Gene-specific and General RNA Regulators in Chloroplasts

叶绿体基因的表达对光合作用至关重要，并且必须与核基因组协调。这种协调是由一套主要是转录后的调节因子实现的，其中一些具有可识别的原核起源，而另一些似乎是在内共生之后独特衍生的。一个引人入胜且尚未解决的问题是，这些来源不同的蛋白质如何相互作用，从而影响叶绿体基因表达的基因特异性和一般控制。本项目以莱茵衣藻（Chlamydomonas reinhardtii）为模型，探讨MCD1和MCD4两种调节蛋白之间的关系。MCD1编码一个大的新蛋白，它似乎是一个多聚体复合体的成员，并且是叶绿体petD mRNA积累所必需的，它通过与petD 5'非翻译区相互作用来保护其免受核糖核分解衰变。MCD4是从携带petd5 ' UTR突变的菌株的抑制因子筛选中分离出来的，该突变使转录物不稳定，但MCD4抑制因子被证明是多效性的，在其他叶绿体转录物中显示出许多缺陷。本研究的具体目的是：(1)通过体外rna结合试验探索MCD1蛋白的生化活性，以确定其是否具有可以解释其对petD特异性的特性；(2)纯化含MCD1的多蛋白复合物，并测定其成分；(3)鉴定多效性基因MCD4；(4)利用遗传杂交研究MCD4与其他RNA代谢因子的特异性和相互作用，特别是其可能调控的两种候选核糖核酸酶，多核苷酸磷酸化酶和RNase J1。该项目探索了植物细胞如何进化以协调不同隔间的活动的一个例子。发生光合作用的叶绿体在古代被捕获为细菌。在这个项目中，两个位于细胞核的基因正在研究中，因为它们调节叶绿体基因，最初是由细菌遗传的，这是光合作用所必需的。这项研究将为未来的科学家提供机会，无论是作为暑期实习生还是大学本科生，都可以获得宝贵的经验，体验生物学研究的乐趣。过去几年，在这些实习生中，代表性不足的群体一直很突出。这个项目的负责人还通过广播节目《微生物世界》、《科学歌厅》和其他工具，参与向非专业公众宣传植物科学。

Chloroplast gene expression is essential for photosynthesis, and has to be coordinated with the nuclear genome. This coordination is achieved by a suite of mostly post-transcriptional regulators, some of which have recognizable prokaryotic origins, and others of which appear to have been uniquely derived following endosymbiosis. A fascinating and largely unresolved question is how these proteins of very different origins interact to effect both gene-specific and general control of chloroplast gene expression. This project uses the green alga Chlamydomonas reinhardtii as a model to explore the relationship between two regulatory proteins, MCD1 and MCD4. MCD1 encodes a large, novel protein which appears to be a member of a multimeric complex, and is specifically required for accumulation of the chloroplast petD mRNA, which it protects from ribonucleolytic decay through interaction with the petD 5' untranslated region. MCD4 was isolated in a suppressor screen of strains carrying petD 5' UTR mutations which destabilize the transcript, however the mcd4 suppressor proved to be pleiotropic, displaying numerous defects in other chloroplast transcripts. The specific aims of this research are to: (1) explore the biochemical activities of the MCD1 protein through in vitro RNA-binding assays, to determine if it has properties which can account for its specificity for petD; (2) purify the multiprotein complex containing MCD1, and determine its components; (3) identify the pleiotropic gene MCD4; and (4) study the specificity and interactions of MCD4 with other RNA metabolic factors using genetic crosses, in particular two candidate ribonucleases which it might regulate, polynucleotide phosphorylase and RNase J1.This project explores an example of how plant cells have evolved to coordinate activities in different compartments. The chloroplast, where photosynthesis occurs, was captured in ancient times as a bacterium. In this project, two genes located in the nucleus are under study, since they regulate chloroplast genes, originally inherited with the bacterium, which are required for photosynthesis. The research will offer opportunities for future scientists, either as summer interns or university undergraduates, to gain valuable experience and taste the excitement of biological research. Underrepresented groups have been prominent among these interns in past years. The leader of this project is also involved in educating the lay public about plant science, through the radio program MicrobeWorld, a Science Cabaret, and other vehicles.