GLYCINE N-METHYLTRANSFERASE IS A MEDIATOR OF CYTOCHROME P4501A1 GENE-EXPRESSION

Cytochrome P4501A1, the isozyme most closely approximating aryl hydrocarbon hydroxylase activity under conditions of induction, is thought to be regulated by several trans-acting factors, including the 4S polycyclic aromatic hydrocarbon-binding protein; this protein has recently been identified as glycine N-methyltransferase (Raha et al, (1994) J. Biol. Chem, 269, 5750-5756), Previous studies had shown that partially purified liver preparations containing the 4S binding protein interacted with 5'-flanking regions of the cytochrome P4501A1 gene, Consequently, the ability of the 4S binding protein to serve as a mediator in the regulation of the cytochrome P4501A1 gene was investigated further, Introduction of an antisense 24-mer oligonucleotide to glycine N-methyltransferase cDNA into rat hepatoma H4IIE cells by lipofectin resulted in a 60% reduction in the benzo(a)pyrene-mediated induction of ethoxyresorufin-O-deethylase activity and protein over the sense and scrambled antisense oligonucleotide controls. In addition, the antisense oligonucleotide caused a marked reduction in the steady-state level of cytochrome P4501A1 mRNA; no such effect was observed with the sense oligonucleotide. Introduction of GNMT polyclonal antibodies into H4IIE cells by a streptolysin-O permeabilization technique markedly reduced both benzo(a)pyrene-binding and benzo(a)pyrene-induced ethoxyresorufin-O-deethylase activities, but had no effect on 2,3,7,8-tetrachlorodibenzo-p-dioxin induction, Collectively, these findings suggest that, in addition to the Ah (dioxin) receptor, glycine N-methyltransferase appears to be both a polycyclic aromatic hydrocarbon-binding protein and a mediator of the induction of the cytochrome P4501A1 gene by polycyclic hydrocarbons such as benzo(a)pyrene. (C) 1995 Academic Press, Inc.