Recombinant mouse phosphatidylinositol transfer protein (PI-TP)β is a substrate for protein kinase C (PKC)-dependent phosphorylation in vitro. Based on site-directed mutagenesis and two-dimensional tryptic peptide mapping, Ser262 was identified as the major site of phosphorylation and Ser165 as a minor phosphorylation site. The phospholipid transfer activities of wild-type PI-TPβ and PI-TPβ(S262A) were identical, whereas PI-TPβ(S165A) was completely inactive. PKC-dependent phosphorylation of Ser262 also had no effect on the transfer activity of PI-TPβ. To investigate the role of Ser262 in the functioning of PI-TPβ, wtPI-TPβ and PI-TPβ(S262A) were overexpressed in NIH3T3 fibroblast cells. Two-dimensional PAGE analysis of cell lysates was used to separate PI-TPβ from its phosphorylated form. After Western blotting, wtPI-TPβ was found to be 85% phosphorylated, whereas PI-TPβ(S262A) was not phosphorylated. In the presence of the PKC inhibitor GF 109203X, the phosphorylated form of wtPI-TPβ was strongly reduced. Immunolocalization showed that wtPI-TPβ was predominantly associated with the Golgi membranes. In the presence of the PKC inhibitor, wtPI-TPβ was distributed throughout the cell similar to what was observed for PI-TPβ(S262A). In contrast to wtPI-TPβ overexpressors, cells overexpressing PI-TPβ(S262A) were unable to rapidly replenish sphingomyelin in the plasma membrane upon degradation by sphingomyelinase. This implies that PKC-dependent association with the Golgi complex is a prerequisite for PI-TPβ to express its effect on sphingomyelin metabolism. Recombinant mouse phosphatidylinositol transfer protein (PI-TP)β is a substrate for protein kinase C (PKC)-dependent phosphorylation in vitro. Based on site-directed mutagenesis and two-dimensional tryptic peptide mapping, Ser262 was identified as the major site of phosphorylation and Ser165 as a minor phosphorylation site. The phospholipid transfer activities of wild-type PI-TPβ and PI-TPβ(S262A) were identical, whereas PI-TPβ(S165A) was completely inactive. PKC-dependent phosphorylation of Ser262 also had no effect on the transfer activity of PI-TPβ. To investigate the role of Ser262 in the functioning of PI-TPβ, wtPI-TPβ and PI-TPβ(S262A) were overexpressed in NIH3T3 fibroblast cells. Two-dimensional PAGE analysis of cell lysates was used to separate PI-TPβ from its phosphorylated form. After Western blotting, wtPI-TPβ was found to be 85% phosphorylated, whereas PI-TPβ(S262A) was not phosphorylated. In the presence of the PKC inhibitor GF 109203X, the phosphorylated form of wtPI-TPβ was strongly reduced. Immunolocalization showed that wtPI-TPβ was predominantly associated with the Golgi membranes. In the presence of the PKC inhibitor, wtPI-TPβ was distributed throughout the cell similar to what was observed for PI-TPβ(S262A). In contrast to wtPI-TPβ overexpressors, cells overexpressing PI-TPβ(S262A) were unable to rapidly replenish sphingomyelin in the plasma membrane upon degradation by sphingomyelinase. This implies that PKC-dependent association with the Golgi complex is a prerequisite for PI-TPβ to express its effect on sphingomyelin metabolism. phosphatidylinositol transfer protein wild-type PI-TP phosphatidylinositol phosphatidylcholine inositol sphingomyelin protein kinase C phosphatidic acid phosphatidylserine trinitrophenyl phosphatidylethanolamine goat anti-rabbit peroxidase bacterial sphingomyelinase phosphate-buffered saline 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid phospholipase A2 pyrene guanosine 5′-O-(thiotriphosphate) L-1-tosylamido-2-phenylethyl chloromethyl ketone Eukaryotic phosphatidylinositol transfer proteins (PI-TPs)1 belong to a family of highly conserved proteins that are able to transfer phospholipids between membranes or from membrane to enzyme (1Wirtz K.W.A. Biochem. J. 1997; 324: 353-360Crossref PubMed Scopus (174) Google Scholar). In mammalian tissues at least two isoforms, PI-TPα and PI-TPβ, are found. PI-TPα is able to transfer phosphatidylinositol (PI) and, to a lesser extent, phosphatidylcholine (PC) (2Wirtz K.W.A. Annu. Rev. Biochem. 