Stringent response and bmp expression in B. burgdorferi

伯氏疏螺旋体的严格反应和 bmp 表达

• 批准号: 6617152
• 负责人: Felipe Cardenas Cabello
• 金额: $ 16.1万
• 依托单位: NEW YORK MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-07-01 至 2008-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6617152
• 关键词: Borrelia; bacterial antigens; bacterial genetics; gene expression; genetic transcription; laboratory mouse; laboratory rabbit; laboratory rat; microorganism culture; physiologic stressor; ticks; tissue /cell culture; virulence

DESCRIPTION (provided by applicant): In previous studies we have shown that the B. burgdorferi bmp chromosomal gene cluster encoding the paralogous Bmp membrane lipoproteins is present in all B. burgdorferi sensu lato. This cluster forms two complex transcriptional units that are transcribed into variable monocistronic and polycistronic messages to generate different bmp mRNA and gene product concentrations. Nutritional stress in Borrelia burgdorferi triggers the stringent response that modulates expression of many genes and is mediated by the alarmon (p)ppGpp. Our work now indicates that B. burgdorferi has a transcriptionally active rel/Bbu, gene needed for generation of (p)ppGpp and that the presence of (p)ppGpp is associated with the modulation of expression of bmp and other genes under different environmental conditions. The genetic, transcriptional, and biological characteristics of the bmp gene cluster makes it an ideal model to study chromosomal gene regulation both by trans regulators such as (p)ppGpp and sigma factors and by cis DNA sequences. The hypothesis underlying this project is that expression of bmp and other B burgdorferi genes is modulated in trans by the stringent response both directly by (p)ppGpp and indirectly through sigma 54 and sigma S and in cis by the DNA sequences of their 5' upstream promoter regions. The specific aims of this project are: 1) characterize the regulation of rel/Bbu expression and the Rel/Bbu-mediated stringent response on bmp expression during growth of B. burgdorferi in BSKH media, in co-culture with tick cells, and in rat peritoneal culture chambers and ticks; 2) ascertain the roles of sigma S and sigma 54 in modulating bmp expression by the stringent response and their hierarchical interactions in B. burgdorferi in vitro and in vivo using bacterial genetics methods, DNA microarrays and proteomics; and 3) identify putative regulatory proteins binding to 5' upstream DNA sequences of the rpsL, bmpD, bmpC, bmpA and sigma S genes and their DNA binding sites. The proposed experiments will permit us to identify global factors, hierarchical regulatory networks and cis acting DNA sequences involved in the regulation of bmp genes, and will take studies of gene expression and modulation in B. burgdorferi from a descriptive paradigm to a paradigm framed by mechanistic and causal interpretations.

描述（由申请人提供）：在之前的研究中，我们已经证明编码旁系bmp膜脂蛋白的伯氏疏螺旋体bmp染色体基因簇存在于所有的伯氏疏螺旋体中。该簇形成两个复杂的转录单位，转录成可变的单顺反子和多顺反子信息，以产生不同的bmp mRNA和基因产物浓度。伯氏疏螺旋体的营养应激触发了严格的反应，该反应调节了许多基因的表达，并由警报(p)ppGpp介导。我们现在的工作表明，伯氏疏螺旋体具有转录活性的rel/Bbu，这是生成(p)ppGpp所需的基因，并且(p)ppGpp的存在与bmp和其他基因在不同环境条件下的表达调节有关。bmp基因簇的遗传、转录和生物学特性使其成为研究染色体基因调控的理想模型，无论是通过反式调控如(p)ppGpp和sigma因子，还是通过顺式DNA序列。该项目的假设是bmp和其他伯氏疏螺旋体基因的表达在反式中受到(p)ppGpp直接和间接通过sigma 54和sigma S的严格响应的调节，在顺式中受到其5'上游启动子区的DNA序列的调节。本项目的具体目的是：1)表征伯氏疏螺旋体在BSKH培养基、与蜱细胞共培养、大鼠腹膜培养室和蜱中生长过程中rel/Bbu表达的调控以及rel/Bbu介导的对bmp表达的严格响应；2)利用细菌遗传学、DNA微阵列和蛋白质组学等方法，研究sigma S和sigma 54在伯氏疏螺旋体体外和体内的严格反应及其层次相互作用对bmp表达的调节作用；3)鉴定rpsL、bmpD、bmpC、bmpA和sigma S基因5'上游DNA序列的推定调控蛋白及其DNA结合位点。我们提出的实验将使我们能够确定bmp基因调控的全局因素、分层调控网络和顺式作用DNA序列，并将对伯氏疏螺旋体基因表达和调控的研究从描述性范式转变为由机制和因果解释框架的范式。