REGULATION OF CYSTATHIONASE EXPRESSION

本研究的长期目标是了解 哺乳动物细胞中的转硫作用。 转硫途径导致 在将蛋氨酸的硫原子转移到半胱氨酸中。 早期 该途径的酶与转甲基化途径共享， 甲基有助于许多重要的代谢 中间体的 几乎没有分子机制的知识 这是两种途径中酶表达的基础。 我们的具体目标是阐明分子机制， 调节胱硫醚酶（CSE）的表达， 转硫作用 我们将研究CSE在发育中的大鼠肝脏中的调节， 其显示了与人肝脏相似的CSE表达的时间模式。 CSE蛋白质合成和降解速率将在不同温度下测定。 发展阶段 功能性CSE mRNA的浓度将是 确定每个阶段和大鼠脑组织，其中CSE活性是 很低 为了更详细地了解CSE的遗传调控， 我们将分离代表CSE mRNA的cDNA克隆，使用我们的特异性引物， 用于文库筛选和/或多核糖体免疫选择的抗血清。 使用 克隆的cDNA，我们将分析转录和转录后 CSE基因表达的调控以及决定基因的数量 CSE基因的拷贝数、内含子的存在和甲基化状态。 有许多临床和基本问题， 转硫酶 出生时缺乏CSE可能使半胱氨酸成为一种 新生儿必需氨基酸。 酶活性遗传缺失 导致胱硫醚尿症， 迟钝 通过转硫作用在肝脏中产生半胱氨酸， 对维持谷胱甘肽水平至关重要。 该硫醇是主要的 重要的是保护细胞免受辐射和有毒药物， 对乙酰氨基酚和化疗剂。 所以，理解 对转硫作用的调控可能对以下方面产生深远的影响： 人体健康 （B）

The long-term objective of this research is to understand the regulation of transsulfuration in mammalian cells. The transsulfuration pathway results in the transfer of the sulfur atom of methionine into cysteine. The early enzymes of the path are shared with the transmethylation pathway by which methyl groups are contributed to a number of important metabolic intermediates. There is almost no knowledge of the molecular mechanisms which underlie expression of enzymes in either path. Our specific objectives are to elucidate the molecular mechanisms which regulate expression of cystathionase (CSE), the terminal enzyme of transsulfuration. We shall examine CSE regulation in developing rat liver, which shows a similar temporal pattern of CSE expression to human liver. CSE protein synthesis and degradation rates will be determined at different stages of development. The concentration of functional CSE mRNA will be ascertained for each stage and for rat brain tissue, where CSE activity is very low. To obtain more detailed understanding of CSE genetic regulation, we shall isolate cDNA clones representing CSE mRNA, employing our specific antisera for library screening and/or polysome immunoselection. Using the cloned cDNA, we shall analyze transcriptional and post-transcriptional regulation of CSE gene expression as well as determine the number of gene copies, presence of introns, and methylation states of the CSE gene(s). There are numerous clinical and basic problems that relate to transsulfuration enzymes. Absence of CSE at birth may make cysteine an essential amino acid for the neonate. Genetic absence of enzyme activity results in cystathioninuria which has been associated with mental retardation. The hepatic production of cyteine by transsulfuration is crucial in maintaining glutathione levels. This thiol is of major importance in protecting cells from radiation and toxic drugs including acetaminophen and chemotherapeutic agents. Thus, an understanding of the regulation of transsulfuration could have far reaching implications for human health. (B)