EFFECT OF AMINO-ACID LIMITATION ON THE EXPRESSION OF 19 GENES IN RAT HEPATOMA-CELLS

We showed previously that the abundance of serum albumin mRNA is decre ased in H4-II-E rat hepatoma cells limited for a single essential amin o acid (phenylalanine, methionine, leucine, or tryptophan). To define the specificity of this phenomenon, we examined the effect of amino ac id limitation on the abundance of mRNAs for 19 genes in the H4-II-E ce lls. These genes included six genes whose expression is either complet ely liver-specific or highly enriched in the liver compared with other tissues [albumin, transthyretin (TTR), transferrin, carbamyl phosphat e synthetase-I, urate oxidase, class I alcohol dehydrogenase], as well as a number of ubiquitously expressed ''housekeeping'' genes. The res ults indicated that the 19 genes could be divided into three classes b ased on their response to amino acid limitation. Class I genes (the si x liver-specific genes and alpha-tubulin) exhibit decreased expression in response to amino acid limitation. The expression of class II gene s [beta(2)- microglobulin, hypoxanthine-guanine phosphoribosyl transfe rase (HPRT), H-ferritin, ubiquitin (UbB), insulin-like growth factor b inding protein-4, HNF-1 alpha] is not significantly affected by amino acid limitation. Class III genes [gadd153, beta-actin, ubiquitin (UbC) , phosphoglycerate kinase-1, C/EBP alpha, C/EBP beta] exhibit increase d expression in response to amino acid limitation. Thus, specific indu ctive as well as repressive effects on gene expression are quite commo n in amino acid-limited cells. The observation that all six genes whos e expression is liver-specific exhibited decreased expression in amino acid-limited cells suggests a common mode of regulation of these gene s by amino acid availability. The strong induction by amino acid limit ation of the C/EBP inhibitor gadd153 is of interest in this regard, as increased levels of gadd153 could interfere with C/EBP, which is requ ired for high expression of most liver-specific genes. To investigate further the molecular mechanism for the decrease in albumin mRNA abund ance, albumin nuclear transcript levels were quantified in control and tryptophan-limited cells. Tryptophan limitation caused a decrease in albumin nuclear transcript abundance, and this decrease preceded the d ecrease in albumin mRNA, suggesting that the decrease in albumin mRNA was caused at least partly by a decrease in albumin gene transcription . Additional experiments with actinomycin D indicated that albumin mRN A was also destabilized in the tryptophan-limited cells. Thus, the ove rall results indicate that the decrease in albumin mRNA in the tryptop han-limited cells is caused by a specific decrease in albumin nuclear transcript abundance and destabilization of albumin mRNA.