The distal enhancer implicated in the developmental regulation of the tyrosine aminotransferase gene is bound by liver-specific and ubiquitous factors.

Abstract

Tyrosine aminotransferase gene expression is confined to parenchymal cells of the liver, is inducible by glucocorticoids and glucagon, and is repressed by insulin. Three enhancers control this tissue-specific and hormone-dependent activity, one of which, located at -11 kb, is implicated in establishing an active expression domain. We have studied in detail this important regulatory element and have identified a 221-bp fragment containing critical enhancer sequences which stimulated the heterologous thymidine kinase promoter more than 100-fold in hepatoma cells. Within this region, we have characterized two essential liver-specific enhancer domains, one of which was bound by proteins of the hepatocyte nuclear factor 3 (HNF3) family. Analyses with the dedifferentiated hepatoma cell line HTC suggested that HNF3a and/or -y, but not HNF31, are involved in activating the tyrosine aminotransferase gene via the -11-kb enhancer. Genomic footprinting and in vitro protein-DNA binding studies documented cell-type-specific binding of ubiquitous factors to the second essential enhancer domain, which by itself stimulated the thymidine kinase promoter preferentially in hepatoma cells.These results will allow further characterization of the role of these enhancer sequences in developmental activation of the tyrosine aminotransferase gene.Differential gene expression is the basis of cellular diversity in complex organisms. Work from many laboratories has established that gene activity is regulated by the combinatorial action of sequence-specific DNA-binding proteins which, together with the basic transcription apparatus, control the frequency of transcription initiation at the promoter. Binding sites for transcriptional activator proteins, which mediate cell-type-specific, hormone-dependent, and developmentally regulated gene expression, can be found in the vicinity of the transcription start site and in enhancer sequences located several kilobases upstream or downstream (33, 43). More recently, cofactors or adaptors which couple activator proteins to the basal transcription factors by protein-protein interaction have been described (reference 42 and references therein).Studies on liver-specific genes have led to the identification of several liver-enriched transcriptional activators, the majority of which can be grouped into four families (see references 14 and 61 for reviews): hepatocyte nuclear factor 1 (HNF1) hierarchical order of liver-specific transcription factors (37,65).Tyrosine aminotransferase (TAT) is a gluconeogenic enzyme which has been studied as a classical system for more than 20 years (reviewed in reference 51). The TAT gene is switched on in the parenchymal cells of the liver shortly after birth, when hormone levels change dramatically (26, 28). Gene activity is induced by glucocorticoids and by glucagon, acting via its intracellular mediator cyclic AMP (cAMP) (25,29,58), and is repressed by insulin (60). Our analyses have defined three enhancers which are responsible for the liverspecific and hormone-inducibl...

The DNA-binding specificity of the hepatocyte nuclear factor 3/forkhead domain is influenced by amino-acid residues adjacent to the recognition helix.

“…3A). Although many of the endogenous HNF-3 sites appear to be specific for recognition, a subset of the HNF-3 sites may bind other family members (3,7,22,27,29,30,36,43,50,51,56). The IN-WR).…”

“…We also used HNF-3-specific antibodies with gel shift assays to confirm that none of the HFH-1 #25 DNA-protein complexes in the lung was due to HNF-3 recognition (data not shown). Further (27), albumin (ALB) (36), apolipoprotein B (APOB) (3), tyrosine aminotransferase (TAT) (43,51), phosphoenolpyruvate carboxykinase (PEPCK) (22), aldolase B (ALDB) (50), and lung clara cell secretory protein CCIO (56). Note that the HNF-3 #1 (two consecutive G residues) and HNF-3 #2 sites possess weak affinity for the HNF-3 proteins.…”

Targeted Disruption of the Gene Encoding Hepatocyte Nuclear Factor 3γ Results in Reduced Transcription of Hepatocyte-Specific Genes

“…5C, the reduced mRNA levels for TAT, PEPCK, and Tf are indeed caused by a decrease in the transcription rate of these genes. Thus, although many transcription factors have been shown to bind to the promoters and enhancers of these genes (16,20,21), HNF3␥ is an important factor in their regulation.…”

Transducin-like Enhancer of Split Proteins, the Human Homologs of Drosophila Groucho, Interact with Hepatic Nuclear Factor 3β

“…Subsequent studies showed that HNF3 is responsible for the basal expression of many liver-specific genes, such as the genes for albumin, apolipoprotein B, apolipoprotein A1, tyrosine aminotransferase, and phosphoenolpyruvate carboxykinase (PEPCK) (GTP:oxaloacetate caboxy-lyase (transphosphorylating), EC 4.1.1.32) (2)(3)(4)(5). The expression of these HNF3-regulated genes is abolished when a dominant negative HNF3 mutant is stably transfected into the mhAT3F hepatoma cell line (5), and the targeted disruption of HNF3␥ in mice results in decreased expression of these genes (6).…”