1991; 60: 73-99Crossref PubMed Scopus (377) Google Scholar, 3van Paridon P.A. Visser A.J. Wirtz K.W.A. Biochim. Biophys. Acta. 1987; 898: 172-180Crossref PubMed Scopus (64) Google Scholar, 4Helmkamp Jr G.M. Harvey M.S. Wirtz K.W.A. Van Deenen L.L.M. J. Biol. Chem. 1974; 249: 6382-6389Abstract Full Text PDF PubMed Google Scholar, 5Helmkamp Jr G.M. Chem. Phys. Lipids. 1985; 38: 3-16Crossref PubMed Scopus (34) Google Scholar, 6Helmkamp Jr G.M. J. Bioenerg. Biomembr. 1986; 18: 71-91Crossref PubMed Scopus (56) Google Scholar) and is mainly localized in the cytosol and in the nucleus (7de Vries K.J. Westerman J. Bastiaens P.I. Jovin T.M. Wirtz K.W.A. Snoek G.T. Exp. Cell Res. 1996; 227: 33-39Crossref PubMed Scopus (81) Google Scholar). Similar to PI-TPα, PI-TPβ is able to transfer PI and PC but is also able to transfer sphingomyelin (SM) (8de Vries K.J. Heinrichs A.A. Cunningham E. Brunink F. Westerman J. Somerharju P.J. Cockcroft S. Wirtz K.W.A. Snoek G.T. Biochem. J. 1995; 310: 643-649Crossref PubMed Scopus (89) Google Scholar). PI-TPβ is mainly associated with the Golgi apparatus (7de Vries K.J. Westerman J. Bastiaens P.I. Jovin T.M. Wirtz K.W.A. Snoek G.T. Exp. Cell Res. 1996; 227: 33-39Crossref PubMed Scopus (81) Google Scholar). The primary sequences of PI-TPα and PI-TPβ are very similar, with an identity of 77% and a similarity of 94% (9Tanaka S. Hosaka K. J. Biochem. (Tokyo). 1994; 115: 981-984Crossref PubMed Scopus (84) Google Scholar).To date, little is known about the exact cellular function of PI-TPα and PI-TPβ. In a cell-free system containing trans-Golgi membranes, both PI-TPα and PI-TPβ stimulated the formation of constitutive secretory vesicles and immature granules (10Ohashi M. de Vries K.J. Frank R. Snoek G.T. Bankaitis V. Wirtz K.W.A. Huttner W.B. Nature. 1995; 377: 544-547Crossref PubMed Scopus (168) Google Scholar). In permeabilized, cytosol-depleted HL60 cells, both isoforms restored GTPγS-stimulated protein secretion and phospholipase C-mediated inositol lipid signaling (11Fensome A. Cunningham E. Prosser S. Tan S.K. Swigart P. Thomas G. Hsuan J. Cockcroft S. Curr. Biol. 1996; 6: 730-738Abstract Full Text Full Text PDF PubMed Google Scholar, 12Cunningham E. Tan S.K. Swigart P. Hsuan J. Bankaitis V. Cockcroft S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 6589-6593Crossref PubMed Scopus (99) Google Scholar). In these assays, PI-TPα and PI-TPβ were found to function equally well despite their different biochemical properties and cellular localizations. On the other hand, NIH3T3 cells with increased expression of either PI-TPα or PI-TPβ demonstrated remarkable differences in lipid metabolic pathways. Cells overexpressing PI-TPα (SPIα cells) showed an enhanced PLA2-mediated PI degradation resulting in increased levels of lyso-PI, glycerophosphoinositol, Ins(1)P, and Ins(2)P (13Snoek G.T. Berrie C.P. Geijtenbeek T.B.H. van der Helm H.A. Cadee J.A. Iurisci C. Corda D. Wirtz K.W.A. J. Biol. Chem. 1999; 274: 35393-35399Abstract Full Text Full Text PDF PubMed Scopus (37) Google Scholar). This was not observed in cells overexpressing PI-TPβ (SPIβ cells). However, in SPIβ cells (but not in SPIα cells) it was shown that under conditions in which plasma membrane SM was hydrolyzed to ceramide by exogenous sphingomyelinase, PI-TPβ was involved in maintaining the steady-state levels of SM (14van Tiel C.M. Luberto C. Snoek G.T. Hannun Y.A. Wirtz K.W.A. Biochem. J. 2000; 346: 537-543Crossref PubMed Scopus (31) Google Scholar). It was recently postulated that PI-TPβ plays a key role in SM metabolism, making it a potential regulator of pathways for diacylglycerol production and consumption in the mammalian trans-Golgi network (15Bankaitis PubMed Scopus Google Scholar). of the PI-TPβ in to in K. J. S. Bankaitis