Proline Is Required for Transcriptional Control of the Aromatic Hydrocarbon‐inducible P1450 Gene in C57BL/6 Mouse Monolayer‐cultured Hepatocytes

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Abstract
Expression of aryl hydrocarbon hydroxylase (AHH) and the corresponding gene, CyplA1 or P1450 in mice, in C57BL/6 mouse hepatocytes in primary culture was investigated after exposure to benz[a]‐anthracene with respect to proline‐related metabolic regulation. When the cells were cultivated in complete Waymouth MB752/1 (Way), prominent induction of AHH by benz[a]anthracene was observed, whereas the induction was inefficient in the same but proline‐deflcient medium [Way‐(‐pro)]. Constitutive AHH activities decreased with increasing culture period. P1450 gene transcripts were slightly expressed when the medium was changed, independently of whether the cells were cultivated in either Way or Way(‐pro), followed by decrease within 24 h and no apparent induction of AHH. However, treatment with Δ1‐pyrroline‐S‐carboxylic acid (P5C), a biosynthetic precursor for proline, dose‐dependently increased basal AHH activities in the cells cultivated in Way‐(‐pro). Benz[a]anthracene induction of AHH in cells cultivated in Way(‐pro) was additively increased in the presence of P5C as much as with proline. Treatment with o‐aminobenzaldehyde, which inactivates P5C, drastically reduced the induced AHH activities in hepatocytes cultivated in either P5C‐added Way (‐pro) or Way medium. Benz[a]anthracene induced both P1450 and P3450 mRNAs. Neither proline nor P5C increased the induced transcripts within 12 h after the start of benz[a]‐anthracene treatment, but the two compounds increased the amounts of P1450 mRNA found at later time points. After treatment with actinomycin D, the half‐life of the induced P1450 mRNA was approximately 12 h, being independent of the presence of either proline or P5C. Our observations suggest that induction of AHH after treatment with polycyclic aromatic hydrocarbon is dependent on proline‐related metabolism which influences the transcriptional process of P1450 gene expression.