联苯培养条件下红球菌R04转录表达和苯甲酸代谢途径解析

Transcriptomic and benzoate metabolic pathways of Rhodococcus sp. R04 cultured in biphenyl

摘要:

摘要:【目的】研究不同碳源，特别是联苯条件下红球菌的细胞转录应答，以挖掘与多氯联苯(PCBs)转运、代谢及其调控相关的基因，为进一步全面理解PCB微生物降解的分子机制奠定基础。【方法】以一株多氯联苯降解菌红球菌(Rhodococcus sp.R04)为材料，分别提取不同碳源(乙醇、葡萄糖和联苯)培养条件下菌体的总RNA，反转录合成cDNA。采用高通量测序法分别对这三种样品进行转录组测序，分析测序数据得出全基因组表达模式，并对不同条件下的基因表达进行差示分析，进而对联苯代谢网络和红球菌中其他基因的转录调节和代谢应答反应做出相关性分析。Q-RT-PCR分析不同碳源培养条件下的基因表达情况。【结果】测序结果表明，与葡萄糖和乙醇相比，联苯培养条件下明显上调(log2 Ratio 1)基因个数分别为375和332个。与葡萄糖相比，联苯培养条件下，相关基因上调表达量与Q-RT-PCR实验结果基本一致。功能分类获得细胞组分、分子功能和生物学过程三大类别160多个细小分支的差示表达基因，部分基因参与联苯代谢转录调控、联苯转运、抗氧化应激反应以及信号传导通路系统等多种生理过程。参与联苯上游代谢途径的众多同工酶基因中，只有bphC2和bphD1在联苯中大量上调表达，其余同工酶在联苯中基本量不变或下调表达。转录组注释及差示分析推测，红球菌R04中苯甲酸的代谢主要是通过儿茶酚邻位途径、间位途径以及原儿茶酸途径三条代谢途径完成。【结论】与葡萄糖和乙醇相比，红球菌R04在联苯培养条件下基因表达差异明显，这为我们进一步解析多氯联苯代谢特征和代谢调控提供理论依据。

关键词:关键词:联苯代谢，苯甲酸代谢，转录应答，相关性分析

Abstract:

Abstract:［Objective］Studying transcriptional characteristics of Rhodococcus sp.R04 is to find the genes that participate in the transportation，metabolism and regulation of polychlorinated biphenyls (PCBs)，and disclose the molecular mechanism of biodegradation of PCBs． ［Methods］ Strain R04 was separately cultivated on ethanol，glucose and biphenyl. The total RNA of the above different cultures was extracted，and the cDNA was obtained by reverse transcription and determined by high-throughput sequencing. The data obtained by sequencing were analyzed to find the correlation among the PCBs metabolic network，gene transcription regulation and metabolic response.［Results］ The sequencing results showed that 375 genes were up-regulated during grown on biphenyl，relative to growth on glucose，and 332 genes were up-regulated，relative to growth on ethanol． Those genes were found to participate in multiple biological processes of biphenyl metabolite． Among the genes relative to biphenyl/PCBs degradation，the genes located on the gene cluster in the upper biphenyl pathway were significantly up-regulated，while bphC4 and bphD2 were up-regulated slightly．By contrast，the other genes encoding BphC and BphD isozymes in the biphenyl pathway were down-regulated even if growth on biphenyl.［Conclusion］Transcriptomic analysis suggested that benzoate was degraded via ortho cleavage pathway，meta cleavage pathway or protocatechuic acid pathway，which provide more valuable data for us to reveal the characteristics and regulation of downstream metabolic pathways of biphenyl and PCBs.

Key words:Keywords: biphenyl metabolic，benzoate metabolic，transcription response，correlation analysis