Molecular mechanism of thyroid hormone receptor function during metamorphosis

变态过程中甲状腺激素受体功能的分子机制

• 批准号: 10459124
• 负责人: Yun-Bo Shi
• 金额: $ 79.62万
• 依托单位: EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10459124
• 关键词: Adult; Amphibia; Amputation; Animals; Aorta; Apoptosis; Biological; Biological Metamorphosis; Birth; Cell Cycle; Cell Death; Cell Differentiation process; Cell Proliferation; ChIP-seq; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Connective Tissue; Defect; Development; Developmental Process; Diploidy; Dorsal; Embryo; Epithelial; Epithelial Cells; Gene Expression; Gene Transfer Techniques; Genes; Genetic Transcription; Genetic study; Goals; Histone Acetylation; Histone H4; Hormone Responsive; Hormones; Human; In Vitro; Individual; Intestines; Knock-out; Knowledge; Ligands; Limb structure; Mammals; Mediating; Modeling; Molecular; Molecular Genetics; Morphogenesis; Morphology; Muscle; NCOA3 gene; Natural regeneration; Ontology; Organ; Organ Culture Techniques; Organ Model; Pathway Analysis; Pathway interactions; Phenotype; Play; Process; RNA; Rana; Receptor Gene; Receptor Signaling; Refractory; Regenerative Medicine; Regenerative capacity; Regenerative research; Replacement Therapy; Research; Role; Signal Pathway; South African; Spinal Cord; Stem Cell Development; Steroid Receptors; Structure; System; Tadpoles; Tail; Thyroid Function Tests; Thyroid Hormone Receptor; Thyroid Hormones; Tissues; Tubular formation; Uterus; Vertebrates; Xenopus; Xenopus laevis; adult stem cell; base; chromatin immunoprecipitation; cofactor; epithelial stem cell; feeding; histone acetyltransferase; hormonal signals; in vivo; interest; intestinal maturation; knockout animal; knockout gene; model development; programs; promoter; receptor; receptor function; recruit; regenerative tissue; self-renewal; stem cell proliferation; stem cells; steroid hormone receptor; tissue regeneration; tissue/cell culture; toad; transcription activator-like effector nucleases; transcriptome sequencing

ANALYSIS OF THYROID HORMONE RECEPTOR KNOCKOUT TADPOLES REVEALS THAT THE ACTIVATION OF CELL CYCLE PROGRAM IS INVOLVED IN THYROID HORMONE-INDUCED LARVAL EPITHELIAL CELL DEATH AND ADULT INTESTINAL STEM CELL DEVELOPMENT DURING XENOPUS TROPICALIS METAMORPHOSIS. We have recently knocked out the TR genes, TR and TR, individually or both in Xenopus tropicalis and analyzed its effect on tadpole development and metamorphosis of different organs. Of interest is intestinal remodeling, which involves near complete degeneration of the larval epithelium through apoptosis. Concurrently, adult intestinal stem cells are formed de novo and subsequently give rise to the self-renewing adult epithelial system, resembling intestinal maturation around birth in mammals. We observed that both TR and TR play important roles for intestinal remodeling. To understand the underlying molecular mechanism, we recently studied the function of endogenous TR in the tadpole intestine by using knockout animals and RNA-seq analysis. We observed that removing endogenous TR caused defects in intestinal remodeling, including drastically reduced larval epithelial cell death and adult intestinal stem cell proliferation. Using RNA-seq on intestinal RNA from premetamorphic wild type and TR knockout tadpoles treated with or without TH for 1 day, prior to any detectable TH-induced cell death and stem cell formation in the tadpole intestine, we identified over 1500 genes regulated by TH treatment of the wild type but not TR knockout tadpoles. Gene ontology and biological pathway analyses revealed that surprisingly, these TR-regulated genes were highly enriched with cell cycle-related genes, in addition to genes related to stem cells and apoptosis. Our findings suggest that TR-mediated TH-activation of the cell cycle program is involved in larval epithelial cell death and adult epithelial stem cell development during intestinal remodeling. A ROLE OF ENDOGENOUS HISTONE ACETYLTRANSFERASE STEROID HORMONE RECEPTOR COACTIVATOR (SRC) 3 IN THYROID HORMONE SIGNALING DURING XENOPUS INTESTINAL METAMORPHOSIS. We have shown previously that during metamorphosis, liganded TR recruits coactivator complexes that include steroid receptor coactivator SRC3, which is a histone acetyltransferase, to TH responsive promoters. To investigate the functions of endogenous coactivators such as SRC3 during metamorphosis, we have generated Xenopus tropicalis animals lacking a functional SRC3 gene and analyzed the resulting phenotype. While removing SRC3 had no apparent effect on external development and animal gross morphology, the SRC3 (-/-) tadpoles displayed a reduction in the acetylation of histone H4 in the intestine comparing to that in wild type animals. Furthermore, the expression of TR target genes was also reduced in SRC3 (-/-) tadpoles during intestinal remodeling. Importantly, SRC3 (-/-) tadpoles had inhibited/delayed intestinal remodeling during natural and TH-induced metamorphosis, including reduced adult intestinal stem cell proliferation and apoptosis of larval epithelial cells. Our results thus demonstrate that SRC3 is a critical component of the TR-signaling pathway in vivo during intestinal remodeling. EVOLUTIONARY DIVERGENCE IN TAIL REGENERATION BETWEEN XENOPUS LAEVIS AND XENOPUS TROPICALIS. Tissue regeneration is of fast-growing importance in the development of biomedicine, particularly organ replacement therapies. Unfortunately, many human organs cannot regenerate. Anuran Xenopus laevis has been used as a model to study regeneration as many tadpole organs can regenerate. In particular, the tail, which consists of many axial and paraxial tissues, such as spinal cord, dorsal aorta and muscle, commonly present in vertebrates, can fully regenerate when amputated at late embryonic stages and most of the tadpole stages. Interestingly, between stage 45 when feeding begins to stage 47, the pseudotetraploid Xenopus laevis tail cannot regenerate after amputation. This period, termed refractory period, has been known for about 20 years. The underlying molecular and genetic bases are unclear in part due to the difficulty to carry out genetic studies in this pseudo-tetraploid species. The availability of the highly related but diploid Xenopus tropicalis offers an opportunity to study the molecular and genetic mechanisms of tail regeneration during this refractory period. We compared tail regeneration between Xenopus laevis and Xenopus tropicalis and found surprisingly that Xenopus tropicalis lacked the refractory period. Further molecular and genetic studies, more feasible in this diploid species, should reveal the basis for this evolutionary divergence in tail regeneration between two related species and facilitate the understanding how tissue regenerative capacity is controlled. In addition, it is well known that many tadpole tissues lose their regenerative capacity during metamorphosis, suggesting a role of TH and TR in regeneration. Making use of our recently generated TR knockout animals, we plan to also investigate if and how TH and TR regulate tissue regeneration. Such studies should have important implications for human regenerative medicine.

