EGF andv-Fps-induced diglyceride production: Regulation via phospholipase D and phosphatidic acid phosphatase.
Item
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Title
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EGF andv-Fps-induced diglyceride production: Regulation via phospholipase D and phosphatidic acid phosphatase.
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Identifier
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AAI9707110
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identifier
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9707110
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Creator
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Jiang, Youwei.
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Contributor
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Adviser: David A. Foster
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Date
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1996
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Language
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English
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Publisher
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City University of New York.
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Subject
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Chemistry, Biochemistry
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Abstract
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Activating the protein-tyrosine kinase v-Fps results in a rapid increase in diglyceride (DG) in cells expressing a temperature-sensitive v-Fps (ts v-Fps). The v-Fps-induced increase in DG is detected only when cellular phospholipids are prelabeled with ({dollar}\sp3{dollar}H) -myristate, which is incorporated primarily into phosphatidylcholine (PC). PC is predominantly a substrate for phospholipase D (PLD). The primary metabolite of PLD is phosphatidic acid (PA) which can be converted to DG by phosphatidic acid phosphatase (PAP). The cells expressing ts v-Fps have elevated levels of PA and PLD activity at both permissive and non-permissive temperatures for v-Fps relative to the parental 3Y1 cells. These data suggest that PLD is constitutive activated in v-Fps-transformed cells, and the activated PLD alone is not sufficient to regulate v-Fps-induced increase in DG. If we inhibit the conversion of PA to DG, the v-Fps-induced increase in DG is blocked. Upon shifting from the non-permissive to the permissive temperature for ts v-Fps, the membrane PAP activity is increased. These data suggest that PAP regulates v-Fps-induced increase in DG. All above evidences indicate that v-Fps induces activation of PLD/PAP signaling pathway to generate DG.;Epidermal growth factor (EGF), like v-Fps, also activates PLD/PAP signaling pathway to generate DG. To further establish PAP as a regulatory site for DG production, we investigated the effect of EGF on PAP activity in A431 cells, which overexpress EGF receptors (EGFR). Upon EGF stimulation, PAP activity in cell lysates is elevated. Protein Kinase C (PKC) inhibitors block the EGF-induced increase in PAP activity. Consistent with a role for PKC, EGF induces a sustained increase in PAP activity in PKC {dollar}\varepsilon{dollar} immunoprecipitates, but not in PKC {dollar}\alpha{dollar} or PKC {dollar}\zeta{dollar} immunoprecipitates. Concomitant with the increased PAP activity in PKC {dollar}\varepsilon,{dollar} there is a corresponding decrease in PAP activity in EGFR immunoprecipitates. AG1478 induces a dramatic decrease in PAP activity in both EGFR and PKC {dollar}\varepsilon{dollar} immunoprecipitates. It correspondingly induces a decrease of PAP activity in the membrane fractions and an increase of PAP activity in the cytosol fractions. AG1478 also blockes EGF-induced DG production. Based on these data, we propose a novel mechanism for PAP activity in regulating EGF-induced DG production, in which EGF induces PAP to dissociate from membrane EGFR and associate with PKC {dollar}\varepsilon{dollar} to localize PAP to PA substrate.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
