Phosphoinositide-specific Inositol Polyphosphate 5-Phosphatase IV Inhibits Akt/Protein Kinase B Phosphorylation and Leads to Apoptotic Cell Death

Phosphoinositide-specific inositol polyphosphate 5- phosphatase IV has the affinity for PI(3,4,5)P3 (K m = 0.65 μM) that is approximately 10-fold greater than the other inositol polyphosphate 5-phosphatases, which use this substrate including SHIP, OCRL, and 5ptase II, suggesting that it may be important in controlling intracellular levels of this metabolite. We created cell lines stably expressing the enzyme to study its effect on cell function. We found that overexpression of 5ptase IV in 293 cells caused the rapid depletion of both PI(4,5)P2 and PI(3,4,5)P3 in cells with corresponding increases in the products, PI(4)P and PI(3,4)P2, changing the balance of two phosphoinositol products of phosphoinositide 3-kinase, PI(3,4)P2 and PI(3,4,5)P3, in the cell. One of the targets of these phosphoinositides is the serine/threonine kinase Akt, which plays an important role in the control of apoptosis. We were able to address the relative roles of PI(3,4)P2 and PI(3,4,5)P3 in the activation of Akt by selective depletion of these phosphoinositides in cells stably transfected with 5ptase IV and inositol polyphosphate 4-phosphatase (4ptase I). In cells transfected with 4ptase I, the level of PI(3,4)P2 was reduced, and PI(3,4,5)P3 was increased. Expression of the two enzymes had the opposite effect on the phosphorylation of Akt in response to stimulation with growth factors or heat shock. Akt phosphorylation was inhibited in cells expressing 5ptase IV but increased in 4ptase I cells and correlated with the intracellular level of PI(3,4,5)P3 and not that of PI(3,4)P2. The inhibition of Akt phosphorylation in cells expressing 5ptase IV makes them highly susceptible to FAS-induced apoptosis, whereas overexpressing of the 4ptase I protects cells from apoptosis. Our results place 5ptase IV as a relevant biological regulator of PI3K/Akt pathway in cells. Phosphoinositide-specific inositol polyphosphate 5- phosphatase IV has the affinity for PI(3,4,5)P3 (K m = 0.65 μM) that is approximately 10-fold greater than the other inositol polyphosphate 5-phosphatases, which use this substrate including SHIP, OCRL, and 5ptase II, suggesting that it may be important in controlling intracellular levels of this metabolite. We created cell lines stably expressing the enzyme to study its effect on cell function. We found that overexpression of 5ptase IV in 293 cells caused the rapid depletion of both PI(4,5)P2 and PI(3,4,5)P3 in cells with corresponding increases in the products, PI(4)P and PI(3,4)P2, changing the balance of two phosphoinositol products of phosphoinositide 3-kinase, PI(3,4)P2 and PI(3,4,5)P3, in the cell. One of the targets of these phosphoinositides is the serine/threonine kinase Akt, which plays an important role in the control of apoptosis. We were able to address the relative roles of PI(3,4)P2 and PI(3,4,5)P3 in the activation of Akt by selective depletion of these phosphoinositides in cells stably transfected with 5ptase IV and inositol polyphosphate 4-phosphatase (4ptase I). In cells transfected with 4ptase I, the level of PI(3,4)P2 was reduced, and PI(3,4,5)P3 was increased. Expression of the two enzymes had the opposite effect on the phosphorylation of Akt in response to stimulation with growth factors or heat shock. Akt phosphorylation was inhibited in cells expressing 5ptase IV but increased in 4ptase I cells and correlated with the intracellular level of PI(3,4,5)P3 and not that of PI(3,4)P2. The inhibition of Akt phosphorylation in cells expressing 5ptase IV makes them highly susceptible to FAS-induced apoptosis, whereas overexpressing of the 4ptase I protects cells from apoptosis. Our results place 5ptase IV as a relevant biological regulator of PI3K/Akt