Regulatory mechanisms of gene expression of hepatic enzyme by insulin and fructose

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Regulation of the expression of the hepatic L-type pyruvate kinase gene by insulin and fructose was studied in diabetic rats. Insulin stimulated transcription of the pyruvate kinase L gene. Cycloheximide inhibited the induction caused by insulin, suggesting that insulin may stimulate transcription of the pyruvate kinase L gene by stimulating synthesis of some unknown protein. On the other hand, feeding fructose had no effect on transcription of the pyruvate kinase L gene. Since increases in the levels of putative nuclear RNA precursor species of the pyruvate kinase L after fructose feeding preceded changes in the levels of cytosolic pyruvate kinase L mRNA, fructose may increase the levels of pyruvate kinase L mRNA by stabilizing nuclear RNA species. The effect of fructose on pyruvate kinase L was not mediated by hormones such as glucagon, glucocorticoid and thyroid hormones, and thus seems to be directly on the liver.Comparison of the nucleotide sequence of rat R-type isozyme mRNA with that of the L-type isozyme showed that the R-type isozyme mRNA had an identical nucleotide sequence to that of the L-type except in the 5' terminal region including the coding sequence: the sequence upstream of the 5th coding residue of the L-type was replaced by a 98-nucleotide coding sequence plus a 5' untranslated region in the R-type isozyme. By sequence analysis of a genomic clone of LPK30 twelve exons were identified, of which the first and second exons coded the R- and L-specific sequences, respectively. The remaining exons present downstream coded amino acids common to the two isozymes. Thus, we suggest that the two isozyme mRNAs are generated by use of different promoters.