甲状腺激素受体敲除t的分析表明，细胞周期程序的激活参与甲状腺激素诱导的幼虫上皮细胞死亡和Xenopus tropicalis metamorphosis期间的成人肠道干细胞发育。最近，我们在热带爪蟾中分别或两者都淘汰了TR基因，TR和TR，并分析了其对不同器官t的影响和变态的影响。感兴趣的是肠重塑，涉及通过凋亡几乎完全变性幼虫上皮。同时，成年的肠道干细胞是从头形成的，随后引起了自我更新的成年上皮系统，类似于哺乳动物出生周围的肠道成熟。我们观察到TR和TR都起着重要的作用，可以进行肠重塑。为了了解潜在的分子机制，我们最近通过使用基因敲除动物和RNA-Seq分析研究了内源性TR在t肠中的功能。我们观察到，去除内源性TR会导致肠重塑的缺陷，包括大幅度降低的幼虫上皮细胞死亡和成人肠道干细胞增殖。使用RNA-Seq在肠道RNA上从有或没有TH的肠道RNA上使用或不带有T的TR敲除t，在t the tadpole肠中进行任何可检测到的TH诱导的细胞死亡和干细胞形成之前，我们在1500多个基因中鉴定了由TH治疗的1500多个基因，但未受到野生类型的治疗，但没有野生型敲除丁基tadpoles。基因本体论和生物途径分析表明，除了与干细胞和凋亡相关的基因外，这些TR调节基因具有高度富含细胞周期相关基因的高度富集。我们的发现表明，在肠道重塑期间，TR介导的细胞周期程序的TH介导的TH激活与幼虫上皮细胞死亡和成年上皮干细胞发育有关。 内源性组蛋白乙酰转移酶类固醇受体共激活因子（SRC）3在Xenopus肠道变质过程中甲状腺激素信号传导中的作用。我们先前已经表明，在变态过程中，配体TR募集的共激活剂复合物包括类固醇受体共激活剂SRC3（是组蛋白的乙酰转移酶）与反应型启动子。为了研究变态过程中内源性共激活因子（例如SRC3）的功能，我们已经产生了缺乏功能性SRC3基因的热带爪蟾动物，并分析了所得的表型。虽然去除SRC3对外部发育和动物总形态没有明显的影响，但与野生型动物相比，SRC3（ - / - ）t在肠中显示了组蛋白H4的乙酰化H4的乙酰化降低。此外，在肠重塑期间，在SRC3（ - / - ）t中，TR靶基因的表达也降低。重要的是，在天然和TH诱导的变形过程中，SRC3（ - / - ）t抑制了/延迟的肠道重塑，包括减少成年肠干细胞增殖和幼虫上皮细胞的凋亡。因此，我们的结果表明，在肠重塑期间，SRC3是体内TR-Signaling途径的关键组成部分。 Xenopus laevis和Xenopus tropicalis之间的尾部再生的进化差异。组织再生在生物医学（尤其是器官替代疗法）的发展中具有快速增长的重要性。不幸的是，许多人体器官无法再生。 Anuran Xenopus laevis已被用作研究再生的模型，因为许多t骨器官都可以再生。特别是，尾巴由许多轴向组织和近似组织组成，例如脊髓，背主动脉和肌肉（通常存在于脊椎动物中），当在晚期胚胎阶段和大多数tadpole阶段截肢时可以完全再生。有趣的是，在第45阶段开始喂养第47阶段时，截肢后的假倍倍倍诺植物Laevis尾巴无法再生。这一时期称为难治时期，已闻名约20年。基本的分子和遗传碱基尚不清楚，部分原因是在这种伪四倍体物种中很难进行遗传研究。在此难治时期，高度相关但二倍体爪蟾的tropicalis的可用性为研究尾巴再生的分子和遗传机制提供了机会。我们比较了Xenopus laevis和Xenopus tropicalis之间的尾巴再生，并出人意料地发现，热带爪蟾缺乏难治时期。进一步的分子和遗传研究，在这种二倍体物种中更可行，应揭示两个相关物种之间这种进化差异的基础，并促进理解如何控制组织再生能力。此外，众所周知，许多t t组织在变形过程中失去了再生能力，这表明TH和TR在再生中的作用。 利用我们最近产生的TR淘汰动物，我们计划还研究TH和TR如何调节组织再生。这样的研究应该对人类再生医学具有重要意义。