Regulation of the synthesis of tyrosine aminotransferase: the relationship to mRNATAT

The activity of the hepatic enzyme tyrosine aminotransferase (TAT) is the sum of many diverse regulatory factors. These include the developmental stage of the animal, the hormonal and nutritional environment of the animal (or tissue culture cell), other extrinsic and intrinsic regulatory cycles and factors (including cytoplasmic substances), and chromatin structure. Although TAT is subject to a number of post-translational modifications, alterations in catalytic activity always parallel changes in enzyme amount. In a few instances this is due to a selective change in TAT degradation, but most are due to changes in the rate of aminotransferase synthesis. Recent studies have shown that TAT synthesis is generally directly correlated with the activity, and presumably amount, of the mRNA that codes for tyrosine aminotransferase.     

The increase in hepatic tyrosine aminotransferase activity by ethanol administration involves an acceleration of enzyme synthesis

The present study was conducted to examine the nature of the increase in tyrosine aminotransferase (TAT) activity by acute ethanol administration. A significant rise in aminotransferase activity was observed as early as 1 hr after intact rats were gavaged with ethanol. Ethanol administration also increased TAT activity in adrenalectomized rats. Inhibition of ethanol metabolism by pyrazole administration had no effect on the ethanol-induced increase in TAT activity. Immunochemical analyses revealed that the enhancement of TAT activity in ethanol-fed rats correlated with an increase in aminotransferase protein. Measurement of the rate of TAT synthesis showed that in ethanol-fed rats, [3H]leucine was incorporated into the aminotransferase protein at a higher rate than in controls by a factor which was similar to the enhancement in enzyme activity. Our findings indicate that an acceleration of TAT synthesis fully accounts for the increase in TAT activity during the early stage of enzyme induction. TAT induction by ethanol administration is not dependent upon an increase in adrenal corticosteroid production, nor does it require ethanol metabolism.     

Comparison of the tyrosine aminotransferase cDNA and genomic DNA sequences of normal mink and mink affected with tyrosinemia type II

Type II tyrosinemia, designated Richner-Hanhart syndrome in humans, is a hereditary metabolic disorder with autosomal recessive inheritance characterized by a deficiency of tyrosine aminotransferase activity. Mutations occur in the human tyrosine aminotransferase gene, resulting in high levels of tyrosine and disease. Type II tyrosinemia occurs in mink, and our hypothesis was that it would also be associated with mutation(s) in the tyrosine aminotransferase gene. Therefore, the transcribed cDNA and the genomic tyrosine aminotransferase gene were sequenced from normal and affected mink. The gene extended over 11.9 kb and had 12 exons coding for a predicted 454-amino-acid protein with 93% homology with human tyrosine aminotransferase. FISH analysis mapped the gene to chromosome 8 using the Mandahl and Fredga (1975) nomenclature and chromosome 5 using the Christensen et al. (1996) nomenclature. The hypothesis was rejected because sequence analysis disclosed no mutations in either cDNA or introns that were associated with affected mink. This suggests that an unlinked gene regulatory mutation may be the cause of tyrosinemia in mink.     

Isolation, characterization and chromosomal mapping of the mouse tyrosine aminotransferase gene

The tyrosine aminotransferase (TAT) gene is expressed in a tissue and developmental-specific manner. In addition, this gene is regulated by glucocorticoid and polypeptide hormones and its expression is affected when a regulatory region near the albino locus of the mouse is deleted. In order to allow studies of the molecular effects of these deletion mutations we have isolated and characterized the mouse TAT gene. The gene is 9.2 x 10(3) bases in length and consists of 12 exons which give rise to a 2.3 x 10(3) base long messenger RNA. The DNA sequence at the 5' end of the gene was determined and compared with the corresponding sequence of the rat tyrosine aminotransferase gene. The sequence comparison showed extensive homology over the entire region sequenced. In addition, DNA: DNA heteroduplex studies between the mouse and rat tyrosine aminotransferase genes revealed that this homology extends over the entire gene and its flanking sequences. The mouse tyrosine aminotransferase gene has been mapped distal to the serum esterase-1 locus on mouse chromosome 8, using a restriction fragment length polymorphism between two mouse species. Since the albino deletions are located on mouse chromosome 7, the assignment of the TAT gene to chromosome 8 suggests that a regulatory factor(s) affecting TAT gene expression acts in trans.     

