Phosphatidylinositol kinase is activated in membranes derived from cells treated with epidermal growth factor.

Walker, Duncan; Pike, Linda J.

doi:10.1073/pnas.84.21.7513

Cited by 47 publications

(20 citation statements)

References 36 publications

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“…Evidence for a functional relationship between p2lras and PKC has been previously reported (12). Activation of PKC could be the link between PC and PI metabolism, as judged from previous observations implying a role of PKC in regulation of PI kinase activity (26). The results presented in this study are consistent with a complex regulatory mechanism for phospholipid metabolism in which p2lras proteins have an important regulatory function.…”

Section: Discussionsupporting

confidence: 81%

Diacylglycerol production in Xenopus laevis oocytes after microinjection of p21ras proteins is a consequence of activation of phosphatidylcholine metabolism.

Lacal¹

1990

Mol. Cell. Biol.

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Microinjection of p21Hal proteins into Xenopus laevis oocytes induces a rapid increase of 1,2-diacylglycerol (DAG) levels. The observed alterations in DAG levels were consistent with the ability of the protein to induce maturation, measured by germinal vesicle breakdown (GVBD). Both the increase in DAG levels and GVBD activity were dependent on the ability of the proteins to undergo membrane translocation. Alterations of DAG levels or GVBD activity did not correlate with changes in the levels of inositol phosphates. However, at nminimal doses sufficient to achieve maximal biological response, a biphasic increase in the amounts of phosphocholine and CDP-choline was observed. The first burst of phosphocholine and CDP-choline preceded the increase in DAG levels. The second peak paralleled the appearance of DAG. Choline kinase activity was also increased in oocyte extracts after p2l1 microinjection. These results suggest that both the synthesis and degradation of phosphatidylcholine are activated after microinjection of ras proteins into Xenopus oocytes, resulting in a net production of DAG.ras genes code for 21-kilodalton proteins that are thought to play a critical role in the regulation of signal transduction mechanisms in a variety of cell systems (for reviews, see references 1 and 14). The p21 proteins bind GTP and GDP with high affinity and are able to hydrolyze GTP with a very low intrinsic efficiency. Recently, a cytoplasmic protein designated GTPase-activating protein has been shown to increase both in vivo and in vitro the intrinsic GTPase of the normal protein by a factor of at least 100-fold, with no effect on the GTPase activity of mutant p21 proteins (23). Singlepoint mutations at defined positions alter biochemical and biological properties of the p2lras products. Such alterations are thought to play an important role in malignant transformation. Despite intensive investigations to determine the pathway in which ras proteins function, their role as regulating factors remains elusive.ras genes are highly conserved in evolution from yeast to mammals, with a great degree of homology of their products (1, 14). Several systems have been used to investigate the biochemical properties of the ras proteins, including yeast, Dictyostelium discoideum, Drosophila melanogaster, and Xenopus laevis. Microinjection of recombinant p2lras proteins into X. laevis oocytes has been shown to induce maturation. This effect seems to be different from the mechanism induced by progesterone, since it is independent of the regulation of the levels of cyclic AMP (3). I have used X. laevis oocytes for analysis of alterations in the phospholipid metabolism induced by ras proteins. In a previous study, it was reported that microinjection of ras proteins into Xenopus oocytes induced a rapid formation of 1,2-diacylglycerol (DAG) and inositol phosphates (IPs) (11). However, there are some controversial studies on the involvement of p2lras proteins in the regulation of a phospholipase C (PLC) (7,9,13,21,25,29). In this study, the relati...

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Section: Discussionsupporting

confidence: 81%

Diacylglycerol production in Xenopus laevis oocytes after microinjection of p21ras proteins is a consequence of activation of phosphatidylcholine metabolism.

Lacal¹

1990

Mol. Cell. Biol.

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“…This speculation is further supported by gel exclusion chromatography analysis, which suggests that the PIPK␤ exists as large multimeric complexes. 3 Previous studies indicate that one or more phosphoinositide kinase activities partially co-purify with EGFR (16) and that EGF stimulates phosphoinositide kinase activity in several cell types (23). To determine whether PIPK␤ and PIPK␤-truncated are physically associated with EGFR, cells expressing PIPK␤, PIPK␤-truncated, or both constructs were stimulated with EGF, and immunoprecipitation studies were carried out as described in the legend to Fig.…”

Section: Phosphatidylinositol-4-phosphate 5-kinase-1␤ and Endocytosismentioning

confidence: 99%

Phosphatidylinositol-4-phosphate 5-Kinase-1β Is Essential for Epidermal Growth Factor Receptor-mediated Endocytosis

Barbieri¹,

Heath²,

Peters³

et al. 2001

Journal of Biological Chemistry

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Phosphatidylinositol-4,5-bisphosphate (PIP 2 ) is known to play an important role in signal transduction and membrane trafficking. We show that one enzyme responsible for PIP 2 production, phosphatidylinositol-4-phosphate 5-kinase type 1␤ (PIPK␤), is essential for epidermal growth factor receptor (EGFR)-mediated endocytosis. Expression of murine PIPK␤ in NR6 cells expressing EGFR strikingly increased receptor internalization. Moreover, the kinase was shown to form an immunoprecipitable complex with EGFR. Expression of either a truncated kinase or a kinase dead mutant inhibited EGFR endocytosis and also blocked the membrane recruitment of PIPK␤ and both clathrin light chain and dynamin. Our results delineate a novel mechanism by which PIPK␤ regulates receptor-mediated endocytosis and receptor tyrosine kinase membrane traffic.

