REGULATION OF RAS AND RHO GTPASES BY POST-TRANSLATIONAL MODIFICATION: CYSTEINE OXIDATION AND UBIQUITINATION Public Deposited

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  • March 20, 2019
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  • Hobbs, Guy Aaron
    • Affiliation: School of Medicine, Department of Biochemistry and Biophysics
Abstract
  • Ras superfamily GTPases cycle between active GTP-bound and inactive GDP-bound forms to regulate a multitude of cellular processes, including cell growth, differentiation, and apoptosis. The activation state of Ras superfamily GTPases is regulated by protein modulatory agents that accelerate the slow intrinsic rates of GDP dissociation and GTP hydrolysis. In addition, evidence for the redox regulation of Ras superfamily GTPases is growing, and current data supports regulation by the generation of a thiyl radical. Oxidation and reduction events are critical to physiological and pathological processes and are highly regulated. In Ras GTPases, the redox-sensitive cysteine is in the NKCD motif, which is a motif that is critical for nucleotide binding. However, we show that oxidation that occurs absent of the generation of a thiyl radical does not result in Ras activation. In the Rho subfamily of GTPases, there is increasing in vitro and cell-based data supporting the oxidant-mediated regulation of Rho GTPases that contain a redox-sensitive cysteine at the end of the conserved phosphoryl-binding loop motif (GXXXXG[S/T]C). While this motif is distinct from Ras, our data suggest that Rho GTPases are regulated by free radical-generation as well as oxidation of the thiol. Specifically for RhoA and other Rho GTPases that have two cysteine residues within the phosphoryl-binding loop motif (GXXCXG[S/T]C), oxidation can result in internal disulfide bond formation that can inactivate these GTPases. In addition, we characterized relevant cellular oxidation states of RhoA and show that oxidation can result in the activation and deactivation of RhoA. Thus, from data presented herein, it is evident that Ras and Rho family GTPases are uniquely regulated by free radical and non-radical oxidants, and the modulation of activity of the GTPase due to oxidation is dependent on the position(s) of the redox-sensitive thiol. While Ras GTPases are only activated by free radicals because the redox-sensitive thiol does not make interactions with the bound nucleotide, in Rho GTPases, the redox-sensitive thiol makes direct interactions with the bound nucleotide, which results in activation upon modification of the redox-sensitive thiol in RhoA and related GTPases.
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  • In Copyright
Advisor
  • Campbell, Sharon
Degree
  • Doctor of Philosophy
Graduation year
  • 2013
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