Inactivation of the Phosphoinositide Phosphatases Sac1p and Inp54p Leads to Accumulation of Phosphatidylinositol 4,5-Bisphosphate on Vacuole Membranes and Vacuolar Fusion Defects

Fenny Wiradjaja; Lisa M. Ooms; Sabina Tahirovic; Ellie Kuhne; Rodney J. Devenish; Alan L. Munn; Robert C. Piper; Peter Mayinger; Christina A. Mitchell

doi:10.1074/jbc.M701038200

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Inactivation of the Phosphoinositide Phosphatases Sac1p and Inp54p Leads to Accumulation of Phosphatidylinositol 4,5-Bisphosphate on Vacuole Membranes and Vacuolar Fusion Defects

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Inactivation of the Phosphoinositide Phosphatases Sac1p and Inp54p Leads to Accumulation of Phosphatidylinositol 4,5-Bisphosphate on Vacuole Membranes and Vacuolar Fusion Defects

Fenny Wiradjaja, Lisa M. Ooms, Sabina Tahirovic, Ellie Kuhne, Rodney J. Devenish, Alan L. Munn, Robert C. Piper, Peter Mayinger and Christina A. Mitchell

The Journal of biological chemistry, Vol.282(22), pp.16295-16307

06/01/2007

DOI: 10.1074/jbc.M701038200

PMID: 17392273

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Abstract

Phosphoinositides direct membrane trafficking, facilitating the recruitment of effectors to specific membranes. In yeast phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) isproposed to regulate vacuolar fusion; however, in intact cells this phosphoinositide can only be detected at the plasma membrane. In Saccharomyces cerevisiae the 5-phosphatase, Inp54p, dephosphorylates PtdIns(4,5)P2 forming PtdIns(4)P, a substrate for the phosphatase Sac1p, which hydrolyzes (PtdIns(4)P). We investigated the role these phosphatases in regulating PtdIns(4,5)P2 subcellular distribution. PtdIns(4,5)P2 bioprobes exhibited loss of plasma membrane localization and instead labeled a subset of fragmented vacuoles in Δsac1 Δinp54 and sac1ts Δinp54 mutants. Furthermore, sac1ts Δinp54 mutants exhibited vacuolar fusion defects, which were rescued by latrunculin A treatment, or by inactivation of Mss4p, a PtdIns(4)P 5-kinase that synthesizes plasma membrane PtdIns(4,5)P2. Under these conditions PtdIns(4,5)P2 was not detected on vacuole membranes, and vacuole morphology was normal, indicating vacuolar PtdIns(4,5)P2 derives from Mss4p-generated plasma membrane PtdIns(4,5)P2. Δsac1 Δinp54 mutants exhibited delayed carboxypeptidase Y sorting, cargo-selective secretion defects, and defects in vacuole function. These studies reveal PtdIns(4,5)P2 hydrolysis by lipid phosphatases governs its spatial distribution, and loss of phosphatase activity may result in PtdIns(4,5)P2 accumulation on vacuole membranes leading to vacuolar fragmentation/fusion defects.

Details

Title: Subtitle: Inactivation of the Phosphoinositide Phosphatases Sac1p and Inp54p Leads to Accumulation of Phosphatidylinositol 4,5-Bisphosphate on Vacuole Membranes and Vacuolar Fusion Defects
Creators: Fenny Wiradjaja - Monash University
Lisa M. Ooms - Monash University
Sabina Tahirovic - Heidelberg University
Ellie Kuhne - Department of Biochemistry and Molecular Biology, Monash University, Clayton 3800, Victoria, Australia
Rodney J. Devenish - Monash University
Alan L. Munn - University of Queensland
Robert C. Piper - University of Iowa
Peter Mayinger - Oregon Health & Science University
Christina A. Mitchell - Monash University
Resource Type: Journal article
Publication Details: The Journal of biological chemistry, Vol.282(22), pp.16295-16307
DOI: 10.1074/jbc.M701038200
PMID: 17392273
NLM abbreviation: J Biol Chem
ISSN: 0021-9258
eISSN: 1083-351X
Publisher: Elsevier Inc
Language: English
Date published: 06/01/2007
Academic Unit: Molecular Physiology and Biophysics; Medicine Administration; Internal Medicine
Record Identifier: 9984297506102771

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