EFFECT OF DIETARY-PROTEIN RESTRICTION ON LIVER TRANSCRIPTION FACTORS

The transcription of several genes that are preferentially expressed i n the liver, including the serum albumin, transthyretin and carbamyl p hosphate synthetase-I genes, is specifically decreased in animals cons uming inadequate amounts of dietary protein. The high level of transcr iption of these genes in the liver is directed in part by a number of liver-enriched transcription factors, including hepatocyte nuclear fac tors (HNF)-1, -3, and -4, and proteins of the CCAAT/enhancer-binding p rotein (C/EBP) family. In the present study, we investigated the possi bility that the co-ordinate decrease in transcription of the nutrition ally sensitive genes in protein-deprived rats results from altered act ivity of one or more of the liver-enriched transcription factors. For HNF-4, Western blots indicated no change in the level of nuclear HNF-4 protein in liver of protein-deprived animals, whereas we observed a 4 0% reduction in the DNA binding activity of HNF-4 as measured by elect rophoretic mobility shift assay (EMSA). Furthermore, the binding affin ity of HNF4 for DNA was unaltered by dietary protein deprivation, whil e the number of HNF-4 molecules able to bind to DNA (B-max) was reduce d, as determined by Scatchard analysis. This indicates that in the pro tein-restricted rats a portion of the pool of HNF4 protein is inactiva ted or otherwise prevented from binding to DNA. The overall DNA bindin g activity of C/EBP alpha and beta Was increased in protein-restricted animals. This change occurred in the absence of a change in the amoun t of the full-length forms of these two proteins, quantified by Wester n blotting. Interestingly, dietary protein restriction specifically in creased the level of a truncated form of C/EBP beta (liver-enriched tr anscriptional inhibitory protein, LIP), which is a potent dominant neg ative inhibitor of C/EBP function. Analysis of HNF-3 DNA-binding activ ity by EMSA revealed that HNF-3 alpha and beta DNA binding was increas ed and that HNF-3 gamma DNA-binding activity was unchanged in protein- restricted animals. We also detected two apparently novel shift comple xes with the HNF-3 probe by EMSA, both of which were decreased in prot ein-restricted animals. HNF-1 DNA-binding activity was increased by di etary protein restriction. We also examined the effect of protein rest riction on the DNA-binding activity of two ubiquitous transcription fa ctors, NF1 and Spl. The DNA binding activity of the major NF1 isoforms was unchanged whereas the binding activity of Sp 1 was increased in t he protein-restricted animals. In summary, restriction of dietary prot ein resulted in a number of specific changes in the DNA-binding activi ty of various transcription factors. Because transcriptional activatio n typically involves the synergistic action of more than one transcrip tion factor, small changes in the amount/activity of several factors c ould have a strong net effect on the transcription of many genes.