Regulatory sequence responsible for insulin destabilization of cytochrome P450 2B1 (CYP2B1) mRNA

Abstract

Diabetes has been reported to increase CYP2E1 (cytochrome P450) and CYP2B1 expression at both the mRNA and protein levels in rat livers. This increase has been attributed to mRNA stabilization and can be reversed by daily insulin treatment. In a previous study, we showed that this hormone directly down-regulates CYP2E1 and 2B1 expression through a post-transcriptional mechanism in rat hepatoma cell lines. We then aimed to identify the molecular mechanisms involved in this regulation. We first identified a 16-mer sequence that we later showed to be the actual functional target of insulin on the rat CYP2E1 mRNA. Similar work was performed with CYP2B1. We first investigated the presence of mRNA-protein interactions. Using cytoplasmic proteins of Fao cells treated or not with insulin (0.1 microM) and the full-length CYP2B1 mRNA as a probe, a major CYP2B1 RNA-protein complex was observed with RNase T1 protection experiments. With the use of different CYP2B1 mRNA probes and by means of competition experiments with antisense oligonucleotides, a protein fixation site was located on a 16-nt sequence in the 5' part of the coding region. This sequence has a hairpin loop structure, shows 80% sequence identity with a structure previously identified on CYP2E1 and is also responsible for the post-transcriptional effects of insulin on this mRNA. Protein(s) bound to both CYP2B1 and CYP2E1 sequences are cytosolic and have an apparent molecular mass of 60 kDa. The protein(s) that bind(s) to both these sequences and the insulin transduction signal involved in this regulation remain(s) to identified.

PPARα‐dependent induction of the energy homeostasis‐regulating nuclear receptor NR1i3 (CAR) in rat hepatocytes: Potential role in starvation adaptation

“…This discrepancy might be attributable to additional posttranscriptional control mechanisms by which phenobarbital could affect CYP2B1 mRNA stability or translation. Evidence that CYP2B1 mRNA abundance is regulated on the posttranscriptional level was recently provided [21]. The synthesis of functional CYP2B protein also depends on the adequate supply with heme whose synthesis is controlled by amino levulinate synthase 1 (ALAS‐1) an enzyme that is induced by phenobarbital in a CAR‐independent manner [22].…”

Molecular basis for a functional role of cytochrome P450 2E1 in non‐alcoholic steatohepatitis

“…11 Such stabilization of the CYP2E1 mRNA is reversed by insulin, 12 as Truong et al revealed the presence of a 16-nucleotide sequence in the 5′ region of the CYP2E1 mRNA that is responsible for insulin-mediated destabilization of the mRNA. 13 In this issue of the Journal of Gastroenterology and Hepatology, Ethirvel and colleagues demonstrate that transgenic mice overexpressing CYP2E1 developed severer experimental steatohepatitis than non-transgenic control mice. 14 Overexpression of CYP2E1 correlated with upregulation of antioxidant enzymes, including superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and heme oxygenase-1 (HO-1) at the mRNA level, presumably in response to increased oxidative stress.…”

“…Such stabilization of the CYP2E1 mRNA is reversed by insulin, 12 as Truong et al. revealed the presence of a 16‐nucleotide sequence in the 5′ region of the CYP2E1 mRNA that is responsible for insulin‐mediated destabilization of the mRNA 13 …”

Cyp2e1

“…The stabilization of the CYP2E1 mRNA that is reversed by insulin (Woodcroft et al, 2002) is less well understood. Recent studies revealed the presence of a 16-nucleotide sequence in the 5Ј region of the CYP2E1 mRNA that is responsible for insulin-mediated destabilization of the mRNA (Truong et al, 2005). This sequence bound a 60-kDa cytosolic protein; however, the identity of this protein and the mechanism by which it is activated by insulin and destabilizes the CYP2E1 mRNA are not known.…”