pathway in cells. Phosphatidylinositol lipids, also referred to as phosphoinositides, are involved in the regulation of an array of cellular processes, including cell growth, differentiation, cell motility, and apoptosis. The intracellular levels of phosphoinositides are modulated by an intricate interplay between the action of phosphoinositide kinases and phosphatases. In response to stimulation, phosphoinositide kinases, such as PI3K 1PI3Kphosphoinositide 3-kinase5ptaseinositol polyphosphate 5-phospatase4ptaseinositol polyphosphate 4-phospatasePI(4)Pphosphatidylinositol 4-phosphatePI(34)P2, phosphatidylinositol 3,4-bisphosphatePI(45)P2, phosphatidylinositol 4,5-bisphosphate, PI(3,4,5)P3, phosphatidylinositol 3,4,5-trisphosphateFBSfetal bovine serumLDHlactate dehydrogenase and PIPK, produce phosphatidylinositol second messengers phosphatidylinositol 3,4-bisphosphate (PI(3,4)P2), phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2), and phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P3) (1Rameh L.E. Cantley L.C. J. Biol. Chem. 1999; 274: 8347-8350Abstract Full Text Full Text PDF PubMed Scopus (852) Google Scholar, 2Park S.J. Itoh T. Takenawa T. J. Biol. Chem. 2001; 276: 4781-4787Abstract Full Text Full Text PDF PubMed Scopus (74) Google Scholar, 3Anderson R.A. Boronenkov I.V. Doughman S.D. Kunz J. Loijens J.C. J. Biol. Chem. 1999; 274: 9907-9910Abstract Full Text Full Text PDF PubMed Scopus (240) Google Scholar). Inositol polyphosphate phosphatases terminate these signals by hydrolysis of their corresponding substrates. Inositol polyphosphate 5-phosphatases (5ptases) remove the phosphate group at the D5 position, whereas inositol polyphosphate 4-phosphatases (4ptases) hydrolyze the D4 phosphate. The intracellular levels of PI(4,5)P2 and PI(3,4,5)P3 may be determined by 5ptases, and 4ptases could control the intracellular level of PI(3,4)P2 (4Majerus P.W. Kisseleva M.V. Norris F.A. J. Biol. Chem. 1999; 274: 10669-10672Abstract Full Text Full Text PDF PubMed Scopus (193) Google Scholar, 5Erneux C. Govaerts C. Communi D. Pesesse X. Biochim. Biophys. Acta. 1998; 1436: 185-199Crossref PubMed Scopus (126) Google Scholar). One of the best characterized targets of PI3K products, PI(3,4)P2and PI(3,4,5)P3, is the protein kinase Akt/protein kinase B (6Downward J. Curr. Opin. Cell Biol. 1998; 10: 262-267Crossref PubMed Scopus (1188) Google Scholar). Stimulation of PI3K results in the activation of Akt by phosphorylation. Recruitment of Akt to the cell membrane and phosphorylation by phosphoinositide-dependent kinase-1 are mediated by PI(3,4)P2 and PI(3,4,5)P3 (7Vanhaesebroeck B. Alessi D.R. Biochem. J. 2000; 346: 561-576Crossref PubMed Scopus (1399) Google Scholar,8Coffer P.J. Jin J. Woodgett J.R. Biochem. J. 1998; 335: 1-13Crossref PubMed Scopus (969) Google Scholar). phosphoinositide 3-kinase inositol polyphosphate 5-phospatase inositol polyphosphate 4-phospatase phosphatidylinositol 4-phosphate 4)P2, phosphatidylinositol 3,4-bisphosphate 5)P2, phosphatidylinositol 4,5-bisphosphate, PI(3,4,5)P3, phosphatidylinositol 3,4,5-trisphosphate fetal bovine serum lactate dehydrogenase We have identified an inositol polyphosphate phosphatase with a unique substrate specificity, inositol polyphosphate 5-phosphatase type IV (5ptase IV) (9Kisseleva M.V. Wilson M.P. Majerus P.W. J. Biol. Chem. 2000; 275: 20110-20116Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar). Only the lipid substrates, PI(4,5)P2 and PI(3,4,5)P3 are used by this enzyme in vitro. The 5ptase IV has a remarkably high affinity for PI(3,4,5)P3, making it a potential candidate for the regulation of the PI(3,4,5)P3 level in cells. We have established human embryonic kidney 293 cell lines stably transfected with the human 5ptase IV gene under a tetracycline-inducible promoter to determine the substrate specificity of this enzyme in vivo and