GENE EXPRESSION DURING DEVELOPMENT OF MYXOBACTERIA

粘细菌发育过程中的基因表达

基本信息

批准号：
2684708
负责人：
SUMIKO INOUYE
金额：
$ 28.81万
依托单位：
UNIV OF MED/DENT NJ-R W JOHNSON MED SCH
依托单位国家：
美国
项目类别：
财政年份：
1987
资助国家：
美国
起止时间：
1987-04-01 至 2000-03-31
项目状态：
已结题

项目摘要

Myxococcus xanthus is a gram negative soil bacterium which undergoes remarkable morphogenesis forming multi-cellular fruiting bodies upon depletion of nutrient. This unique developmental feature together with its social behavior provides and excellent prokaryotic system for studying intercellular as well as intracellular communication and signal transduction during differentiation. Our recent finding that M. xanthus contains a large family of eukaryotic-like protein serine/threonine kinases also raises an interesting possibility that protein serine/threonine kinase cascades unknown in the prokaryotes play a major role in environmental adaptation and morphogenesis of M. xanthus. The wealth of knowledge on M. xanthus accumulated by our group as well as others during the past 20 years allows us to readily study this bacterium as an ideal model system for our basic understanding of differentiation and cellular communication. In this proposal, we will pursue the following specific projects: (1) Roles of protein serine/threonine kinases in M. xanthus. So far we have cloned 13 kinase genes. For some genes, the determination of their DNA sequences, identification of their gene products as kinases, and genetic and biochemical analyses of the roles of the gene products have been achieved. We will continue to work on these genes as well as others to establish primary information on their DNA sequences, genetics (deletion, mutations and lacZ fusion) and biochemistry of the enzymes. At the same time, we will initiate our effort to decipher the signalling cascades by identifying cellular substrates for individual kinases and their upstream signals. In particular, Pkn1, a developmentally produced kinase, Pkn2 and Pkn11, putative transmembrane kinases, Pkn5 and Pkn6 seemingly coregulated by sharing a 130-bp upstream fragment between the two genes, and Pkn9 expressed in both vegetative and developmental growth will be investigated. (2) Identification of protein phosphatases associated with protein kinase cascades. The existence of a large family of protein kinases suggests the requirement of phosphatases in their cascades. We will explore to identify such phosphatases by biochemical and genetic means. (3) Development-specific sigma factors. We have so far identified, SigA, the putative vegetative sigma factor, and SigB and SigC that are developmentally regulated. Recently, we have found the fourth putative sigma factor, SigD and possibly the fifth factor, SigE. In particular, we will focus our effort on SigB to elucidate its regulatory cascade-and possible interaction with kinase cascades. (4) Roles of protein W in myxospores. In the course of the study on myxospore germination, we have found the protein of 41.5 kDa to be a major protein associated with the internal structure of the myxospores. Preliminary results indicate that protein W plays an essential role in the myxospore structure. We will continue to work on this rather unusual protein for its development-specific expression, its assembly in myxospores and its functional roles. (5) Development of new genetic systems required for inducible gene expression and for establishing a strain containing multiple independent deletion mutations.

粘粒球Xanthus是一种革兰氏阴性土壤细菌，发生在形成多细胞水果体的显着形态发生营养耗尽。这个独特的发展功能及其社会行为提供了学习的出色核系统细胞间以及细胞内通信和信号分化过程中的转导。我们最近的发现Xanthus 包含大型真核类蛋白丝氨酸/苏氨酸激酶还提出了一种有趣的可能性丝氨酸/苏氨酸激酶在原核生物中未知的级联黄叶菌的环境适应和形态发生中的作用。这关于Xanthus M. 在过去的20年中，其他人则可以轻松研究这种细菌作为我们对差异化基本理解的理想模型系统和蜂窝通信。在此提案中，我们将追求以下特定项目：（1）蛋白质丝氨酸/苏氨酸激酶在Xanthus中的作用。到目前为止，我们克隆了13个激酶基因。对于某些基因，确定其 DNA序列，其基因产物作为激酶的鉴定以及基因产物作用的遗传和生化分析具有已经实现了。我们将继续研究这些基因以及其他基因为了建立有关其DNA序列的主要信息，遗传学（缺失，突变和LACZ融合）和酶的生物化学。在同时，我们将开始努力破译信号传导通过鉴定单个激酶的细胞底物和他们的上游信号。特别是，PKN1是一种开发产生的激酶，PKN2和PKN11，推定的跨膜激酶，PKN5和PKN6 看似通过共享130 bp上游片段的片段来策略两个基因和PKN9在营养和发育生长中均表达将被调查。（2）鉴定与蛋白激酶相关的蛋白磷酸酶级联。大型蛋白激酶的存在表明其级联反应酶的需求。我们将探索通过生化和遗传手段鉴定这种磷酸酶。（3）特定开发的Sigma因子。到目前为止，我们已经确定了Siga，推定的营养西格玛因子，以及sigb和sigc 在开发监管上。最近，我们发现了第四个假定 Sigma因子，SIGD，可能是第五因子，Sige。特别是我们将把我们的努力集中在SIGB上，以阐明其监管级联可能与激酶级联反应。（4）蛋白质W在粘液孢子中的作用。在研究过程中粘液孢子发芽，我们发现41.5 kDa的蛋白质是主要的蛋白质与粘液孢子的内部结构相关。初步结果表明蛋白质W在粘孢子结构。我们将继续致力于这一不寻常的工作蛋白质的发育特异性表达，其组装粘液孢子及其功能作用。（5）开发诱导基因所需的新遗传系统表达并建立包含多个独立的菌株缺失突变。