Developmental and hormonal regulation of the Xenopus liver‐type arginase gene

Abstract

Liver-type L-arginase is a major urea-cycle enzyme which is strongly induced during amphibian metamorphosis, but little is known about the molecular mechanisms underlying this induction. As a first step towards elucidating the possible mechanisms, we have isolated a cDNA clone for Larginase from an adult Xenopus luevis liver cDNA library. Sequence comparison of Xenopus livertype L-arginase cDNA shows a strong conservation at the amino acid level with those of human, rat and yeast. Using a Xenopus arginase cDNA fragment as a hybridization probe, we have shown by Northern blotting that the gene is highly expressed in the liver, and very slightly in kidney and spleen, of adult Xenopus. The expression is developmentally regulated. Only traces of arginase mRNA can be detected in pre-metamorphic tadpoles, but its accumulation increases very markedly at the onset of natural metamorphosis, being maintained at a high concentration constitutively upon completion of this developmental process. Amphibian metamorphosis is under the strict control of thyroid hormones. It is therefore significant that exposure of pre-metamorphic tadpoles (at stages before endogenous thyroid hormone secretion) to exogenous hormone (1 nM triiodothyronine) precociously activated the L-arginase gene. The time course of this precocious hormonal induction paralleled that of serum albumin gene in the liver. Polyclonal antibodies were raised against recombinant Xenopus L-arginase expressed in Escherichiu coli as a fusion protein with glutathione S-transferase in the plasmid expression vector pGEX. Western blotting using this antibody showed that, although arginase mRNA is present in high concentration in Xenopus tadpole liver at the onset of natural metamorphosis, the protein is detected only upon its completion. Our results show a complex transcriptional and post-transcriptional regulation of the Xenopus liver-type L-arginase gene during post-embryonic development. They also demonstrate that this gene can be exploited as a target for thyroid hormones in further studies to analyze the mechanisms underlying the establishment of the adult phenotype during amphibian metamorphosis.

The Metamorphosis of Amphibian Toxicogenomics

“…Use of the X. tropicalis genome as an assembly scaffold had limited utility since X. laevis and R. catesbeiana sequences aligned imperfectly to the X. tropicalis genome with R. catesbeiana , not surprisingly, having the least benefit of alignment. Nevertheless, some transcript identities linked to count frequencies were positively confirmed and the data obtained for the urea cycle enzymes, for example, matched well with previous observations (Helbing et al, 1992; Xu et al, 1993; Iwase et al, 1995). This validates the method for transcripts that are identifiable and quantifiable in this way.…”

Multiple Mechanisms Promote the Retained Expression of Gene Duplicates in the Tetraploid Frog Xenopus laevis

“…Of course, this pattern of substitution could also occur if the rapidly evolving paralog were a pseudogene. However in the case of liver-type arginase, polyclonal antibodies generated from protein translated from cDNA of the rapidly evolving paralog recognize two differently sized proteins in tadpole livers, but only one size in adult liver [41]. Although this would suggest expression of both paralogs at the protein level, cross hybridization to other genes or splice variants is a concern, and further studies are needed to confirm expression and translation of both of these paralogs.…”

Inhibition of apoptotic potency by ligand stimulated thyroid hormone receptors located in mitochondria

“…These investigators suggested that the truncated rTR a DF1 was likely involved in regulating mitochondrial transcriptional activity [9,21]. Thyroid hormone receptors have been shown to play a vital role in both inducing apoptosis and promoting cell proliferation during Xenopus laevis metamorphosis [22][23][24][25]. In a myoblast cell line derived from the tadpole tail (XLT-15), T 3 up-regulated the expression of the CPP32/apopain/ Yama (caspase-3 homolog) gene during tail regression and metamorphosis [26].…”