Biol. PubMed Scopus Google of by which PI-TP activity is PI-TPα was shown to be phosphorylated by protein kinase C in well as in G.T. Westerman J. Wirtz K.W.A. Biochem. J. PubMed Scopus Google Scholar, Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). PI-TPα was phosphorylated on with the form of PI-TPα phosphorylated the form Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). in NIH3T3 cells, PI-TPα was from the cytosol to the Golgi membranes upon phosphorylation with This of PI-TPα with an increased of lyso-PI, the of a G.T. Westerman J. Wirtz K.W.A. Biochem. J. PubMed Scopus Google Scholar, Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). Based on these a was for the of PI-TPα by PKC-dependent In contrast to PI-TPα, PI-TPβ from or not be phosphorylated despite the that it the and an PKC phosphorylation site not in PI-TPα (8de Vries K.J. Heinrichs A.A. Cunningham E. Brunink F. Westerman J. Somerharju P.J. Cockcroft S. Wirtz K.W.A. Snoek G.T. Biochem. J. 1995; 310: 643-649Crossref PubMed Scopus (89) Google Scholar, G.T. Westerman J. Wirtz K.W.A. Biochem. J. PubMed Scopus Google Scholar, de E. van J. Brunink F. Westerman J. Snoek G.T. Wirtz K.W.A. Biochim. Biophys. Acta. 1994; PubMed Scopus Google that PI-TPβ be phosphorylated by PKC in as well as in The major site of phosphorylation was whereas a minor site. site-directed mutagenesis shown that Ser165 is for the lipid transfer activity of the whereas phosphorylation of Ser262 is for the association of PI-TPβ with the Golgi membranes. This is also for the of PI-TPβ to steady-state levels of SM in the plasma shown that PI-TPβ is a substrate for analysis that Ser262 was the phosphorylation site. However, which is to in PI-TPα, was also phosphorylated to a very In a had shown that the PKC-dependent phosphorylation of PI-TPα was to Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). The PI-TPα which is highly to PI-TPβ of Ser262 (9Tanaka S. Hosaka K. J. Biochem. (Tokyo). 1994; 115: 981-984Crossref PubMed Scopus (84) Google Scholar). that phosphorylation of the PI-TP isoforms is for the of their were to the role of Ser262 in the function of peptide of wtPI-TPβ and PI-TPβ(S165A) major which are from the of PI-TPβ(S262A) This that these containing phosphorylated The formation of these is to the presence of tryptic in the acid of the peptide the and to the in van der P. 1991; PubMed Scopus Google Scholar, to to and to The tryptic of PI-TPβ(S262A) showed a peptide containing phosphorylated This was in the peptide of that phosphorylation was to of Ser165 with PI-TPβ(S165A) and the both of which in completely and SM transfer On the other hand, PI-TPβ(S262A) was of the in PI-TPα also an protein Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). the it be that is at the as of the of PI-TPα Jr G.M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). it is that the of transfer activity is to the of to with the membrane However, at the that of with the of the protein in E. The for the of transfer activity observed in PI-TPβ(S165A) and After PKC-dependent the phospholipid transfer activity of PI-TPβ was that phosphorylation of Ser262 had no effect lysates from SPIβ cells two of PI-TPβ that be by analysis it was that 85% of the PI-TPβ at and at of the cells with the PKC inhibitor GF PI-TPβ to strongly that the at the phosphorylated form of PI-TPβ. the from the cells predominantly PI-TPβ at that in PI-TPβ is phosphorylated at that GF and D. P. P. M. V. P. E. F. D. J. J. Biol. Chem. 1991; Full Text PDF PubMed Google Scholar, J. M. M. M. M. S. J. F. Biochem. J. PubMed Scopus Google not which PKC is involved in the phosphorylation of PI-TPβ. it is that PKC is in these cells, it that phosphorylated PI-TPβ in association with the Golgi is a substrate for protein In a PI-TPβ from not be phosphorylated by PKC (8de Vries K.J. Heinrichs A.A. Cunningham E. Brunink F. Westerman J. Somerharju P.J. Cockcroft S. Wirtz K.W.A. Snoek