BACTERIAL FUNCTION INVOLVED IN CELL GROWTH CONTROL

参与细胞生长控制的细菌功能

• 批准号: 5200961
• 负责人: S GOTTESMAN
• 金额: --
• 依托单位: DIVISION OF CANCER BIOLOGY AND DIAGNOSIS
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/5200961
• 关键词: Escherichia coli; adenosinetriphosphatase; bacterial genetics; cell capsule; cell growth regulation; chimeric proteins; cold temperature; endopeptidases; enzyme mechanism; enzyme substrate; gene expression; genetic transcription; molecular chaperones; molecular cloning; open reading frames; protein biosynthesis; protein degradation; stress proteins

We have continued studies on the role that energy-dependent protein degradation plays in regulating gene expression, using E. coli as a model system. Studies on the Lon-dependent degradation of abnormal and naturally unstable proteins has demonstrated that chaperones such as DnaJ, DnaK and GrpE are not necessary for the in vivo degradation of all Lon substrates, but may help promote Lon degradation in some cases by keeping substrates in soluble form. RcsA, a positive regulator of capsule gene transcription and a Lon substrate, requires DnaJ and DnaK to remain active and accessible to the protease; the dependence on heat shock chaperones for activity may provide the basis for the temperature regulation of capsule synthesis. The synthesis of RcsA is regulated at the transcriptional level by silencing by the nucleoid associated HNS protein. We identified a small RNA, DsrA, which, when overproduced, can counteract HNS silencing of rcsA and all other HNS silenced genes. In single copy, dsrA is necessary for the very low temperature expression of capsule and synthesis from dsrB, a gene encoding a small conserved open reading frame. We have continued studies on the basis for Lon substrate selection by examining Lon degradation of GST-N fusions and hybrid proteins between lambda N and the stable Nun protein. These results suggest that regions of both the N and C terminus are sufficient for Lon recognition and degradation, and provide tools for analyzing the elements of recognition. In studies on other energy-dependent proteases in E. coli, we have found that a novel two-component protease (ClpQY) encoded by the heat shock hslU hslV operon can, when overproduced, degrade Lon substrates. It seems possible that ClpQY and the Alp protease, a protease also able to degrade Lon substrates but only thus far detected indirectly in E. coli, are one and the same. The regulation of Alp protease activity is apparently quite complex.

我们继续研究能量依赖蛋白的作用 降解在调节基因表达中发挥作用，使用大肠杆菌作为 模型系统。 异常物质的Lon依赖性降解研究 和天然不稳定的蛋白质已经证明，诸如伴侣 因为 DnaJ、DnaK 和 GrpE 对于体内降解不是必需的 所有 Lon 底物，但可能有助于促进某些 Lon 降解 情况下通过保持底物处于可溶形式。 RcsA，阳性 荚膜基因转录的调节因子和 Lon 底物，需要 DnaJ 和 DnaK 保持活性并易于蛋白酶接触；这 活动对热休克伴侣的依赖性可能提供基础 用于胶囊合成的温度调节。 的合成 RcsA 在转录水平上通过沉默来调节 核仁相关的 HNS 蛋白。 我们鉴定了一个小 RNA，DsrA， 当过量产生时，可以抵消 HNS 对 rcsA 和所有 其他 HNS 沉默基因。 在单一副本中，dsrA 对于 胶囊的极低温表达和 dsrB 的合成 编码一个小的保守开放阅读框的基因。 我们已经继续 通过检查 Lon 来选择 Lon 底物的基础上进行研究 GST-N 融合蛋白和 lambda N 之间的杂合蛋白的降解 稳定的 Nun 蛋白。 这些结果表明，两个区域 N 和 C 末端足以识别 Lon，并且 降解，并提供分析元素的工具 认出。 在对大肠杆菌中其他能量依赖性蛋白酶的研究中。 大肠杆菌中，我们发现了一种新型双组分蛋白酶（ClpQY） 由热休克 hslU hslV 操纵子编码，当过量产生时， 降解 Lon 底物。 ClpQY 和 Alp 似乎有可能 蛋白酶，一种蛋白酶也能够降解 Lon 底物，但仅此而已 到目前为止，在大肠杆菌中间接检测到的结果是相同的。 这 Alp 蛋白酶活性的调节显然相当复杂。