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This investigation was conducted to determine the comparative effect of structurally diverse peroxisome proliferators (PP) on the two-dimensional protein pattern of rat liver whole homogenates. Perfluoro-n-decanoic acid (PFDA), perfluoro-n-octanoic acid (PFOA), clofibrate, and di(2-ethylhexyl)phthalate (DEHP) are all known to cause the proliferation of hepatic peroxisomes and the induction of peroxisomal beta-oxidative and microsomal omega-oxidative enzymes. To clarify the mechanistic differences between these compounds with regard to the liver, we examined the unique patterns of protein alteration produced by in vivo exposure to them. Following exposure to various doses, whole liver homogenates were prepared and separated by two-dimensional gel electrophoresis (2DE) using the ISO-DALT system. Stained gels were digitized and protein patterns analyzed using the Kepler 2D gel analysis system. Immunoglobulin heavy chain binding protein (BiP), also known as 78-kDa glucose-regulated protein (Grp78), was identified immunologically and by comigration of recombinant Grp78. BiP is a luminal endoplasmic reticular protein that functions in the assembly and folding of nascent proteins as they enter the ER. The present results suggest a selective posttranslational modification of BiP following PFDA exposure. Single-dose exposure to PFDA was associated with a notable charge modification of BiP that persists up to 30 days. PFOA, clofibrate, and DEHP had less effect in this regard. The identity of BiP/Grp78 as the halothane hepatitis-associated trifluoroacetylated protein was also demonstrated. The nature of this PFDA-associated protein modification (reactive metabolite conjugation, abnormal ribosylation, or phosphorylation) is currently under investigation.(ABSTRACT TRUNCATED AT 250 WORDS)

This investigation was conducted to determine the comparative effect of structurally diverse peroxisome proliferators (PP) on the two-dimensional protein pattern of rat liver whole homogenates. Perfluoro-n-decanoic acid (PFDA), perfluoro-n-octanoic acid (PFOA), clofibrate, and di(2-ethylhexyl)phthalate(DEHP) are all known to cause the proliferation of hepatic peroxisomes and the induction of peroxisomal beta-oxidative and microsomal omega-oxidative enzymes. To detect potential differences between these compounds with regard to the liver, we examined the unique patterns of protein alteration produced by in vivo exposure to them. Following exposure to various doses, whole liver homogenates were prepared and separated by two-dimensional gel electrophoresis (2DE) using the ISO-DALT System. Stained gels were digitized and protein patterns analyzed using the Kepler 2D Gel Analysis System. Immunoglobulin heavy-chain binding protein (BiP), also known as 78 kD glucose regulated protein (Grp78), was identified immunologically and by comigration of recombinant Grp78. BiP is a luminal endoplasmic reticular (ER) protein that functions in the assembly and folding of nascent proteins as they enter the ER. The present results suggest a selective posttranslational modification of BiP following PFDA exposure. Single-dose exposure to PFDA was associated with a notable charge-modification of BiP that persists up to 30 days. PFOA, clofibrate, and DEHP had less effect in this regard. Our data suggest the likely nature of this PFDA-associated protein modification is associated with protein-phosphorylation. These results document the unique nature of PFDA's hepatotoxicity with respect to classic peroxisome proliferators and support the utility of 2D gel analysis in toxicity testing.