G.T. Biochem. J. 1995; 310: 643-649Crossref PubMed Scopus (89) Google Scholar). protein is to PI-TPβ, it that in also PI-TPβ is mainly in its phosphorylated that in mouse PI-TPβ was predominantly associated with the Golgi (8de Vries K.J. Heinrichs A.A. Cunningham E. Brunink F. Westerman J. Somerharju P.J. Cockcroft S. Wirtz K.W.A. Snoek G.T. Biochem. J. 1995; 310: 643-649Crossref PubMed Scopus (89) Google Scholar). In the that PI-TPβ was associated with the Golgi complex in the SPIβ cells SPIβ cells with GF 109203X, a of PI-TPβ from the Golgi to the was observed similar throughout the was observed for PI-TPβ(S262A) in NIH3T3 cells that Ser262 to be phosphorylated for PI-TPβ to be associated with the Golgi It is to be that the phosphorylation site Ser262 is in PI-TPβ, whereas the phosphorylation site is conserved in identified with the of PI-TP Jr G.M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google contrast to PI-TPβ(S262A) that is overexpressed in mouse is not able to the of SM the of lipid by sphingomyelinase. the protein lipid transfer activity in that the association of PI-TPβ with the Golgi is a prerequisite for PI-TPβ to SM SM and in the Golgi also to the Golgi and phosphorylation of the protein J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). Similar to its PI-TPβ a role in the of vesicles from the Golgi (10Ohashi M. de Vries K.J. Frank R. Snoek G.T. Bankaitis V. Wirtz K.W.A. Huttner W.B. Nature. 1995; 377: 544-547Crossref PubMed Scopus (168) Google Scholar, R. J. Cell Biol. PubMed Scopus Google Scholar). It well that the of SM is to the and of lipid K. D. Rev. Biol. 2000; PubMed Scopus Google Scholar). are PI-TPβ is involved in Eukaryotic phosphatidylinositol transfer proteins (PI-TPs)1 belong to a family of highly conserved proteins that are able to transfer phospholipids between membranes or from membrane to enzyme (1Wirtz K.W.A. Biochem. J. 1997; 324: 353-360Crossref PubMed Scopus (174) Google Scholar). In mammalian tissues at least two isoforms, PI-TPα and PI-TPβ, are found. PI-TPα is able to transfer phosphatidylinositol (PI) and, to a lesser extent, phosphatidylcholine (PC) (2Wirtz K.W.A. Annu. Rev. Biochem. 1991; 60: 73-99Crossref PubMed Scopus (377) Google Scholar, 3van Paridon P.A. Visser A.J. Wirtz K.W.A. Biochim. Biophys. Acta. 1987; 898: 172-180Crossref PubMed Scopus (64) Google Scholar, 4Helmkamp Jr G.M. Harvey M.S. Wirtz K.W.A. Van Deenen L.L.M. J. Biol. Chem. 1974; 249: 6382-6389Abstract Full Text PDF PubMed Google Scholar, 5Helmkamp Jr G.M. Chem. Phys. Lipids. 1985; 38: 3-16Crossref PubMed Scopus (34) Google Scholar, 6Helmkamp Jr G.M. J. Bioenerg. Biomembr. 1986; 18: 71-91Crossref PubMed Scopus (56) Google Scholar) and is mainly localized in the cytosol and in the nucleus (7de Vries K.J. Westerman J. Bastiaens P.I. Jovin T.M. Wirtz K.W.A. Snoek G.T. Exp. Cell Res. 1996; 227: 33-39Crossref PubMed Scopus (81) Google Scholar). Similar to PI-TPα, PI-TPβ is able to transfer PI and PC but is also able to transfer sphingomyelin (SM) (8de Vries K.J. Heinrichs A.A. Cunningham E. Brunink F. Westerman J. Somerharju P.J. Cockcroft S. Wirtz K.W.A. Snoek G.T. Biochem. J. 1995; 310: 643-649Crossref PubMed Scopus (89) Google Scholar). PI-TPβ is mainly associated with the Golgi apparatus (7de Vries K.J. Westerman J. Bastiaens P.I. Jovin T.M. Wirtz K.W.A. Snoek G.T. Exp. Cell Res. 1996; 227: 33-39Crossref PubMed Scopus (81) Google Scholar). The primary sequences of PI-TPα and PI-TPβ are very similar, with an identity of 77% and a similarity of 94% (9Tanaka S. Hosaka K. J. Biochem. (Tokyo). 1994; 115: 981-984Crossref PubMed Scopus (84) Google Scholar). To date, little is known about the exact cellular function of PI-TPα and PI-TPβ. In a cell-free system containing trans-Golgi membranes, both PI-TPα and PI-TPβ stimulated the formation of constitutive secretory vesicles and immature granules (10Ohashi M. de Vries K.J. Frank R. Snoek G.T. Bankaitis V. Wirtz K.W.A. Huttner W.B. Nature. 