The effect of sodium butyrate on tyrosine aminotransferase induction in primary cultures of normal adult rat hepatocytes

Treatment of primary cultures of adult rat hepatocytes with 5 mM butyrate inhibited the spontaneous decrease in basal activity and mRNA levels of tyrosine aminotransferase (TAT) that occurred during culture (Staecker et al., submitted). We report here that butyrate treatment of primary cultures of rat hepatocytes initially inhibited the induction of TAT. This inhibition was followed by a period of accelerated TAT induction. TAT induction in butyrate-treated primary cultures of adult rat hepatocytes occurred only after metabolism of butyrate by the cultured hepatocytes. The accelerated induction of TAT in hepatocyte cultures treated with sodium butyrate was reflected by increased TAT activity and mRNA levels. Cultured hepatocytes rapidly metabolized butyrate, but the addition of more butyrate into cultures after its initial metabolism resulted in a rapid reduction in TAT activity. These findings indicate that butyrate treatment can affect the expression of TAT in primary hepatocyte cultures in both a positive (increased basal TAT expression) and a negative (inhibition of the induced expression of TAT) manner.     

Ontogeny of tyrosine aminotransferase in Xenopus laevis.

The development of tyrosine aminotransferase (TAT) activity in Xenopus laevis embryos was studied. Undivided eggs can transaminate tyrosine to some extent. The enzyme activity increases after hatching on the third day of development. In the early stages of development, the transamination of tyrosine is due to aspartate aminotransferase (ASAT, EC 2.6.1.1), both isoenzymes of which are present in the undivided egg. No specific TAT (EC 2.6.1.5) can be detected until the age of about 1 day, at which time neurulation is complete and the rapid development of the foregut and visceral pouches and arches has begun. The appearance of the enzyme is immediately preceded by a steep increase in the concentration of free tyrosine. Tyrosine aminotransferase is known to be induced by its substrate in the adult liver, and a similar effect may operate in the embryo.     

Assignment of the human tyrosine aminotransferase gene to chromosome 16

Summary The liver enzyme tyrosine aminotransferase (TAT; EC 2.6.1.5) catalyzes the rate-limiting step in the catabolic pathway of tyrosine. Deficiency in TAT enzyme activity underlies the autosomally inherited disorder tyrosinemia II (Richner-Hanhart syndrome). Using a human TAT cDNA clone as hybridization probe, we have determined the chromosomal location of the TAT structural gene by Southern blot analysis of DNAs from a series of human x rodent somatic cell hybrids. The results assign the TAT gene to human chromosome 16.     

Crystal structure of Trypanosoma cruzi tyrosine aminotransferase: substrate specificity is influenced by cofactor binding mode

The crystal structure of tyrosine aminotransferase (TAT) from the parasitic protozoan Trypanosoma cruzi, which belongs to the aminotransferase subfamily Igamma, has been determined at 2.5 A resolution with the R-value R = 15.1%. T. cruzi TAT shares less than 15% sequence identity with aminotransferases of subfamily Ialpha but shows only two larger topological differences to the aspartate aminotransferases (AspATs). First, TAT contains a loop protruding from the enzyme surface in the larger cofactor-binding domain, where the AspATs have a kinked alpha-helix. Second, in the smaller substrate-binding domain, TAT has a four-stranded antiparallel beta-sheet instead of the two-stranded beta-sheet in the AspATs. The position of the aromatic ring of the pyridoxal-5'-phosphate cofactor is very similar to the AspATs but the phosphate group, in contrast, is closer to the substrate-binding site with one of its oxygen atoms pointing toward the substrate. Differences in substrate specificities of T. cruzi TAT and subfamily Ialpha aminotransferases can be attributed by modeling of substrate complexes mainly to this different position of the cofactor-phosphate group. Absence of the arginine, which in the AspATs fixes the substrate side-chain carboxylate group by a salt bridge, contributes to the inability of T. cruzi TAT to transaminate acidic amino acids. The preference of TAT for tyrosine is probably related to the ability of Asn17 in TAT to form a hydrogen bond to the tyrosine side-chain hydroxyl group.     

