennwald
Patrick J
PJ
Bankaitis
Vytas A
VA
D019870:1-Phosphatidylinositol 4-Kinase / Q000502:physiology*; D000199:Actins / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005089:Exocytosis / Q000502:physiology*; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D010716:Phosphatidylinositols / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D049294:Phospholipid Transfer Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D010770:Phosphotransferases / Q000096:biosynthesis / Q000235:genetics; D017853:Phosphotransferases (Alcohol Group Acceptor); D050767:Qc-SNARE Proteins / Q000096:biosynthesis / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000378:metabolism* / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*
D000199:Actins; C483430:CSR1 protein, S cerevisiae; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols; D049294:Phospholipid Transfer Proteins; D050767:Qc-SNARE Proteins; C483438:SEC14 protein, S cerevisiae; C090117:SEC9 protein, S cerevisiae; C483439:SFH5 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); D019870:1-Phosphatidylinositol 4-Kinase; C112465:STT4 protein, S cerevisiae; C489795:MSS4 protein, S cerevisiae; D010739:Phospholipase D