1995; 377: 544-547Crossref PubMed Scopus (168) Google Scholar). In permeabilized, cytosol-depleted HL60 cells, both isoforms restored GTPγS-stimulated protein secretion and phospholipase C-mediated inositol lipid signaling (11Fensome A. Cunningham E. Prosser S. Tan S.K. Swigart P. Thomas G. Hsuan J. Cockcroft S. Curr. Biol. 1996; 6: 730-738Abstract Full Text Full Text PDF PubMed Google Scholar, 12Cunningham E. Tan S.K. Swigart P. Hsuan J. Bankaitis V. Cockcroft S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 6589-6593Crossref PubMed Scopus (99) Google Scholar). In these assays, PI-TPα and PI-TPβ were found to function equally well despite their different biochemical properties and cellular localizations. On the other hand, NIH3T3 cells with increased expression of either PI-TPα or PI-TPβ demonstrated remarkable differences in lipid metabolic pathways. Cells overexpressing PI-TPα (SPIα cells) showed an enhanced PLA2-mediated PI degradation resulting in increased levels of lyso-PI, glycerophosphoinositol, Ins(1)P, and Ins(2)P (13Snoek G.T. Berrie C.P. Geijtenbeek T.B.H. van der Helm H.A. Cadee J.A. Iurisci C. Corda D. Wirtz K.W.A. J. Biol. Chem. 1999; 274: 35393-35399Abstract Full Text Full Text PDF PubMed Scopus (37) Google Scholar). This was not observed in cells overexpressing PI-TPβ (SPIβ cells). However, in SPIβ cells (but not in SPIα cells) it was shown that under conditions in which plasma membrane SM was hydrolyzed to ceramide by exogenous sphingomyelinase, PI-TPβ was involved in maintaining the steady-state levels of SM (14van Tiel C.M. Luberto C. Snoek G.T. Hannun Y.A. Wirtz K.W.A. Biochem. J. 2000; 346: 537-543Crossref PubMed Scopus (31) Google Scholar). It was recently postulated that PI-TPβ plays a key role in SM metabolism, making it a potential regulator of pathways for diacylglycerol production and consumption in the mammalian trans-Golgi network (15Bankaitis PubMed Scopus Google Scholar). of the PI-TPβ in to in K. J. S. Bankaitis Biol. PubMed Scopus Google Scholar). In of by which PI-TP activity is PI-TPα was shown to be phosphorylated by protein kinase C in well as in G.T. Westerman J. Wirtz K.W.A. Biochem. J. PubMed Scopus Google Scholar, Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). PI-TPα was phosphorylated on with the form of PI-TPα phosphorylated the form Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). in NIH3T3 cells, PI-TPα was from the cytosol to the Golgi membranes upon phosphorylation with This of PI-TPα with an increased of lyso-PI, the of a G.T. Westerman J. Wirtz K.W.A. Biochem. J. PubMed Scopus Google Scholar, Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). Based on these a was for the of PI-TPα by PKC-dependent In contrast to PI-TPα, PI-TPβ from or not be phosphorylated despite the that it the and an PKC phosphorylation site not in PI-TPα (8de Vries K.J. Heinrichs A.A. Cunningham E. Brunink F. Westerman J. Somerharju P.J. Cockcroft S. Wirtz K.W.A. Snoek G.T. Biochem. J. 1995; 310: 643-649Crossref PubMed Scopus (89) Google Scholar, G.T. Westerman J. Wirtz K.W.A. Biochem. J. PubMed Scopus Google Scholar, de E. van J. Brunink F. Westerman J. Snoek G.T. Wirtz K.W.A. Biochim. Biophys. Acta. 1994; PubMed Scopus Google Scholar). In that PI-TPβ be phosphorylated by PKC in as well as in The major site of phosphorylation was whereas a minor site. site-directed mutagenesis shown that Ser165 is for the lipid transfer activity of the whereas phosphorylation of Ser262 is for the association of PI-TPβ with the Golgi membranes. This is also for the of PI-TPβ to steady-state levels of SM in the plasma shown that PI-TPβ is a substrate for analysis that Ser262 was the phosphorylation site. However, which is to in PI-TPα, was also phosphorylated to a very In a had shown that the PKC-dependent phosphorylation of PI-TPα was to Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). The PI-TPα which is highly to PI-TPβ of Ser262 (9Tanaka S. Hosaka K. J. Biochem. (Tokyo). 