Regulation ofras signaling by Gs-alpha.
Item
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Title
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Regulation ofras signaling by Gs-alpha.
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Identifier
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AAI9417446
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identifier
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9417446
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Creator
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Chen, Jianghao.
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Contributor
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Adviser: Ravi Iyengar
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Date
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1994
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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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Biology, Molecular | Biology, Cell
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Abstract
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Mutant activated {dollar}\alpha{dollar}-subunits of G proteins ({dollar}\alpha\sp*{dollar}) have been postulated to be oncoproteins. {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar} and {dollar}\alpha\sb{lcub}\rm i{rcub}\sp{lcub}*{rcub}{dollar} have been found in human pituitary and ovarian tumors respectively. {dollar}\alpha\sb{lcub}\rm i{rcub}\sp{lcub}*{rcub}{dollar}, {dollar}\alpha\sb{lcub}\rm o{rcub}\sp{lcub}*{rcub}{dollar} and {dollar}\alpha\sb{lcub}\rm q{rcub}\sp{lcub}*{rcub}{dollar} are oncogenic as assessed by in vivo and in vitro experiments. Although {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar} was the first mutant {dollar}\alpha{dollar}-subunit identified in tumors, it has not been shown to have oncogenic potential. I have examined whether {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar} had this transforming ability. Two cell types, NIH-3T3 and RAT-1, that show different proliferative responses to cAMP were used in my study. In NIH-3T3 cells, increases or decreases in cellular cAMP levels alone do not have any effects on cell proliferation. On the other hand, lowering of cellular cAMP levels strongly stimulates cell proliferation in RAT-1 cells. Clonal NIH-3T3 and RAT-1 cell lines that expressed {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar} were established and examined. Expression of {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar} in these cell lines modestly raises cellular cAMP levels; however, this expression did not result in transformation of these cells and did not affect mitogenesis. {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar} may be nontransforming by itself because it is a weak oncogene that functions best in the presence of another oncogene. Hence, I tested whether {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar} promotes H-ras-induced transformation. Surprisingly, {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar} substantially blocked ras-induced transformation of NIH-3T3 cells and inhibited ras-stimulated DNA synthesis in these cells. Expression of {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar} did not affect transfection efficiencies or expression of H-ras protein. Similarly, H-ras-induced transformation of RAT-1 cells is also blocked by {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar}.;I studied the possible mechanisms which underlie the inhibitory effects of {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar} on ras-induced transformation. It has been demonstrated that proliferative signals from ras are routed through the MAP kinase pathway. I found that ras-stimulated MAP kinase activity was significantly reduced by {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar}. Furthermore, I have also shown that a cAMP analog, 8 Br-cAMP, suppressed ras-induced transformation of NIH-3T3 cells and decreased MAP kinase activity. The effect of {dollar}\alpha\sb{lcub}\rm s{rcub}\sp{lcub}*{rcub}{dollar} on the ras-induced transformation can be relieved by dominant negative regulatory subunits of protein kinase A. These data suggest that {dollar}\alpha\sb{lcub}\rm s{rcub}{dollar} inhibits proliferative signals from ras by stimulating cAMP production and activating protein kinase A. The antiproliferative effect of {dollar}\alpha\sb{lcub}\rm s{rcub}{dollar} may be achieved by attenuation of ras signaling through the MAP kinase pathway.
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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.
