Phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] plays a complex role in generating intracellular signalling molecules, and also in regulating actin-binding proteins, vesicular trafficking and vacuolar fusion. Four inositol polyphosphate 5-phosphatases (hereafter called 5-phosphatases) have been identified in Saccharomyces cerevisiae: Inp51p, Inp52p, Inp53p and Inp54p. Each enzyme contains a 5-phosphatase domain which hydrolyses PtdIns(4,5)P2, forming PtdIns4P, while Inp52p and Inp53p also express a polyphosphoinositide phosphatase domain within the Sac1-like domain. Disruption of any two yeast 5-phosphatases containing a Sac1-like domain results in abnormalities in actin polymerization, plasma membrane, vacuolar morphology and bud-site selection. Triple null mutant 5-phosphatase strains are non-viable. To investigate the role of PtdIns(4,5)P2 in mediating the phenotype of double and triple 5-phosphatase null mutant yeast, we determined whether a mammalian PtdIns(4,5)P2 5-phosphatase, 5-phosphatase II, which lacks polyphosphoinositide phosphatase activity, could correct the phenotype of triple 5-phosphatase null mutant yeast and restore cellular PtdIns(4,5)P2 levels to near basal values. Mammalian 5-phosphatase II expressed under an inducible promoter corrected the growth, cell wall, vacuolar and actin polymerization defects of the triple 5-phosphatase null mutant yeast strains. Cellular PtdIns(4,5)P2 levels in various 5-phosphatase double null mutant strains demonstrated significant accumulation (4.5-, 3- and 2-fold for ∆inp51∆inp53, ∆inp51∆inp52 and ∆inp52∆inp53 double null mutants respectively), which was corrected significantly following 5-phosphatase II expression. Collectively, these studies demonstrate the functional and cellular consequences of PtdIns(4,5)P2 accumulation and the evolutionary conservation of function between mammalian and yeast PtdIns(4,5)P2 5-phosphatases.
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Research Article|
April 24 2001
Mammalian inositol polyphosphate 5-phosphatase II can compensate for the absence of all three yeast Sac1-like-domain-containing 5-phosphatases
Cindy J. O'MALLEY;
Cindy J. O'MALLEY
1
*Department of Biochemistry and Molecular Biology, Monash University, Melbourne, Victoria 3800, Australia
1To whom correspondence should be sent, at present address: Lymphocyte Signalling and Development Laboratory, Molecular Immunology Programme, The Babraham Institute, Babraham, Cambridge CB2 4AT, U.K. (e-mail cindy.omalley@bbsrc.ac.uk).
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Brad K. McCOLL;
Brad K. McCOLL
†Sir Donald and Lady Trescowthick Research Laboratories, Peter MacCallum Institute, St Andrews Place, Melbourne Victoria 3000, Australia
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Anne M. KONG;
Anne M. KONG
*Department of Biochemistry and Molecular Biology, Monash University, Melbourne, Victoria 3800, Australia
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Sarah L. ELLIS;
Sarah L. ELLIS
†Sir Donald and Lady Trescowthick Research Laboratories, Peter MacCallum Institute, St Andrews Place, Melbourne Victoria 3000, Australia
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A. Primrose W. WIJAYARATNAM;
A. Primrose W. WIJAYARATNAM
†Sir Donald and Lady Trescowthick Research Laboratories, Peter MacCallum Institute, St Andrews Place, Melbourne Victoria 3000, Australia
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Joe SAMBROOK;
Joe SAMBROOK
†Sir Donald and Lady Trescowthick Research Laboratories, Peter MacCallum Institute, St Andrews Place, Melbourne Victoria 3000, Australia
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Christina A. MITCHELL
Christina A. MITCHELL
*Department of Biochemistry and Molecular Biology, Monash University, Melbourne, Victoria 3800, Australia
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Publisher: Portland Press Ltd
Received:
October 25 2000
Revision Received:
January 11 2001
Accepted:
February 21 2001
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London © 2001
2001
Biochem J (2001) 355 (3): 805–817.
Article history
Received:
October 25 2000
Revision Received:
January 11 2001
Accepted:
February 21 2001
Citation
Cindy J. O'MALLEY, Brad K. McCOLL, Anne M. KONG, Sarah L. ELLIS, A. Primrose W. WIJAYARATNAM, Joe SAMBROOK, Christina A. MITCHELL; Mammalian inositol polyphosphate 5-phosphatase II can compensate for the absence of all three yeast Sac1-like-domain-containing 5-phosphatases. Biochem J 1 May 2001; 355 (3): 805–817. doi: https://doi.org/10.1042/bj3550805
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