Carried out a Genome-wide identification of thyroid hormone receptor targets in the remodeling intestine during Xenopus tropicalis metamorphosis: We have been studying how TH regulates adult stem cell development by using TH-regulated intestinal metamorphosis as a model system. A key step in this is to identify and functionally characterize direct TH target genes during this process. Prior to the release of a well-annotated Xenopus genome, we generated a set of genomic microarray chips covering about 8000 bp flanking each predicted transcription start site in Xenopus tropicalis genome. We used these chips for chromatin immunoprecipitation assays (ChIP-on-chip) on the intestine of premetamorphic tadpoles treated with or without TH for genome wide identification of TR binding sites. We have now completed and published this study. Our work has led to the identification of 278 candidate direct TR target genes. We have further provided evidence that these genes are regulated by TH and likely involved in the TH-induced formation of adult intestinal stem cells during metamorphosis. Discovered that EVI and MDS/EVI are required for adult intestinal stem cell formation during postembryonic vertebrate development: As indicated above, we have been using intestinal metamorphosis as a model to study the development of adult organ-specific stem cells in vertebrates. During metamorphosis, the larval epithelial cells in the tadpole intestine undergo apoptosis in response to the rising concentration of TH. Concurrently, a small fraction of the epithelial cells, for yet unknown mechanism, undergoes TH-dependent dedifferentiation to become adult stem cells. We have previously carried out a microarray analysis to identify genes that are regulated by TH in the intestinal epithelium. Among the genes discovered are the genes ectopic viral integration site 1 (EVI) and its variant myelodysplastic syndrome 1 (MDS)/EVI. They both encode zinc-finger proteins that have been recognized as important oncogenes in various types of cancer. In contrast to the established role of EVI and MDS/EVI in cancer development, their potential function during vertebrate postembryonic development, especially in organ-specific adult stem cells are unclear. We have now shown that high levels of EVI and MDS/EVI are expressed in the intestine at the climax of metamorphosis and induced by TH. By using the TALEN gene editing technology, we have knocked out both EVI and MDS/EVI and shown that EVI and MDS/EVI are not essential for embryogenesis and premetamorphosis in Xenopus tropicalis. On the other hand, knocking out EVI and MDS/EVI causes severely retardation in the growth and development of the tadpoles during metamorphosis and leads to tadpole lethality at climax of metamorphosis. Furthermore, the homozygous knockout animals have reduced adult intestinal epithelial stem cell proliferation at the end of metamorphosis (for the few surviving ones that survive through metamorphosis) or during TH induced metamorphosis. These findings reveal a novel role of EVI and/or MDS/EVI in regulating the formation and/or proliferation of adult intestinal adult stem cells during postembryonic development in vertebrates.
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