to study the physiological consequences of 5ptase IV overexpression. Following overexpression of 5ptase IV in these cells, we measured changes in the levels of phosphoinositides and the effects of 5ptase IV protein expression on cell growth and apoptosis. We have determined Akt phosphorylation in cells expressing either 4ptase I or 5ptase IV and have studied the relative role of the two substrate phospholipids in Akt phosphorylation. The T-Rex tetracycline-regulated system (Invitrogen) was used to create cell lines stably expressing inositol phosphatases. cDNA encoding Myc-tagged full-length 5ptase IV (GenBankTMAF187891) was subcloned into theBamHI site of pcDNA 4/TO vector (Invitrogen). Full-length 4ptase I (GenBankTMU26398) was subcloned intoHindIII/EcoRI sites of the vector. 293T-Rex cells (Invitrogen) stably expressing the tetracycline repressor were transfected with either plasmid and selected with 400 μg of zeocin/ml for 4 weeks. Isolated clones were grown in Dulbecco's modified Eagle's medium supplemented with 2 mm glutamine, 1% penicillin/streptomycin, 10% Tet system-approved fetal bovine serum (FBS) (CLONTECH), 5 μg of blasticidin/ml, and 300 μg of zeocin/ml. An addition of 100 ng of tetracycline/ml to the growth medium induced the expression of the inositol phosphatase proteins. Cells were labeled in 35-mm tissue culture dishes in 1 ml of medium containing 400 μCi of [32P]orthophosphate (PerkinElmer Life Sciences) for 2–4 h at 37 °C in a phosphate-free Dulbecco's modified Eagle's medium (Invitrogen) containing 5% FBS dialyzed against 0.85% NaCl. No serum was added to the labeling medium of cells stimulated with growth factors. Cells were harvested, spun, and lysed by an addition of 1.8 ml of methanol:chloroform:8% HClO4 (20:10:1). After the addition of 500 μl of chloroform and 500 μl of 1% HClO4, the lower organic phase was collected, washed twice with 1% HClO4, and evaporated. Deacylation was as described previously (10Lips D.L. Majerus P.W. J. Biol. Chem. 1989; 264: 19911-19915Abstract Full Text PDF PubMed Google Scholar). The glycerophosphorylinositol derivatives were resolved on an anion-exchange column (PartiSphere Sax, 4.6 × 125 mm, Whatman) with the gradient of 0–1.25 mNaH2PO4, pH 4.5. The linear gradient rose to 7% B over 30 min. A 1-min step to 15% was followed by a linear gradient to 30% B at 60 min followed by a linear gradient to 60% B at 80 min. Finally, pump B was increased to 100% over 5 min. Radiolabeled deacylated phospholipids were detected by a β-RAM in-flow detection system (In/US Systems, Inc.). Cells were seeded at a density of 2000 cells/well in a 96-well plate. After 24 h, protein expression was induced by the addition of 100 ng of tetracycline/ml to the growth medium. The number of viable cells was determined using the CellTiter 96 AQueous Assay (Promega). Four wells were analyzed for each time point. Cells stably transfected with either 5ptase 4ptase I, or an vector were grown in the of tetracycline for an of of the were also with of Cells were washed twice with FBS in and in at 4 °C Cells were in containing mm and A and for 2 h at 4 Cells were analyzed by cell Assay was used to the effect of 5ptase IV overexpression on apoptosis. or 5ptase IV cells were seeded on a at a density of 2 × expression was induced by the addition of 100 ng of tetracycline/ml in Dulbecco's modified Eagle's medium supplemented with 5% was analyzed to the by the the of lactate dehydrogenase in the of cells was used to the of apoptosis. in the was measured with detection Cells were lysed with containing mm pH 1 mm 1% mm 5 mm mm mm 1 each of and and 1 containing 60 μg of protein were resolved by and to protein was by of the membrane with After in 5% bovine serum in for 1 h, were at °C with the in the After with containing detection was with and The of Akt detected with or was to the of Akt