Survival signals in human cancer cells mediated by phospholipase D and mTOR
Item
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Title
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Survival signals in human cancer cells mediated by phospholipase D and mTOR
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Identifier
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d_2009_2013:5792cf71bbeb:10029
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identifier
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10017
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Creator
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Toschi, Alfredo,
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Contributor
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David Foster
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Date
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2009
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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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Molecular biology
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Abstract
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Phospholipase D (PLD), which is commonly elevated in renal and other cancers, provides a survival signal that suppresses apoptosis induced by serum deprivation in renal cancer cells. Hypoxia-Inducible Factors alpha (HIFalpha), important effectors of hypoxic response, have been shown to play a pivotal role in the tumorigenesis of renal cancer cells that lack the von Hipple Lindau tumor suppressor gene (VHL), a critical mediator of HIFalpha proteolytic degradation. We report here a role for PLD as another regulatory component of HIFalpha expression in renal cancer cells where accumulation of both HIF1alpha and HIF2alpha in require functional PLD for efficient translation, independently from pVHL expression.;The expression of HIF1alpha has been widely shown to be dependent on mTOR, the mammalian target of rapamycin and its sensitivity to rapamycin has been established. In contrast, HIF2alpha has been reported to be insensitive to rapamycin. mTOR, a critical node for control of cell growth and survival, exists in two complexes, mTORC1 and mTORC2, which are differentially sensitive to rapamycin. We report here that while HIF2alpha is insensitive to rapamycin in renal cancer cells, HIF2alpha expression is still dependent on mTOR expression and we are able to show in the fourth chapter of this work that while HIF1alpha is dependent on both mTORC1 and mTORC2, HIF2alpha depends solely on the rapamycin resistant mTORC2.;However, while much is known about the regulation of mTORC1, little is known about the regulation of mTORC2. PLD and its metabolite phosphatidic acid (PA) have been implicated in the regulation mTOR but its role has been controversial. In light of the concomitant regulation by PLD and mTOR of HIFalpha expression, we investigated the role of PLD in the regulation mTOR activity. We report in the fifth chapter of this work that PA, in competition with rapamycin, is required for functional mTORC1 and mTORC2 complex formation. Suppression of PLD prevented phosphorylation of the mTORC1 substrate S6 kinase at Thr389 and the mTORC2 substrate Akt at Ser473. Suppression of PLD also blocked insulin-stimulated phosphorylation of Akt and the mTORC2- and Akt-dependent phosphorylation of PRAS40 indicating that PA is required for the association of mTOR with Raptor to form mTORC1 and mTOR with Rictor to form mTORC2. The effect of PA was competitive with rapamycin with much higher concentrations of rapamycin needed to compete with the PA-mTORC2 interaction than with the PA-mTORC1 interaction. However, suppressing PA production substantially increased the sensitivity of mTORC2 to rapamycin. The data provided here reveal a PA requirement for the stabilization of both mTORC1 and mTORC2 complexes. The competition between PA and rapamycin for mTOR suggests a mechanism for the suppression of mTOR by rapamycin and explains the rapamycin resistance of mTORC2 and HIF2alpha.;The last part of this work implicates PLD in the regulation of another hallmark of cancer cells: aerobic glycolisis. The metabolic shift from oxidative phosphorylation to aerobic glycolysis, also known as the "Warburg effect", is thought to provide a means for cancer cells to survive under conditions where oxygen is limited and to generate metabolites necessary for cell growth. A shift to aerobic glycolysis is also a response to hypoxia, which stimulates the accumulation of HIFalpha necessary for the expression of proteins involved in glucose uptake and glycolysis.;We are able to show here that the metabolic shift from oxidative phosphorylation to aerobic glycolysis in human cancer cells is dependent on the elevated PLD activity in breast and renal cancer cells. Intriguingly, the effect of PLD on the Warburg phenotype was dependent on mTORC1 in breast cancer cells and on mTORC2 in renal cancer cells, consistently with a role for PLD in activating mTOR. We are able to conclude that elevated PLD signaling, which is common in human cancer cells, is critical for the activation of mTOR complexes and accounts for mTORC2 insensitivity to Rapamycin. Moreover, elevated PLD activity is required for the expression of HIFalpha and the consequent transcriptional activation of many genes involved in tumorigenesis including genes involved in the metabolic shift to aerobic glycolysis. Taken together, this data provides evidence that targeting PLD could prove therapeutically significant in cancers with elevated PLD activity such as renal and breast cancer.
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Type
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dissertation
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Source
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2009_2013.csv
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degree
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Ph.D.
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Program
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Biology
