PNNL

Abstract

We have previously reported that CYP3A crosslinks with polyubiquitinated proteins in microsomes from nicardipine-treated rats in a process distinct from the classical polyubiquitination process. In order to further examine the role of proteasome in CYP3A degradation, we investigated the affects of proteasome inhibitors lactacystin, MG132, proteasome inhibitor 1 and hemin in primary cultures of rat hepatocytes. Opposite to what would be expected if the proteasome was responsible for degradation for CYP3A, the proteasome inhibitors caused a reduction of the major ~55 kDa CYP3A band. Hemin caused an increase in high molecular mass (HMM) CYP3A bands in microsomes. Since the other proteasome inhibitors did not have a similar effect, it seems unlikely that these HMM bands were the result of proteasome inhibition. Although we were unable to detect CYP3A in cytosolic fractions, an increase in the HMM CYP3A bands was also observed in a detergent-insoluble fraction of the 10,000 g pellet. The present of HMM CYP3A in this fraction suggests that CYP3A is forming a large protein complex that is likely to be degraded by autophagic pathways independent of the proteasome.