1994; 115: 981-984Crossref PubMed Scopus (84) Google Scholar). that phosphorylation of the PI-TP isoforms is for the of their were to the role of Ser262 in the function of peptide of wtPI-TPβ and PI-TPβ(S165A) major which are from the of PI-TPβ(S262A) This that these containing phosphorylated The formation of these is to the presence of tryptic in the acid of the peptide the and to the in van der P. 1991; PubMed Scopus Google Scholar, to to and to The tryptic of PI-TPβ(S262A) showed a peptide containing phosphorylated This was in the peptide of that phosphorylation was to of Ser165 with PI-TPβ(S165A) and the both of which in completely and SM transfer On the other hand, PI-TPβ(S262A) was of the in PI-TPα also an protein Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). the it be that is at the as of the of PI-TPα Jr G.M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). it is that the of transfer activity is to the of to with the membrane However, at the that of with the of the protein in E. The for the of transfer activity observed in PI-TPβ(S165A) and After PKC-dependent the phospholipid transfer activity of PI-TPβ was that phosphorylation of Ser262 had no effect lysates from SPIβ cells two of PI-TPβ that be by analysis it was that 85% of the PI-TPβ at and at of the cells with the PKC inhibitor GF PI-TPβ to strongly that the at the phosphorylated form of PI-TPβ. the from the cells predominantly PI-TPβ at that in PI-TPβ is phosphorylated at that GF and D. P. P. M. V. P. E. F. D. J. J. Biol. Chem. 1991; Full Text PDF PubMed Google Scholar, J. M. M. M. M. S. J. F. Biochem. J. PubMed Scopus Google not which PKC is involved in the phosphorylation of PI-TPβ. it is that PKC is in these cells, it that phosphorylated PI-TPβ in association with the Golgi is a substrate for protein In a PI-TPβ from not be phosphorylated by PKC (8de Vries K.J. Heinrichs A.A. Cunningham E. Brunink F. Westerman J. Somerharju P.J. Cockcroft S. Wirtz K.W.A. Snoek G.T. Biochem. J. 1995; 310: 643-649Crossref PubMed Scopus (89) Google Scholar). protein is to PI-TPβ, it that in also PI-TPβ is mainly in its phosphorylated that in mouse PI-TPβ was predominantly associated with the Golgi (8de Vries K.J. Heinrichs A.A. Cunningham E. Brunink F. Westerman J. Somerharju P.J. Cockcroft S. Wirtz K.W.A. Snoek G.T. Biochem. J. 1995; 310: 643-649Crossref PubMed Scopus (89) Google Scholar). In the that PI-TPβ was associated with the Golgi complex in the SPIβ cells SPIβ cells with GF 109203X, a of PI-TPβ from the Golgi to the was observed similar throughout the was observed for PI-TPβ(S262A) in NIH3T3 cells that Ser262 to be phosphorylated for PI-TPβ to be associated with the Golgi It is to be that the phosphorylation site Ser262 is in PI-TPβ, whereas the phosphorylation site is conserved in identified with the of PI-TP Jr G.M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google contrast to PI-TPβ(S262A) that is overexpressed in mouse is not able to the of SM the of lipid by sphingomyelinase. the protein lipid transfer activity in that the association of PI-TPβ with the Golgi is a prerequisite for PI-TPβ to SM SM and in the Golgi also to the Golgi and phosphorylation of the protein J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). Similar to its PI-TPβ a role in the of vesicles from the Golgi (10Ohashi M. de Vries K.J. Frank R. Snoek G.T. Bankaitis V. Wirtz K.W.A. Huttner W.B. Nature. 1995; 377: 544-547Crossref PubMed Scopus (168) Google Scholar, R. J. Cell Biol. PubMed Scopus Google Scholar). It well that the of SM is to the and of lipid K. D. Rev. Biol. 2000; PubMed Scopus Google Scholar). are PI-TPβ is involved in In shown that PI-TPβ is a substrate for analysis that Ser262 was the phosphorylation site. However, which is to in PI-TPα, was also phosphorylated to a very In a had shown that the PKC-dependent phosphorylation of PI-TPα was to Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). The PI-TPα which is highly to PI-TPβ of Ser262 (9Tanaka S. Hosaka K. J. Biochem. (Tokyo). 