No abstract

The impact of nutrients on gene expression has become an area of considerable interest as the number of genes coding for key regulatory proteins in metabolic pathways are studied in detail. This has been greatly aided by a number of new techniques developed to study gene transcription in animals. We will use as an example studies on the regulation of transcription of the gene coding for P-enolpyruvate carboxykinase, a key enzyme in hepatic and renal gluconeogenesis. The promoter for P-enolpyruvate carboxykinase contains a number of regulatory elements within 500 bp of the start-site of gene transcription that are required for the response of the gene to metabolic signals. These elements bind tissue-specific transcription factors in complex patterns of interactions, which result in the coordinate control of P-enolpyruvate carboxykinase gene expression. An analysis of the regulation of transcription of this gene involves the use of a number of techniques ranging from gene transfection into cells in culture to the introduction of chimeric genes containing the P-enolpyruvate carboxykinase promoter into transgenic mice. This review presents a progress report on the current status of research on the nutritional and hormonal regulation of transcription of the P-enolpyruvate carboxykinase gene.

Gluconeogenesis in the chicken has unique features due in part to the presence of two isozymes of PEPCK, a cytosolic form, PEPCK-C, and a mitochondrial form, PEPCK-M, which have novel patterns of expression. Here we show that, in contrast to mammals, in which PEPCK-C is not present in liver until after birth, avian PEPCK-C is expressed throughout embryonic life with mRNA levels gradually decreasing as development proceeds and becoming negligible at time of hatching. In addition two distinct mRNAs for PEPCK-M are expressed during development with specific patterns that vary among individual birds. These differences are likely to be genetic, as hormonal treatment of a chicken hepatoma cell line indicates that whereas the mRNA levels for PEPCK-C are hormonally regulated, the expression of PEPCK-M mRNA is unresponsive.

Hepatic expression of the gene for phosphoenolpyruvate carboxykinase (GTP) (PEPCK-C) (EC 4.1.1.32) in birds occurs prior to birth and decreases to negligible levels before hatching, whereas in mammals the gene for PEPCK-C in the liver is expressed at birth and is active throughout the life of the animal. The administration of cyclic AMP to adult chickens results in the induction of transcription of the gene for PEPCK-C and the transient accumulation of PEPCK-C mRNA in the liver. DNase I footprint analysis of 330 bp of the avian PEPCK-C promoter immediately 5' of the start-site of transcription indicated the presence of several protein binding domains, purified CAAT/enhancer binding protein alpha, cAMP regulatory element binding protein and nuclear factor-1 bound to these regions of the promoter. Sequences corresponding to an hepatic nuclear factor-1 binding domain and to the insulin response sequence, previously identified in the rat PEPCK-C promoter, were also found in the chicken PEPCK-C promoter. Co-transfection of an expression vector for CAAT/enhancer binding protein alpha or CAAT/enhancer binding protein beta markedly stimulated transcription from both the chicken and rat PEPCK-C promoters in human hepatoma cells. Sequences involved in the regulation of gene transcription by cyclic AMP and insulin were found to reside between -210 and +1 of the avian PEPCK-C promoter. In general, transcription from the avian promoter was more sensitive to inhibition by insulin than was noted for the rat PEPCK-C promoter, which may explain in part the lack of expression of the gene for PEPCK-C in the livers of adult birds.