XB-ART-48511Cell Biosci November 13, 2013; 3 (1): 43.
Differential regulation of two histidine ammonia-lyase genes during Xenopus development implicates distinct functions during thyroid hormone-induced formation of adult stem cells.
BACKGROUND: Organ-specific, adult stem cells are essential for organ-homeostasis and tissue repair and regeneration. The formation of such stem cells during vertebrate development remains to be investigated. Frog metamorphosis offers an excellent opportunity to study the formation of adult stem cells as this process involves essentially the transformations of all larval tissues/organs into the adult form. Of particular interest is the remodeling of the intestine. Early studies in Xenopus laevis have shown that this process involves complete degeneration of the larval epithelium and de novo formation of adult stem cells through dedifferentiation of some larval epithelial cells. A major advantage of this metamorphosis model is its total dependence on thyroid hormone (T3). In an effort to identify genes that are important for stem cell development, we have previously carried out tissue-specific microarray analysis of intestinal gene expression during Xenopus laevis metamorphosis. RESULTS: We report the detailed characterization of one of the genes thus identified, the histidine ammonia-lyase (HAL) gene, which encodes an enzyme known as histidase or histidinase. We show that there are two duplicated HAL genes, HAL1 and HAL2, in both Xenopus laevis and Xenopus tropicalis, a highly related but diploid species. Interestingly, only HAL2 is highly upregulated by T3 and appears to be specifically expressed in the adult intestinal progenitor/stem cells while HAL1 is not expressed in the intestine during metamorphosis. Furthermore, when analyzed in whole animals, HAL1 appears to be expressed only during embryogenesis but not metamorphosis while the opposite appears to be true for HAL2. CONCLUSIONS: Our results suggest that the duplicated HAL genes have distinct functions with HAL2 likely involved in the formation and/or proliferation of the adult stem cells during metamorphosis.
PubMed ID: 24499573
PMC ID: PMC3874607
Article link: Cell Biosci
Species referenced: Xenopus laevis
Genes referenced: hal.1 hal.2 mmp20 mmp9.1
Article Images: [+] show captions
|Figure 1. The two Xenopus histidine ammonia lyase (HAL) genes encode proteins highly homologous to mammalian HAL. Sequence alignment was carried out among the human (h), mouse (m) and the three HALs from Xenopus laevis (xl) and two HALs Xenopus tropicalis (xt), a diploid species. Note that the proteins are highly homologous throughout the entire length. The HAL1 and HAL2 from Xenopus laevis and Xenopus tropicalis shared about 84% identity between each other, similar to their homologies to the mammalian HAL.|
|Figure 2. Phylogenetic tree of HALs from different species as shown in Figure1suggests that the two HAL genes were duplicated after the separation of mammals and amphibians but prior to the separation of Xenopus laevis and Xenopus tropicalis or that mammals lost the HAL1 gene during evolution.|
|Figure 3. Upregulation of HAL2 gene only in the epithelium during Xenopus laevis intestinal metamorphosis. The epithelium (Ep) and the rest of the intestine (non-Ep), which is made of mainly the connective tissue, were isolated from tadpoles at stage 56 (the onset of metamorphosis, also referred to as a premetamorphic stage), stage 61 (the climax of metamorphosis), and stage 66 (the end of metamorphosis) . The total RNA from the Ep and non-Ep was used for qRT-PCR analysis of HAL2 gene expression. Note that HAL2 was exclusively expressed in the Ep at the climax of metamorphosis.|
|Figure 4. Upregulation of HAL2 during intestinal metamorphosis. Total RNA isolated from the intestine of animals from premetamorphic stage 54 to the end of metamorphosis (stage 66) was subjected to qRT-PCR analysis for HAL2 gene expression. Note that again HAL2 was found to have little or no expression before or after metamorphosis but was drastically upregulated during metamorphosis.|
|Figure 5. Dramatic upregulation of HAL2 by T3-treatment of premetamorphic tadpoles. Stage 54 premetamorphic tadpoles were treated with 10 nM T3 for 0-7 days and total intestinal RNA was isolated for expression analysis. Note that HAL2 expression was upregulated significantly after 1 day of treatment and the expression continued to increase dramatically during the treatment, peaking after 5 days.|
|Figure 6. In situ hybridization analysis suggests specific expression of HAL2 in the proliferating adult intestinal progenitor/stem cells. ISH on intestinal cross-sections at stages 54 (A), 61 (B), and 66 (C) was carried out with anti-sense probe for HAL2. An enlarged photo of the boxed areas (a, b, c) in the top panels are shown in the lower panels. Note that HAL2 staining was found to be in clusters of cells located in between the dying larval epithelial cells and connective tissue at the climax of metamorphosis, where the proliferating adult epithelial cells are located [8,19,36,48], consistent with the expression data in Figures 3 and 4. AE, adult epithelium; CT, connective tissue; Lu, lumen; LE, larval epithelium; M, muscle; Ty, typhlosole.|
|Figure 7. Distinct expression profiles of HAL1 and HAL2 during development. Total RNA isolated from whole animals from embryonic stage 30 to the end of metamorphosis (stage 66) was subjected to quantitative RT-PCR analysis for HAL1 and HAL2 gene expression. Note that HAL1 was expressed strongly in embryos and repressed after stage 45 when tadpole feeding begins while HAL2 was upregulated by stage 45 and its expression continued to rise till the end of metamorphosis.|
References [+] :
AUERBACH, Histidinemia. A deficiency in histidase resulting in the urinary excretion of histidine and of imidazolepyruvic acid. 1962, Pubmed