in a as detected with × cells were in a and protein expression was induced by in a medium containing 100 ng of was induced by the of with and protein in Dulbecco's modified Eagle's medium supplemented with 1% After for cells were and was using and μl of the The of was measured as a of in the with measured in the of vector cells. Only two lipid inositol PI(4,5)P2 and PI(3,4,5)P3, are of 5ptase IV in (9Kisseleva M.V. Wilson M.P. Majerus P.W. J. Biol. Chem. 2000; 275: 20110-20116Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar). determine the enzyme its substrate specificity in we created cell lines stably expressing Myc-tagged human 5ptase clones with levels of 5ptase IV expression were clones and as in the in overexpression was induced by the addition of 100 ng of tetracycline/ml to the growth h cells were labeled with [32P]orthophosphate for 2–4 h at 37 phosphoinositides were analyzed by anion-exchange high 1 of is are to the in each The intracellular level of PI(4,5)P2 was in cells overexpressing 5ptase IV with the cells transfected with vector 1 of 5ptase IV in an level of In the level of PI(4)P was increased of the expression of 5ptase IV protein 1 PI(4)P and PI(4,5)P2 the labeled phosphoinositides, and PI(3,4,5)P3 was In cells overexpressing 5ptase PI(3,4,5)P3 was to levels 1 as it be as a of hydrolysis of PI(3,4,5)P3 by 5ptase The clones with levels of the enzyme expression not in the effect on the levels of cellular phosphoinositides, and the level of expression was to the intracellular levels of PI(3,4,5)P3 not and with levels of 5ptase IV expression and vector cells were grown for in We used AQueous Assay to cell The vector cell In in the number of cells was in cell lines stably transfected with 5ptase IV of cell growth was with 5ptase IV overexpression may either cell growth or cell We the overexpression of 5ptase IV to cell apoptosis, or was as described under The of a is to the of the of cells expressing 5ptase IV were 24 h of enzyme expression with cells in vector cells 2 of 5ptase IV to apoptosis. with of cells expressing 5ptase IV were and of them were from the tissue culture the of vector cells apoptosis. We the effect of 5ptase IV expression on the cell The number of cells in of the cell was determined by cell at the of protein expression with tetracycline the of the of the were with a in 1998; Google Scholar). that h of 5ptase IV cells have in with the vector cells. 5ptase IV expression increased the number of cells in to whereas of vector cells were in of the 5ptase IV expression increased the number of cells in 5ptase IV expressing cells were in with the of cells transfected with an vector 24 h of with No were between cells stably transfected with 5ptase IV or an vector The was with the One of the involved in the regulation of cell is Akt/protein kinase B (6Downward J. Curr. Opin. Cell Biol. 1998; 10: 262-267Crossref PubMed Scopus (1188) Google Scholar, B. Alessi D.R. Biochem. J. 2000; 346: 561-576Crossref PubMed Scopus (1399) Google Scholar). The kinase of Akt is increased in response to phosphorylation of and by phosphoinositide-dependent The phosphoinositides PI(3,4)P2 and PI(3,4,5)P3 are for activation of Akt R.A. Alessi D.R. J. Biochem. J. 1999; PubMed Scopus Google Scholar, D.R. Curr. Biol. Full Text Full Text PDF PubMed Google Scholar, D. T. PubMed Scopus Google Scholar, D. J. 1998; PubMed Scopus Google Scholar). We the phosphorylation of Akt in cells stably transfected with 5ptase IV or vector. of 5ptase IV protein results in the inhibition of Akt phosphorylation in response to growth stimulation activation was a stimulation with growth not We between the of phosphorylation on and In response to an such as with a growth or heat phosphorylation of Akt is increased. of PI3K followed by an in the intracellular levels of PI(3,4)P2 and PI(3,4,5)P3 this response Biochem. 