Modulation of the substrate specificity of the mammalian phosphatidylinositol 3-kinase by cholesterol sulfate and sulfatide.
The substrate specificity of the purified, mammalian phosphatidylinositol 3-kinase is subject to modulation by detergents, which are able to switch substrate specificity in vitro in favor of PtdInsP2. This effect of the detergents is due to an activation of the phosphatidylinositol biphosphate 3-kinase activity, while the phosphatidylinositol 3-kinase activity is inhibited. The selective inhibition of the phosphatidylinositol 3-kinase activity (p110 alpha/p85 alpha) is shown here also to be observed by employing cholesterol sulfate or sulfatide at low micromolar concentrations, whereas cholesterol and androsterone sulfate fail to inhibit. These naturally occurring sulfated lipids have at these concentrations no effect on the phosphatidylinositol bisphosphate 3-kinase activity but inhibit the manganese-dependent intrinsic protein kinase activity, thus switching substrate specificity toward the more highly phosphorylated inositol lipids. Cholesterol sulfate and sulfatide inhibit the free catalytic subunit p110 alpha but fail to inhibit the homologous phosphatidylinositol 3-kinase from Saccharomyces cerevisiae (Vps34p), suggesting that these sulfated lipids act specifically on the mammalian phosphatidylinositol 3-kinase. Consistent with this specificity, the regulatory subunit (p85), which is not conserved in the yeast enzyme, is found to play an important role for the affinity of these inhibitors. The implications for the phosphatidylinositol 3-kinase activity in vivo are discussed.
Woscholski
R
R
Protein Phosphorylation Laboratory, Imperial Cancer Research Fund, London, UK.
Kodaki
T
T
Palmer
R H
RH
Waterfield
M D
MD
Parker
P J
PJ
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002384:Catalysis; D002460:Cell Line; D002784:Cholesterol / Q000031:analogs & derivatives; D002788:Cholesterol Esters / Q000494:pharmacology*; D003902:Detergents / Q000494:pharmacology*; D019869:Phosphatidylinositol 3-Kinases; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D018411:Spodoptera; D013379:Substrate Specificity; D013433:Sulfoglycosphingolipids / Q000494:pharmacology*
D000255:Adenosine Triphosphate; D002784:Cholesterol; D002788:Cholesterol Esters; D003902:Detergents; D019869:Phosphatidylinositol 3-Kinases; D018129:Phosphatidylinositol Phosphates; D017853:Phosphotransferases (Alcohol Group Acceptor); D011994:Recombinant Proteins; D013433:Sulfoglycosphingolipids; C095807:cholesterol sulfatide; C060980:phosphatidylinositol 3,4-diphosphate; C007045:cholesteryl sulfate