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“…Most notably, PI-4-P is the precursor for PI-4,5-P 2 , which is hydrolyzed by phospholipase C into inositol 1,4,5-trisphosphate and diacylglycerol, important mediators of intracellular calcium release and protein kinase C activity, respectively (40). PI-4-P synthesis can be activated by ligand binding to membrane receptors for neurotransmitters, hormones, and growth factors (41)(42)(43). Therefore, one might expect that the lipid kinase activity of RAFT1 will be tightly regulated.…”

Section: Rapamycin-fkbp12mentioning

confidence: 99%

The Rapamycin and FKBP12 Target (RAFT) Displays Phosphatidylinositol 4-Kinase Activity

Sabatini

Pierchala

Barrow

et al. 1995

Journal of Biological Chemistry

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The immunosuppressant rapamycin prevents cell cycle progression in several mammalian cell lines and the yeast Saccharomyces cerevisiae. In mammalian cells, rapamycin binds to the small FK506-binding protein, FKBP12, allowing the drug-receptor complex to interact with the 289-kDa RAFT1/FRAP proteins. These proteins, along with their yeast homologs, TOR1/DRR1 and TOR2/ DRR2, contain a C-terminal domain with amino acid homology to several phosphatidylinositol (PI) 4-and 3-kinases. However, no direct demonstration of kinase activity for this family of proteins has been reported. We now show that RAFT1, immunoprecipitated from rat brain and MG63 and HEK293 cells, contains PI 4-kinase activity and that rapamycin-FKBP12 has no effect on this activity. Thus, it is likely that, in vivo, rapamycin does not directly inhibit the PI 4-kinase activity and affects the RAFT1/FRAP protein through another mechanism.The ability of rapamycin, a potent immunosuppressant drug, to arrest a variety of mammalian cells and the yeast Saccharomyces cerevisiae in the G 1 stage of the cell cycle (1-4) has made it a valuable tool for studying the intermediate signaling events that convey mitogenic stimuli to the cell nucleus.Rapamycin binds with low nanomolar affinity to the FK506-binding protein (FKBP12), a small soluble protein that is also the intracellular receptor for FK506, a structurally similar immunosuppressant (5-7). Analogous to the mechanism of action of FK506, it is the drug-receptor complex that mediates the effects of the rapamycin, which include inhibition of the 70-kDa S6 kinase (8, 9) and of several cyclin-dependent kinases (2-4). The immunosuppressant effects of FK506 derive from the binding of the FK506-FKBP12 complex to the calcium-activated phosphatase, calcineurin, and the resulting inhibition of its activity (10). Although rapamycin binds to FKBP12, the complex does not interact with calcineurin. Instead, rapamycin-FKBP12 binds to a recently purified and molecularly cloned protein designated rapamycin and FKBP12 target-1 (RAFT1) in rats (11) and FKBP-rapamycin-associated protein (FRAP) in humans (12). Several other groups have subsequently purified and/or cloned the same protein (13-15).RAFT1 is a 2549-amino acid protein with a predicted molecular mass of 289 kDa and is thought to be a mammalian homolog of the products of two yeast genes, TOR1/DRR1 and TOR2/DRR2, which when mutated lead to dominant rapamycin resistance in yeast (1, 16, 17). The C-terminal 600-amino acid domain of RAFT1 and the yeast TOR proteins has homology to several PI 1 kinases, including the p110 subunit of mammalian PI 3-kinase (18); the yeast PI 3-kinase VPS34 (19); two yeast PI 4-kinases, STT4 (20) and PIK1 (21, 22); and the recently cloned mammalian PI 4-kinase, PI4K␣ (23). This homology to known lipid kinases has lead to the suggestion that RAFT1 may also be a PI kinase and that rapamycin exerts its effects by inhibiting this activity (11). However, no direct demonstration of kinase activity for RAFT1/FRAP or the yeast TOR proteins has been r...

show abstract

Phosphatidylinositol kinase is activated in membranes derived from cells treated with epidermal growth factor.

Cited by 47 publications

References 36 publications

Diacylglycerol production in Xenopus laevis oocytes after microinjection of p21ras proteins is a consequence of activation of phosphatidylcholine metabolism.

Diacylglycerol production in Xenopus laevis oocytes after microinjection of p21ras proteins is a consequence of activation of phosphatidylcholine metabolism.

Phosphatidylinositol-4-phosphate 5-Kinase-1β Is Essential for Epidermal Growth Factor Receptor-mediated Endocytosis

The Rapamycin and FKBP12 Target (RAFT) Displays Phosphatidylinositol 4-Kinase Activity

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