A DNA Conformation-Dependent Genetic Switch in Halobacterium

9604443 DasSarma这项研究验证了一种假设，即盐生盐杆菌基因组的特定区域采用超螺旋密度依赖的Z-DNA构象，作为基因表达控制的调节开关。盐生盐杆菌是一种极端嗜盐的古生菌，其左旋Z-DNA构象具有高度负的DNA超螺旋密度、高G+C含量(包括交替的CG序列)和高的细胞质盐浓度(5M)。突变和生化分析相结合将确定编码盐藻紫膜蛋白的bop基因的表达是否受位于bop基因转录起始点上游23个碱基对的潜在Z-DNA序列的B-to-Z DNA构象依赖开关控制。初步证据表明，BOP基因转录在DNA高度负超卷曲条件下被诱导，当DNA旋转酶的抑制解除超卷曲时，BOP基因转录被抑制。因此，DNA旋转酶抑制剂novobiocin将被用来确定DNA超螺旋在介导DNA构象变化和BOP转录中的作用，包括建立一个体外转录系统来研究DNA超螺旋对转录的影响。本研究将评估DNA构象和影响DNA构象的条件在盐生盐杆菌基因表达调控中的作用。盐生盐杆菌是一种存在于极高盐浓度条件下的古细菌。***

9604443 DasSarma This research tests the hypothesis that specific regions of the genome of Halobacterium halobium adopt supercoil density-dependent Z-DNA conformations that function as regulatory switches for the control of gene expression. Halobacterium halobium is an extremely halophilic Archaea in which the left-handed Z-DNA conformation is favored by a highly negative DNA supercoiling density, a high G+C content including alternating CG sequences, and a high cytoplasmic salt concentration (5M). A combination of mutagenic and biochemical analysis will determine if expression of the bop gene encoding the H. halobium purple membrane protein is controlled by B-to-Z DNA conformation-dependent switch at a potential Z-DNA sequence located 23 base pairs upstream from the bop gene transcription start site. Preliminary evidence indicates that bop gene transcription is induced under conditions of highly negative DNA supercoiling, and reduced when supercoiling is relieved by inhibition of DNA gyrase. Therefore, the DNA gyrase inhibitor novobiocin will be used to determine the role of DNA supercoiling in mediating DNA conformational changes and bop transcription, including the development of an in vitro transcription system for studying the effects of DNA supercoiling on transcription. %%% This research will evaluate the role of DNA conformation and conditions that affect DNA conformation in the regulation of gene expression in Halobacterium halobium, a Archaea bacteria that exists under conditions of extremely high salt concentration. ***