1999; PubMed Scopus Google Scholar). We the phosphorylation of Akt in cells stably expressing either 5ptase IV or 4ptase I, an inositol phosphatase that PI(3,4)P2 F.A. Majerus P.W. J. Biol. Chem. Full Text PDF PubMed Google Scholar). In cells, the expression of both inositol phosphatases to a in Akt phosphorylation In 5ptase IV cells, Akt phosphorylation was In cells overexpressing 4ptase I, Akt phosphorylation was were at with the these cell lines as as cells transfected with vector were followed by stimulation with of growth or growth cells were to a heat for min at Akt phosphorylation was inhibited in 5ptase cells but was increased in cells overexpressing 4ptase I stimulation with growth factors or heat the activation of PI3K results in a in the intracellular levels of PI(3,4)P2 and PI(3,4,5)P3 (1Rameh L.E. Cantley L.C. J. Biol. Chem. 1999; 274: 8347-8350Abstract Full Text Full Text PDF PubMed Scopus (852) Google Scholar). hydrolysis of phosphoinositides by either 5ptase IV or 4ptase I the levels of these phospholipids in cells (4Majerus P.W. Kisseleva M.V. Norris F.A. J. Biol. Chem. 1999; 274: 10669-10672Abstract Full Text Full Text PDF PubMed Scopus (193) Google Scholar). cells were labeled with [32P]orthophosphate for 2 h and stimulated with growth The levels of PI(3,4)P2 and PI(3,4,5)P3 are as in A and In cells, the level of these are The levels of PI(3,4)P2 and PI(3,4,5)P3 were detected min of stimulation not In vector cells, the of PI(3,4)P2 was In cells stably transfected with 5ptase the intracellular level of PI(3,4,5)P3 was with the vector cells and by a in the level of PI(3,4)P2. is in a of the PI(3,4,5)P3 to PI(3,4)P2 to The overexpression of 4ptase I had the opposite the level of PI(3,4)P2 was increased and the PI(3,4,5)P3 to PI(3,4)P2 was the intracellular level of PI(3,4,5)P3 than with the inhibition of the Akt phosphorylation in cells overexpressing 5ptase IV and the of Akt phosphorylation in cells overexpressing 4ptase We the effects of overexpression of 4ptase I and 5ptase IV on FAS-induced and the phosphorylation of apoptosis, we measured in the to the membrane 5ptase IV overexpression cell whereas the overexpression of 4ptase I protects cells from h, in the from cells expressing 5ptase IV was than in vector cells, that 5ptase IV expressing cells are susceptible to FAS-induced apoptosis. in the medium from 4ptase cells was results were was not of Akt correlated with the levels of Akt phosphorylation in 5ptase cells with of the cells to FAS-induced apoptosis. of 4ptase I the phosphorylation of Akt, and the activation of Akt FAS-induced Inositol polyphosphate 5-phosphatases control the intracellular levels of and lipid inositol including and PI(3,4,5)P3 of these by 5-phosphatases results in the of by these 5ptase IV the lipid substrates, PI(4,5)P2 and PI(3,4,5)P3 (9Kisseleva M.V. Wilson M.P. Majerus P.W. J. Biol. Chem. 2000; 275: 20110-20116Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar). affinity for m = 0.65 μM) is approximately 10-fold greater than that of the other 5ptases, which use this substrate including SHIP, OCRL, and 5ptase II, suggesting that it may be important in controlling intracellular levels of this metabolite. We created cell lines overexpressing the enzyme in a system to study its effect on cell function. We found that overexpression of 5ptase IV in 293 cells caused the rapid depletion of both PI(4,5)P2 and PI(3,4,5)P3 in cells with corresponding increases in the products, PI(4)P and PI(3,4)P2. The activation of PI3K by a to a of PI(3,4)P2and PI(3,4,5)P3 in cells. One of the of these is to the serine/threonine kinase Akt that plays an important role in the control of apoptosis. inositol Akt to the membrane it is by phosphorylation by phosphoinositide-dependent protein kinase of Akt is by the phosphorylation of and Akt a number of targets that to the inhibition of J. Curr. Opin. Cell Biol. 1998; 10: 262-267Crossref PubMed Scopus (1188) Google Scholar, B. Alessi D.R. Biochem. J. 2000; 346: 561-576Crossref PubMed Scopus (1399) Google D. J. 1998; PubMed Scopus Google Scholar). The role of PI(3,4)P2 in Akt is has that PI(3,4)P2 but not PI(3,4,5)P3 Akt D.R. Cantley L.C. 275: PubMed Scopus Google Scholar, J.R. