Transcriptional profiling of nitrogen fixation and the role of NifA in the diazotrophic endophyte Azoarcus sp. strain BH72.
Transcriptional profiling of nitrogen fixation and the role of NifA in the diazotrophic endophyte Azoarcus sp. strain BH72.
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DOI：
10.1371/journal.pone.0086527
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Reinhold-Hurek B
中科院分区：
文献类型：
--
作者：
Sarkar A;Reinhold-Hurek B
模式内生真菌Azoarcus sp.菌株BH72为其寄主Kallar grass提供固定氮，并在水稻幼苗内生表达氮酶基因。氮的可利用性是调控氮酶基因转录的信号。因此，我们分析了响应氮源差异的全局转录。DNA微阵列，包括70个寡核苷酸代表菌株BH72基因组的3989个开放阅读框，用于转录组研究。比较了微生物培养细胞在N2和铵环境下的转录谱。固定N2后，7.2%的基因表达显著上调，5.8%的基因表达显著下调。对38个上游区域的σ54型启动子序列进行了平行全基因组预测，其中36个序列在N2响应下被调节。除了与N2固定相关的基因被调节外，可能与植物-微生物相互作用相关的基因簇和一些转录因子的表达也显著增强。在n2固定条件下，野生型与NifA插入突变体（NifA是nif基因必需的转录激活因子）的转录组进行比较，发现24.5%的基因组表达受到影响。对29个NifA结合序列进行全基因组预测，这些序列与25个靶基因相匹配，这些基因在微阵列分析中表达差异，其中一些被NifA推定为负调控。对于选定的基因，通过实时RT-PCR研究证实了差异表达。我们的数据表明，固氮条件下的生活是BH72菌株在根中的生活方式的重要组成部分，因为远超过nif调控的广泛基因被调节。此外，菌株BH72中的NifA调控似乎包含了更广泛的细胞功能，而不仅仅是nif基因的调控。
The model endophyte Azoarcus sp. strain BH72 is known to contribute fixed nitrogen to its host Kallar grass and also expresses nitrogenase genes endophytically in rice seedlings. Availability of nitrogen is a signal regulating the transcription of nitrogenase genes. Therefore, we analysed global transcription in response to differences in the nitrogen source. A DNA microarray, comprising 70-mer oligonucleotides representing 3989 open reading frames of the genome of strain BH72, was used for transcriptome studies. Transcription profiles of cells grown microaerobically on N2 versus ammonium were compared. Expression of 7.2% of the genes was significantly up-regulated, and 5.8% down-regulated upon N2 fixation, respectively. A parallel genome-wide prediction of σ54-type promoter elements mapped to the upstream region of 38 sequences of which 36 were modulated under the N2 response. In addition to modulation of genes related to N2 fixation, the expressions of gene clusters that might be related to plant-microbe interaction and of several transcription factors were significantly enhanced. While comparing under N2-fixation conditions the transcriptome of wild type with a nifLA− insertion mutant, NifA being the essential transcriptional activator for nif genes, 24.5% of the genome was found to be affected in expression. A genome-wide prediction of 29 NifA binding sequences matched to 25 of the target genes whose expression was differential during microarray analysis, some of which were putatively negatively regulated by NifA. For selected genes, differential expression was corroborated by real time RT-PCR studies. Our data suggest that life under conditions of nitrogen fixation is an important part of the lifestyle of strain BH72 in roots, as a wide range of genes far beyond the nif regulon is modulated. Moreover, the NifA regulon in strain BH72 appears to encompass a wider range of cellular functions beyond the regulation of nif genes.
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