THYROID HORMONE CONTROL OF A GENE EXPRESSION NETWORK

两栖动物的变态为研究激素如何以组织特异性和发育阶段特异性的方式控制基因表达网络提供了一个极好的机会。在从幼蛙蝌蚪到成蛙的过渡过程中，甲状腺激素控制着新成蛙结构的生长和分化，成蛙现有结构的重塑，以及整个幼蛙器官的甲状腺激素分解。这些形态发生的变化是由核甲状腺激素受体介导的基因表达变化引起的。消减杂交和候选基因方法已经在重吸收的蝌蚪尾巴中发现了20到30个甲状腺激素诱导和抑制的基因，一个激素控制的程序性细胞死亡的例子研究了下游的AM信号事件，最终导致组织特异性的生物反应。此外，目前的方法，如天然铬素。我们建议确定在这种基因表达级联中最后被诱导的基因的甲状腺激素激活的机制。我们选择研究的基因编码一种分泌型基质金属蛋白酶，胶原酶-3，以及一种细胞内死亡诱导蛋白酶，caspase-3。胶原酶-3是濒临死亡的肌肉细胞中上调最强的基因。我们将使用两种互补的方法来识别和表征这两个序列中的Th反应和基本调控元件。对候选调控序列的理想测试是它是否能够支持体内适当的转录调控。因此，我们将最近开发的非洲爪哇转基因技术来筛选最小的基因组片段从胶原酶-3和caspase-3基因中筛选与早期诱导的转录结合的序列，重要的调节序列将在体内使用转基因蝌蚪进行功能表征，从甲状腺高水平与其受体更多地结合到诱导一个概念的过程中，利用增益和激素诱导的基因表达级联反应，在拟议的工作中确定的分子标记可以应用于其他重要但复杂的生物学问题。这些包括甲状腺激素对两栖动物肢体生长和分化的控制，当然还有类固醇和甲状腺激素对成年和发育中的哺乳动物基因网络的控制。

Amphibian metamorphosis presents an excellent opportunity to study how hormones control gene expression networks in a tissue-specific and development stage-specific manner. During the transition from the larval tadpole to the adult frog, thyroid hormone controls the growth and differentiation of new adult structures, the remodeling of existing structures for a an adult frog, thyroid hormone lysis of entire larval organs. These morphogenetic changes are caused by gene expression changes mediated by the nuclear thyroid hormone receptors. Subtractive hybridization and candidate gene approaches have identified between twenty and thirty thyroid hormone induced and repressed genes in the reabsorbing tadpole tail, an example of hormonally controlled programmed cell death examined the downstream am signaling events that culminate in tissue specific biological responses. Furthermore, current methods such native chrromatin. We propose to determine the mechanism of thyroid hormone activation of genes that are among the last to be induced in this gene expression cascade. The genes we have chosen to study encode a secreted matrix metalloproteinase, collagenase-3 , and an intra cellular cell death inducing protease, caspase-3. Collagenase- 3 is the most strongly up-regulated gene in dying muscle cells. We will use two complementary approaches to identify and characterize the Th- responsive and basal regulatory elements in these two sequenced. The ideal test of a candidate regulatory sequence is whether or not it can support appropriate transcriptional regulation in vivo. Thus, we will recently developed transenic technologies in Xenopus laevis to screen for the minimal genomic fragment genomic DNA from the collagenase-3 and caspase-3 genes will be screened for sequences that bind to early induced transcription bind to important regulatory sequences will be functionally characterized in vivo using transgenic tadpoles, employing both gain- and hormone-induced gene expression cascade from the time of thyroid hor more binding to its receptors to the induction of a concepts, and molecular markers identified during the proposed work can be applied to other important yet complex biological problems. These include thyroid hormone control of the growth and differentiation of the amphibian limbs, and certainly steroid and thyroid hormone control of gene networks in adult and developing mammals.