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, D. Biol. PubMed Scopus Google Scholar). in of the activation of phosphoinositide-dependent it that PI(3,4,5)P3 is important R.A. Alessi D.R. J. Biochem. J. 1999; PubMed Scopus Google Scholar, D.R. Curr. Biol. Full Text Full Text PDF PubMed Google Scholar, D. T. PubMed Scopus Google Scholar). products of PI3K are for Akt it be inhibited by PI3K and D.R. Full Text PDF PubMed Scopus Google Scholar, Alessi D.R. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, C. D. Biol. 2000; PubMed Scopus Google Scholar). The activation of Akt in cells the has to the hydrolysis of the PI3K products as a of phosphoinositide of T. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). of Akt in cells be by of C. T. J. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar, D. D. Curr. Biol. 1998; Full Text Full Text PDF PubMed Google Scholar, 1998; PubMed Scopus Google Scholar). In both of these cells are of both PI(3,4)P2and PI(3,4,5)P3 of the relative role of the two A role of PI(3,4,5)P3 in activation of Akt has by the of the effects of the and on Akt activation and C. D. Biol. 2000; PubMed Scopus Google Scholar, J. 2000; PubMed Scopus Google Scholar, T. 1999; PubMed Scopus Google Scholar, T. T. T. T. Biol. 2001; PubMed Scopus Google Scholar). of results in of the intracellular level of PI(3,4,5)P3 T. 1999; PubMed Scopus Google whereas overexpression of to the of the PI(3,4,5)P3 level C. D. Biol. 2000; PubMed Scopus Google Scholar, T. T. T. T. Biol. 2001; PubMed Scopus Google Scholar). and cells from were susceptible to and the activation of these cells with in the of PI(3,4,5)P3 and Akt phosphorylation T. 1999; PubMed Scopus Google Scholar). an inhibition of Akt activation was detected in B cells overexpressing T. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar, D. T. J. Biol. Chem. 1999; 274: Full Text Full Text PDF PubMed Scopus Google Scholar). overexpression of in cells to of Akt and cell C. D. Biol. 2000; PubMed Scopus Google Scholar). An of this study is by the that to Akt In these the levels of PI(3,4,5)P3 were but PI(3,4)P2 We were able to address the relative of PI(3,4)P2 and PI(3,4,5)P3 in the activation of Akt by selective depletion of phosphoinositides in cells stably transfected with 5ptase IV or 4ptase The intracellular level of PI(3,4,5)P3 is by the expression of 5ptase and PI(3,4)P2 is by the overexpression of 4ptase In cells, expression of either of the two inositol phosphatases the level of Akt phosphorylation. A was cell stimulation with growth factors or heat shock. of 5ptase IV inhibited Akt In cells expressing 4ptase I had levels of Akt phosphorylation than vector cells. of phosphoinositides that relative levels of the two products of PI(3,4)P2 and PI(3,4,5)P3, in these cells are in opposite of PI(3,4,5)P3 by in the depletion of PI(3,4,5)P3 and of PI(3,4)P2. In cells transfected with 4ptase I, the level of PI(3,4)P2 was reduced, and PI(3,4,5)P3 was increased. The increased PI(3,4,5)P3 level in 4ptase cells the action of phosphatidylinositol phosphate kinases R.A. Boronenkov I.V. Doughman S.D. Kunz J. Loijens J.C. J. Biol. Chem. 1999; 274: 9907-9910Abstract Full Text Full Text PDF PubMed Scopus (240) Google Scholar). The of PI(3,4,5)P3 from in a by phosphatidylinositol phosphate kinases has in X. Loijens J.C. Boronenkov I.V. Norris F.A. J. Majerus P.W. R.A. