Novel Functions And Physiological Significance Of Ral Interacting Protein - RLIP76 In In Vitro And In Vivo Models

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Title: Novel Functions And Physiological Significance Of Ral Interacting Protein - RLIP76 In In Vitro And In Vivo Models
Author: Zajac, Ewa L
Abstract: This dissertation is divided into four parts . The first part is an introduction of energy dependent xenobiotic transporters and their relevance in multidrug resistance . However , the main focuse is the description of RLIP76 protein , non -ABC transporter , which it the most efficient transporter of glutathione conjugates and anti -cancer drugs . The second part describes materials and methods that were used in two investigated projects . The third part explains the correlation of RLIP76 with signaling protein POB1 and an involvement of these two proteins in apoptosis of lung cancer cells . The effect of recombinant POB1 and its deletion mutant POB11 -512 , on transport activity of RLIP76 towards doxorubicin and dinitrophenyl -glutathione were studied . The results demonstrated that , along with an increased POB1 /RLIP76 ratio , the transport of doxorubicin and dinitrophenyl -glutathione by RLIP76 decreased . Incubation of cancer cells with recombinant POB1 led to an arrest of their cell cycle . Augmentation of cellular POB1 caused increasing intracellular doxorubicin accumulation as well as a decreasing rate of doxorubicin efflux from cells . These results showed for the first time that augmented POB1 inhibits RLIP76 transport function , leading to apoptosis of cancer cells through the accumulation of endogenously formed GSH -conjugates . In the fourth part , it was demonstrated , that RLIP76 , through participation in ligand -receptor complex endocytosis , can mediate insulin -resistance . Insulin sensitivity tests demonstrated that administration of increasing doses of recombinant RLIP76 resulted in development of insulin resistance in RLIP76 wild type (WT ) mice . Double administration of insulin to RLIP76 knockout (KO ) animals caused a dramatic increase of insulin sensitivity . Distribution of hydrocortisone , into RLIP76 KO animals did not have any effect on glucose level . Lack of RLIP76 in KO animals did not change an expression of phosphoenolpyruvate carboxykinase and fructose 1 ,6 -bisphosphatase but specific activities of these enzymes were diminished . Cell culture studies showed that inhibition of RLIP76 resulted in increased glucose uptake , Akt1 phosphorylation , FOXO1 inactivation , and a reduction of endocytosis of insulin and epidermal growth factor . Augmenting RLIP76 by cell transfection caused an opposite effect . These studies demonstrated that RLIP76 antagonizes the effect of insulin , and link oxidative stress with the mechanisms which function to terminate insulin -signaling through ligand -receptor complexes endocytosis .
URI: http : / /hdl .handle .net /10106 /223
Date: 2007-08-23

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Novel Functions And Physiological Significance Of Ral Interacting Protein - RLIP76 In In Vitro And In Vivo Models. Available electronically from http : / /hdl .handle .net /10106 /223 .

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