Heme-Reversible Impairment of CYP2B1/2 Induction in Heme-Depleted Rat Hepatocytes in Primary Culture:Translational Control by a Hepatic alpha-Subunit of the Eukaryotic Initiation Factor Kinase?

摘要

The role of heme in the phenobarbital-mediated induction of CYP2B1/2 was reexamined in rat hepatocytes in monolayer culture,acutely depleted of heme by treatment with either 3,5-dicarbethoxy-2,6-dimethyl-4-ethyl-1,4-dihydropyridine(DDEP)or N-methylprotoporphyrins(NMPP).The findings revealed that such acute hepatic heme depletion markedly impaired CYP2B1/2 protein induction,an effect that was reversible by heme resupplementation.However,TaqMan analyses of hepatic mRNA isolated from these heme-depleted cells revealed that this impairment was not due to faulty transcriptional activation of either CYP2B1 or CYP2B2 gene expression as previously proposed,thereby confirming literature reports that heme is not a transcriptional regulator of the CYP2B1/2 gene.In contrast,the rate of de novo CYP2B1/2 protein synthesis was found to be dramatically inhibited in both DDEP-and NMPP-treated hepatocytes.Concurrently,a marked(>80%)suppression of de novo hepatocellular protein synthesis was also observed,along with a significantly enhanced phosphorylation of the alpha-subunit of the eukaryotic initiation factor elF2(elF2alpha),as monitored by the phosphorylated elF2Ntotal elF2alpha ratio in these heme-depleted cells.Indeed,the parallel reversal of all these three effects by heme supplementation suggests that this impaired CYP2B1 induction most likely stems from blocked translational initiation resulting from the activation of a heme-sensitive elF2alphalpha kinase.Such global suppression of hepatic protein synthesis may disrupt a myriad of vital cellular functions,thereby contributing to the clinical symptoms of acute hepatic heme-deficient states such as the hepatic porphyrias.