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). the effect of the expression of inositol phosphatases on phosphorylation of Akt with the intracellular level of PI(3,4,5)P3 and not that of PI(3,4)P2. In with these changes in Akt we the effect on 5ptase IV expression increased apoptosis, and 4ptase I of PI(4,5)P2 the intracellular substrate for PI3K to produce the of PI(3,4,5)P3 in cells overexpressing 5ptase IV is to hydrolysis of both the substrate PI(4,5)P2 and the PI(3,4,5)P3 of The inhibition of the PI3K/Akt pathway has to in cell C. D. Biol. 2000; PubMed Scopus Google Scholar, 1998; PubMed Scopus Google Scholar, 1999; PubMed Scopus Google Scholar). Expression of inositol phosphatases that hydrolyze the lipid products of and of the cell A of into cells cell in phase 1998; Google Scholar, 1999; PubMed Scopus Google Scholar). Expression of inositol 5-phosphatase to the cell as C. D. Biol. 2000; PubMed Scopus Google Scholar). effect was to of the cell the kinase of and into phase 1998; PubMed Scopus Google Scholar, Curr. Biol. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). of 5ptase IV also cell h of of the 5ptase IV a number of cells in phase with vector cells in a to that with the overexpression of and the role of PI(3,4,5)P3 in the regulation of cell by In the expression of 5ptase IV results in a in the intracellular level of in to an of the cell in phase 24 h of the 5ptase IV protein An of cells in phase is caused by with the in the intracellular level of expression of 5ptase IV results in the cell in and growth inhibition of cells stably transfected with the The inhibition of Akt activation in cells stably expressing 5ptase IV makes them highly susceptible to apoptosis. of with a in 293 cells. Expression of 5ptase IV the effect of The in the of to the membrane was in cells expressing 5ptase IV than in vector cells. the overexpressing the 4ptase I protects cells from FAS-induced apoptosis, and the in the is lower with the with the of FAS-induced in cells stably transfected with the two inositol phosphatases. Akt is not to be involved in mediated by including of Akt with growth the of protects cells from J. Biol. Chem. 1999; 274: Full Text Full Text PDF PubMed Scopus Google Scholar). from are to FAS-induced apoptosis, and Akt was found to be in these cells C. 1999; PubMed Scopus Google Scholar). A of the intracellular level of PI(4,5)P2 in cells overexpressing 5ptase IV may have to the of in that PI(4,5)P2 may be involved in the regulation of by the of T. D. J. Biol. Chem. 2000; 275: Full Text Full Text PDF PubMed Scopus Google Scholar, X. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar). T. D. J. Biol. Chem. 2000; 275: Full Text Full Text PDF PubMed Scopus Google that in vitro. was for the T. D. J. Biol. Chem. 2000; 275: Full Text Full Text PDF PubMed Scopus Google Scholar). In X. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google detected the effect of PI(4,5)P2 to in and the addition of this effect X. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar). be that in both the effect of PI(4,5)P2 was in in and it to be PI(4,5)P2 has effect on in In we that 5ptase a of a of inositol polyphosphate phosphatase of is a in of intracellular PI(3,4,5)P3 by this enzyme to the inhibition of Akt and makes cells highly susceptible to apoptosis. 5ptase IV as a relevant biological regulator of PI3K/Akt pathway in cells. 5ptase IV has the affinity PI(3,4,5)P3 of is in and cell lines including and (9Kisseleva M.V. Wilson M.P. Majerus P.W. J. Biol. Chem. 2000; 275: 20110-20116Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar). A of that two other 5-phosphatases SHIP, to cells, and a are involved in the regulation of PI3K/Akt pathway C. D. Biol. 2000; PubMed Scopus Google Scholar, J. 2000; PubMed Scopus Google Scholar, T. 1999; PubMed Scopus Google Scholar, T. T. T. T. Biol. 2001; PubMed Scopus Google Scholar). be to cells for the or of 5ptase IV and other We for with the cell and Wilson for