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Phosphatidic acid regulates the affinity of the murine phosphatidylinositol 4‐phosphate 5‐kinase‐Iβ for phosphatidylinositol‐4‐phosphate
Author(s) -
JarquinPardo Marta,
Fitzpatrick Abbie,
Galiano Floyd J.,
First Eric A.,
Davis J. Nathan
Publication year - 2006
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.21027
Subject(s) - phosphatidic acid , phosphatidylinositol , phospholipase d , biochemistry , chemistry , phosphorylation , pi , kinase , phosphatidylinositol 4,5 bisphosphate , biophysics , biology , signal transduction , phospholipid , membrane
Type I phosphatidylinositol 4‐phosphate 5‐kinase (PI4P5K) catalyzes the phosphorylation of phosphatidylinositol 4 phosphate [PI(4)P] at carbon 5, producing phosphatidylinositol 4,5 bisphosphate [PI(4,5)P2]. Phosphatidic acid (PA) activates PI4P5K in vitro and plays a central role in the activation of PIP5K pathways in vivo. This report demonstrates that actin fiber formation in murine fibroblasts involves PA activation of PIP5Ks and defines biochemical interactions between PA and the PIP5Ks. Inhibition of phospholipase D production of PA results in the loss of actin fibers. Overexpression of the beta isoform of the type I murine phosphatidylinositol 4‐phosphate 5‐kinase (mPIP5K‐Iβ) maintains actin fiber structure in the face of phospholipase D inhibition. PA activates mPIP5K‐Iβ by direct binding to mPIP5K‐Iβ through both electrostatic and hydrophobic interactions, with the fatty acid acyl chain length and degree of saturation acting as critical determinants of binding and activation. Furthermore, kinetic analysis suggests that phosphorylation of the PI(4)P substrate does not follow classical Michaelis–Menten kinetics. Instead, the kinetic data are consistent with a model in which mPIP5K‐Iβ initially binds to the lipid micelle and subsequently binds the PI(4)P substrate. In addition, the kinetics indicate substrate inhibition, suggesting that mPIP5K‐Iβ contains an inhibitory PI(4)P‐binding site. These results suggest a model in which mPIP5K‐Iβ is surrounded by PI(4)P, but is unable to catalyze its conversion to PI(4,5)P2 unless PA is bound. J. Cell. Biochem. 100: 112–128, 2007. © 2006 Wiley‐Liss, Inc.