1994; 115: 981-984Crossref PubMed Scopus (84) Google Scholar). that phosphorylation of the PI-TP isoforms is for the of their were to the role of Ser262 in the function of PI-TPβ. The peptide of wtPI-TPβ and PI-TPβ(S165A) major which are from the of PI-TPβ(S262A) This that these containing phosphorylated The formation of these is to the presence of tryptic in the acid of the peptide the and to the in van der P. 1991; PubMed Scopus Google Scholar, to to and to The tryptic of PI-TPβ(S262A) showed a peptide containing phosphorylated This was in the peptide of that phosphorylation was to of Ser165 with PI-TPβ(S165A) and the both of which in completely and SM transfer On the other hand, PI-TPβ(S262A) was of the in PI-TPα also an protein Tiel C.M. Westerman J. M. Wirtz K.W.A. Snoek G.T. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus (31) Google Scholar). the it be that is at the as of the of PI-TPα Jr G.M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). it is that the of transfer activity is to the of to with the membrane However, at the that of with the of the protein in E. The for the of transfer activity observed in PI-TPβ(S165A) and After PKC-dependent the phospholipid transfer activity of PI-TPβ was that phosphorylation of Ser262 had no effect The lysates from SPIβ cells two of PI-TPβ that be by analysis it was that 85% of the PI-TPβ at and at of the cells with the PKC inhibitor GF PI-TPβ to strongly that the at the phosphorylated form of PI-TPβ. the from the cells predominantly PI-TPβ at that in PI-TPβ is phosphorylated at that GF and D. P. P. M. V. P. E. F. D. J. J. Biol. Chem. 1991; Full Text PDF PubMed Google Scholar, J. M. M. M. M. S. J. F. Biochem. J. PubMed Scopus Google not which PKC is involved in the phosphorylation of PI-TPβ. it is that PKC is in these cells, it that phosphorylated PI-TPβ in association with the Golgi is a substrate for protein In a PI-TPβ from not be phosphorylated by PKC (8de Vries K.J. Heinrichs A.A. Cunningham E. Brunink F. Westerman J. Somerharju P.J. Cockcroft S. Wirtz K.W.A. Snoek G.T. Biochem. J. 1995; 310: 643-649Crossref PubMed Scopus (89) Google Scholar). protein is to PI-TPβ, it that in also PI-TPβ is mainly in its phosphorylated form. It that in mouse PI-TPβ was predominantly associated with the Golgi (8de Vries K.J. Heinrichs A.A. Cunningham E. Brunink F. Westerman J. Somerharju P.J. Cockcroft S. Wirtz K.W.A. Snoek G.T. Biochem. J. 1995; 310: 643-649Crossref PubMed Scopus (89) Google Scholar). In the that PI-TPβ was associated with the Golgi complex in the SPIβ cells SPIβ cells with GF 109203X, a of PI-TPβ from the Golgi to the was observed similar throughout the was observed for PI-TPβ(S262A) in NIH3T3 cells that Ser262 to be phosphorylated for PI-TPβ to be associated with the Golgi It is to be that the phosphorylation site Ser262 is in PI-TPβ, whereas the phosphorylation site is conserved in identified with the of PI-TP Jr G.M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). In contrast to PI-TPβ(S262A) that is overexpressed in mouse is not able to the of SM the of lipid by sphingomyelinase. the protein lipid transfer activity in that the association of PI-TPβ with the Golgi is a prerequisite for PI-TPβ to SM SM and in the Golgi also to the Golgi and phosphorylation of the protein J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). Similar to its PI-TPβ a role in the of vesicles from the Golgi (10Ohashi M. de Vries K.J. Frank R. Snoek G.T. Bankaitis V. Wirtz K.W.A. Huttner W.B. Nature. 1995; 377: 544-547Crossref PubMed Scopus (168) Google Scholar, R. J. Cell Biol. PubMed Scopus Google Scholar). It well that the of SM is to the and of lipid K. D. Rev. Biol. 2000; PubMed Scopus Google Scholar). are PI-TPβ is involved in
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