REGULATION OF SPORULATION-ASSOCIATED GENE TRANSCRIPTION

基因转录对环境反应的诱导 信号是遗传学的一个核心但仍知之甚少的方面 监管。芽胞形成细菌枯草芽孢杆菌将 在以下情况下激活一系列与产孢子相关的基因 营养不良，并提供了一个极好的系统 研究转录调控机制，这些机制是 对营养环境做出反应。我现在是 研究三个产孢子相关基因的转录； 早期诱导的产孢子基因。SpoOH、SpoVG和一个基因 参与多肽抗生素TycA的合成。全部显示 处于一种常见形式的依赖的负面控制之下 在一个被称为abrB的遗传基因座上。细胞内GTP水平也 似乎在转录调控中发挥作用 产孢子基因。这项提案中概述的主要目标 目标是： 1)通过以下方式定义顺式作用转录调控位点 SpoVG和SpoOH基因启动子的突变分析。我们 也会检查其他基因的表达情况 预计会受到abrB(即编码 中性蛋白酶NPR)。 2)确定与GTP-和abrB-相关的调节器- 依赖的负面控制，最终目标是建立 一种连接促进产孢量的代谢事件的途径 随着基因转录的激活。我们将从 分离导致spoVG、tycA、 和SpoOH是构成的和/或独立于负极的 控制力。 3)利用转座子TN分离abrB基因(S) 917插入，我们已经在abrB基因座上发现。克隆的DNA 转座子的两侧将被用来进行二倍体分析 以确定abrB基因座是否由一个以上的 吉恩。我们还将利用这一DNA获得完整的abrB基因(S) 然后将对其进行排序，以确定主要的 其产品结构(S)。克隆的DNA将被用来创建 AbrB缺失突变体。我们还将使用abrB DNA来 监测abrB的表达并确定其基因 对其表达的要求。ABB产品(S)将是 在B枯草杆菌细胞中过量生产，以检验高表达的效果 AbrB水平对SpoVG、SpoOH和tycA转录的影响 对产孢量的影响。AbrB DNA将被诱变 并重新引入枯草杆菌，希望获得和 干扰孢子形成的abrB突变的特征 入会仪式。

The induction of gene transcription in response to environmental signals is a central but still poorly understand aspect of genetic regulation. The spore-forming bacterium Bacillus subtilis will activate a diverse collection of sporulation-associated genes when nutritionally deprived and provides an excellent system with which to study transcriptional regulatory mechanisms which are responsive to the nutritional environment. I am currently studying the transcription of three sporulation-associated genes; the early-induced sporulation genes. spoOH and spoVG and a gene involved in the synthesis of a peptide antibiotic, tycA. All appear to be under a common form of negative control that is dependent on a genetic locus known as abrB. Intracellular levels of GTP also appear to play a role in the transcriptional regulation of sporulation genes. The main objectives outlined in this proposal are to: 1) define cis-acting transcriptional regulatory sites through mutational analysis of the spoVG and spoOH gene promoters. We will also examine the expression of other genes which we would expect to be influenced by abrB (i.e., the gene encoding the neutral protease npr). 2) identify regulators that are associated with GTP- and abrB- dependent negative control with the ultimate goal of establishing a pathway linking the metabolic events which promote sporulation with the activation of gene transcription. We will begin by isolating mutations which cause the expression of spoVG, tycA, and spoOH to be constitutive and/or independent of negative control. 3) isolate the abrB gene(s) by taking advantage of a transposon Tn 917 insertion we have discovered in the abrB locus. Cloned DNA flanking the transposon will be used to perform diploid analysis in order to determine if the abrB locus is composed of more than one gene. We will also use this DNA to obtain the intact abrB gene(s) which will then be sequenced in order to determine the primary structure of its product(s). The cloned DNA will be used to create a abrB deletion mutant. We will also use the abrB DNA to monitor the expression of abrB and to determine the genetic requirements for its expression. The abrB product(s) will be overproduced in B subtilis cells to examine the effect of high AbrB levels on the transcription of spoVG spoOH and tycA as well as the effect on sporulation. The abrB DNA will be mutagenized and reintroduced into B. subtilis in hopes of obtaining and characterizing abrB mutations which interfere with sporulation initiation.