Role of glucocorticoids and resident liver macrophages in induction of tyrosine aminotransferase

Administration of cortisol to an animal induces tyrosine aminotransferase (TAT) in the liver. A similar effect was observed after stimulation of resident liver macrophages (Kupffer cells) by dextran sulfate. Actinomycin D completely blocks enzyme induction both by cortisol and dextran sulfate, whereas their combined effect gives an additive result. In primary culture of hepatocytes, dextran sulfate inhibits TAT activity, but conditioned macrophage medium reliably increases enzyme activity in hepatocytes. However, incubation of isolated macrophages in the presence of dextran sulfate and such medium transfer into hepatocyte culture results in even more pronounced increase in TAT activity. In a combined culture of hepatocytes and non-parenchymal liver cells, reproducing intercellular interactions in vitro, cortisol and non-parenchymal cells exhibit an additive effect on TAT activity. These results show that liver macrophages release a factor of unknown nature launching the mechanism of TAT induction independently of cortisol, a classic TAT inducer.     

Susceptibility of mink to certain viral animal diseases foreign to the United States.

Mink (Mustela vison) were inoculated with viruses: African horse sickness (AHS), African swine fever (ASF), bovine herpes virus II (BHV2), foot-and-mouth disease (FMD), goat pox (GP), hog cholera (HC), peste des petits ruminants (PPR), rinderpest (RP), swine vesicular disease (SVD), vesicular exanthema of swine (VES) and vesicular stomatitis (VS). Their susceptibility was measured by development of clinical signs, virus isolation and detection of precipitin and/or virus neutralizing antibodies. SVD virus produced a lesion in one mink. Virus was isolated from mink inoculated with SVD, FMD and BHV2. Neutralizing and/or precipitin antibodies were detected in mink inoculated with ASF, FMD, GP, RP, SVD and VS viruses. Mink were not susceptible to AHS, HC, PPR and VES viruses.     

Induction of tyrosine aminotransferase by pyridoxine in Drosophila hydei

Salivary glands incubated in various concentrations of pyridoxine (Vitamin B₆) show increasing tyrosine aminotransferase (TAT) activity at concentrations up to 10^–5 M and then decreasing activity up to 10^–2 M but in all cases the activity is greater than that of the controls. This increase in activity is demonstrable for up to 6 h, the longest period tested, and is dependent on the synthesis of new mRNA. A similar increase in TAT activity is observed in salivary glands subjected to heat shock. Antibodies prepared against purified tyrosine aminotransferase precipitate a peptide of the same molecular weight (40 KD) as that induced by pyridoxine.     

Rosmarinic acid formation and differential expression of tyrosine aminotransferase isoforms in Anchusa officinalis cell suspension cultures

Time-course changes in rosmarinic acid (RA) formation and activities of tyrosine aminotransferase (TAT) isoforms were examined in Anchusa officinalis suspension cultures. Three TAT isoforms (TAT-1, TAT-3, TAT-4) were resolved by Mono-Q anion-exchange column chromatography. The proportion of the TAT-3 activity within the total TAT activity remained high regardless of the growth stage of the cultured cells. TAT-1 activity was positively correlated with the rate of RA biosynthesis during linear growth stage of the culture cycle, while TAT-4 activity was rapidly induced in conjunction with transfer to fresh medium coincident with a transient increase in RA synthesis. Based on these results, as well as the substrate specificity of each TAT isoform, it was concluded that both TAT-1 and TAT-4 are closely involved in RA biosynthesis. TAT-1 controls conversion of tyrosine to 4-hydroxyphenyl pyruvate, and TAT-4 acts by participating in the formation of tyrosine and phenylalanine via prephenate.Abbreviations PAL phenylalanine ammonia-lyase - TAT tyrosine aminotransferase - RA rosmarinic acid     

"Superinduction" of tyrosine aminotransferase by actinomycin D: a reevaluation.

Reexamination of the effects of actinomycin D (AMD) on the intracellular level and rate of synthesis of tyrosine aminotransferase (TAT) in hepatoma tissue culture (HTC) cells reveals that much apparent controversy can be resolved with acknowledgment of the multi-faceted nature of this inhibitor's action. AMD can slow overall protein synthesis and inhibit the degradation of both TAT and its mRNA as well as block the synthesis of RNA. The extent of these secondary actions of the inhibitor depend somewhat upon the growth condition of the cells. The effects of cordycepin (3'-deoxyadenosine) on the metabolism of TAT and its mRNA are also complex, but differ in several respects from those of AMD.     

