Table_2_Transcriptional Response of Streptomyces coelicolor to Rapid Chromosome Relaxation or Long-Term Supercoiling Imbalance.xlsx

Negative DNA supercoiling allows chromosome condensation and facilitates DNA unwinding, which is required for the occurrence of DNA transaction processes, i.e., DNA replication, transcription and recombination. In bacteria, changes in chromosome supercoiling impact global gene expression; however, the limited studies on the global transcriptional response have focused mostly on pathogenic species and have reported various fractions of affected genes. Furthermore, the transcriptional response to long-term supercoiling imbalance is still poorly understood. Here, we address the transcriptional response to both novobiocin-induced rapid chromosome relaxation or long-term topological imbalance, both increased and decreased supercoiling, in environmental antibiotic-producing bacteria belonging to the Streptomyces genus. During the Streptomyces complex developmental cycle, multiple copies of GC-rich linear chromosomes present in hyphal cells undergo profound topological changes, from being loosely condensed in vegetative hyphae, to being highly compacted in spores. Moreover, changes in chromosomal supercoiling have been suggested to be associated with the control of antibiotic production and environmental stress response. Remarkably, in S. coelicolor, a model Streptomyces species, topoisomerase I (TopA) is solely responsible for the removal of negative DNA supercoils. Using a S. coelicolor strain in which topA transcription is under the control of an inducible promoter, we identified genes involved in the transcriptional response to long-term supercoiling imbalance. The affected genes are preferentially organized in several clusters, and a supercoiling-hypersensitive cluster (SHC) was found to be located in the core of the S. coelicolor chromosome. The transcripts affected by long-term topological imbalance encompassed genes encoding nucleoid-associated proteins, DNA repair proteins and transcriptional regulators, including multiple developmental regulators. Moreover, using a gyrase inhibitor, we identified those genes that were directly affected by novobiocin, and found this was correlated with increased AT content in their promoter regions. In contrast to the genes affected by long-term supercoiling changes, among the novobiocin-sensitive genes, a significant fraction encoded for proteins associated with membrane transport or secondary metabolite synthesis. Collectively, our results show that long-term supercoiling imbalance globally regulates gene transcription and has the potential to impact development, secondary metabolism and DNA repair, amongst others.

负DNA超螺旋（Negative DNA supercoiling）可介导染色体凝缩，并促进DNA解旋，而后者是DNA各类代谢过程——即DNA复制、转录与重组——发生的必要条件。在细菌中，染色体超螺旋状态的改变会影响全局基因表达；然而，目前针对全局转录响应的相关研究较为有限，且多聚焦于病原菌物种，同时已报道的受影响基因占比也各不相同。此外，人们对长期超螺旋失衡的转录响应仍知之甚少。本研究聚焦链霉菌属（Streptomyces）环境产抗生素细菌，探究其对新生霉素（novobiocin）诱导的快速染色体解旋，以及长期拓扑失衡（包括超螺旋升高与降低）的转录响应。链霉菌的复杂发育周期中，菌丝细胞内存在多拷贝的GC富集型线性染色体，其拓扑结构会发生剧烈变化：从营养菌丝中松散凝缩的状态，转变为孢子中高度压缩的状态。此外，有研究表明染色体超螺旋的改变与抗生素合成调控及环境应激响应相关。值得注意的是，作为链霉菌的模式物种，天蓝色链霉菌（S. coelicolor）的拓扑异构酶I（TopA，topoisomerase I）是唯一负责去除负DNA超螺旋的酶。本研究利用一个topA转录受诱导型启动子调控的天蓝色链霉菌菌株，鉴定出参与长期超螺旋失衡转录响应的基因。受影响的基因优先以多个基因簇的形式组织排布，其中一个超螺旋敏感簇（supercoiling-hypersensitive cluster, SHC）被定位于天蓝色链霉菌染色体的核心区域。受长期拓扑失衡影响的转录本涵盖了编码类核结合蛋白（nucleoid-associated proteins）、DNA修复蛋白以及转录调控因子（包括多种发育调控因子）的基因。此外，通过使用旋转酶抑制剂（gyrase inhibitor），本研究鉴定出了受新生霉素直接影响的基因，并发现这些基因的启动子区域AT含量升高与其敏感性存在相关性。与受长期超螺旋改变影响的基因不同，在新生霉素敏感基因中，有相当一部分编码与膜转运或次级代谢产物合成相关的蛋白质。综上，本研究结果表明，长期超螺旋失衡可全局性调控基因转录，并有可能影响发育、次级代谢以及DNA修复等诸多生命过程。