Evolution of Intra-specific Regulatory Networks in a Multipartite Bacterial Genome

Reconstruction of the regulatory network is an important step in understanding how organisms control the expression of gene products and therefore phenotypes. Recent studies have pointed out the importance of regulatory network plasticity in bacterial adaptation and evolution. The evolution of such networks within and outside the species boundary is however still obscure. Sinorhizobium meliloti is an ideal species for such study, having three large replicons, many genomes available and a significant knowledge of its transcription factors (TF). Each replicon has a specific functional and evolutionary mark; which might also emerge from the analysis of their regulatory signatures. Here we have studied the plasticity of the regulatory network within and outside the S. meliloti species, looking for the presence of 41 TFs binding motifs in 51 strains and 5 related rhizobial species. We have detected a preference of several TFs for one of the three replicons, and the function of regulated genes was found to be in accordance with the overall replicon functional signature: house-keeping functions for the chromosome, metabolism for the chromid, symbiosis for the megaplasmid. This therefore suggests a replicon-specific wiring of the regulatory network in the S. meliloti species. At the same time a significant part of the predicted regulatory network is shared between the chromosome and the chromid, thus adding an additional layer by which the chromid integrates itself in the core genome. Furthermore, the regulatory network distance was found to be correlated with both promoter regions and accessory genome evolution inside the species, indicating that both pangenome compartments are involved in the regulatory network evolution. We also observed that genes which are not included in the species regulatory network are more likely to belong to the accessory genome, indicating that regulatory interactions should also be considered to predict gene conservation in bacterial pangenomes.调控网络的重构是解析生物体如何调控基因产物表达进而决定表型的关键环节。近期研究指出，调控网络可塑性在细菌适应与进化过程中具有重要意义。然而，这类网络在物种边界内外的进化机制仍不甚明晰。苜蓿中华根瘤菌（Sinorhizobium meliloti）是开展此类研究的理想模式物种：其拥有三个大型复制子，已公开的基因组数量众多，且对其转录因子（Transcription Factor, TF）的研究基础较为深厚。每个复制子均具有独特的功能与进化特征，这或许可通过分析其调控特征得以揭示。本研究聚焦苜蓿中华根瘤菌物种内外的调控网络可塑性，对51株菌株及5个相关根瘤菌物种的41个TF结合基序进行了筛查。研究发现，多个TF倾向于结合三个复制子中的某一个，且受调控基因的功能与对应复制子的功能特征相符：染色体对应持家功能，类染色体质粒（chromid）对应代谢功能，超大质粒（megaplasmid）对应共生功能。上述结果表明，苜蓿中华根瘤菌的调控网络存在复制子特异性的布线模式。与此同时，染色体与类染色体质粒间共享了相当比例的预测调控网络，这为类染色体质粒整合进入核心基因组提供了额外的调控层级。进一步分析发现，调控网络的进化距离与物种内的启动子区域及附属基因组（accessory genome）进化均呈显著相关，提示泛基因组（pangenome）的两个组分均参与了调控网络的进化过程。此外，本研究还观察到，未纳入物种调控网络的基因更大概率属于附属基因组，这表明在预测细菌泛基因组内的基因保守性时，也应将调控相互作用纳入考量范畴。