A tyrosine aminotransferase involved in tocopherol synthesis in Arabidopsis

The metabolic function of the predicted Arabidopsis tyrosine aminotransferase (TAT) encoded by the At5g53970 gene was studied using two independent knock-out mutants. Gas chromatography-mass spectrometry based metabolic profiling revealed a specific increase in tyrosine levels, supporting the proposed function of At5g53970 as a tyrosine-specific aminotransferase not involved in tyrosine biosynthesis, but rather in utilization of tyrosine for other metabolic pathways. The TAT activity of the At5g53970-encoded protein was verified by complementation of the Escherichia coli tyrosine auxotrophic mutant DL39, and in vitro activity of recombinantly expressed and purified At5g53970 was found to be specific for tyrosine. To investigate the physiological role of At5g53970, the consequences of reduction in tyrosine utilization on metabolic pathways having tyrosine as a substrate were analysed. We found that tocopherols were substantially reduced in the mutants and we conclude that At5g53970 encodes a TAT important for the synthesis of tocopherols in Arabidopsis.     

Induction of tyrosine aminotransferase by triamcinolone during rat liver development in vitro

Embryonic rat liver 12–21 days old exhibits low but significant tyrosine aminotransferase (TAT) activity. In organ culture for 24 hr in a nutrient medium, there is an increase in TAT levels. Addition of glucocorticoids increases TAT levels at all embryonic ages. The magnitude of the produced TAT level increases with developmental age. Glucagon also increases embryonic liver TAT, but insulin and growth hormone (somatotropin) had little effect.     

Relaxed Evolution in the Tyrosine Aminotransferase Gene Tat in Old World Fruit Bats (Chiroptera: Pteropodidae)

Frugivorous and nectarivorous bats fuel their metabolism mostly by using carbohydrates and allocate the restricted amounts of ingested proteins mainly for anabolic protein syntheses rather than for catabolic energy production. Thus, it is possible that genes involved in protein (amino acid) catabolism may have undergone relaxed evolution in these fruit- and nectar-eating bats. The tyrosine aminotransferase (TAT, encoded by the Tat gene) is the rate-limiting enzyme in the tyrosine catabolic pathway. To test whether the Tat gene has undergone relaxed evolution in the fruit- and nectar-eating bats, we obtained the Tat coding region from 20 bat species including four Old World fruit bats (Pteropodidae) and two New World fruit bats (Phyllostomidae). Phylogenetic reconstructions revealed a gene tree in which all echolocating bats (including the New World fruit bats) formed a monophyletic group. The phylogenetic conflict appears to stem from accelerated TAT protein sequence evolution in the Old World fruit bats. Our molecular evolutionary analyses confirmed a change in the selection pressure acting on Tat, which was likely caused by a relaxation of the evolutionary constraints on the Tat gene in the Old World fruit bats. Hepatic TAT activity assays showed that TAT activities in species of the Old World fruit bats are significantly lower than those of insectivorous bats and omnivorous mice, which was not caused by a change in TAT protein levels in the liver. Our study provides unambiguous evidence that the Tat gene has undergone relaxed evolution in the Old World fruit bats in response to changes in their metabolism due to the evolution of their special diet.     

NADH-coupled spectrophotometric assay of tyrosine aminotransferase

A rapid spectrophotometric method for estimation of tyrosine aminotransferase activity (TAT) is described, based on a coupled reaction with NADH-dependent aromatic ketoacid reductase. 3-iodo-L-tyrosine, upon TAT action, is transformed into 3-iodo-4-hydroxyphenylpyruvate which quickly reacts with NADH in the presence of aromatic ketoacid reductase; oxidation rates at 340 nm are linear with protein concentration over the whole range of purification steps of TAT. This new method, for its sensitivity, easy performance and possibility of a continuous monitoring of TAT reaction, may be considered comparable to the more diffuse spectrophotometric standard method, and also as an alternative, advantageous procedure in some instances. The method for purification of the coupled aromatic ketoacid reductase is also described.