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继2015年年底,我们报道了苦参(Sophora flavescens)可以与多属种不同的根瘤菌结瘤固氮的现象之后(见博文:一种豆科植物能与多少种根瘤菌共生?http://blog.sciencenet.cn/blog-3533-945737.html ),又经过两年的深入研究,我们又进一步从基因突变体、结瘤共生表型、结瘤因子结构、Tn5插入突变体等角度解释了苦参与不同根瘤菌混杂性的互作共生机制。论文题目是:“苦参与不同根瘤菌之间的非专一性共生(Non-specific symbiosis between Sophora flavescens and different rhizobia )”,日前已在MolecularPlant-Microbe Interaction (MPMI)上在线发表(http://apsjournals.apsnet.org/doi/abs/10.1094/MPMI-05-17-0117-R?journalCode=mpmi,2017年9月20日)。
在这篇论文中,我们删除或插入突变了来自四种豆科植物(大豆、锦鸡儿、菜豆、紫云英)的6种根瘤菌(Ensifer/Sinorhizobium fredii、Bradyrhizobium diazoefficiens、Rhizobium yanglingense、Mesorhizobium amorphae、Rhizobium etli、Mesorhizobium huakuii)中与结瘤因子骨架合成和外围基团修饰的相关基因nodC、nodE、nodM、nodZ、noeI、nodH、nodS,突变体在苦参及原宿主上的结瘤试验证明,除Bradyrhizobium diazoefficiens外,这些根瘤菌中的nodC、nodE、nodM基因删除后,都不能在苦参及原宿主上结瘤了。说明苦参及原宿主与根瘤菌的结瘤,均需要这些基因的存在,或者说是严格依赖于结瘤因子的存在。而参与结瘤因子结构修饰的基因(nodZ、noeI、nodH、nodS)删除后,则不影响这些菌株与苦参与原宿主的结瘤,但却改变了根瘤数量、固氮活性等。
来自于大豆的两种根瘤菌Sinorhizobium fredii和Bradyrhizobium diazoefficiens中,前者可以与苦参结瘤,但后者不能与苦参结有效根瘤,或者只能形成小的根瘤凸起。我们构建了Bradyrhizobium diazoefficiens的Tn5转座子突变体库,发现有14个突变体能与苦参结正常的根瘤了。而这些被突变的基因,在Bradyrhizobium diazoefficiens中参与物质的代谢、物质跨膜转运、细菌运动等功能,与之前报道的控制结瘤宿主范围的基因功能不同。
论文还获得了只产生“五聚体寡糖和脂酰基链,没有修饰的基本结瘤因子结构”的根瘤菌突变体,并对结瘤因子结构进行了鉴定,发现该突变体仍然能与原宿主大豆和苦参结瘤。但五聚体寡糖的单体——N-酰基葡萄糖胺不能诱导植物结瘤,也不能互补nodC突变体的菌株结瘤。该项工作进一步说明供试的宿主是严格依赖于结瘤因子的完整结构。
论文英文摘要如下:
We explored the genetic basis of thepromiscuous symbiosis of Sophoraflavescens with diverse rhizobia. To determine the impact of Nod factors(NFs) on the symbiosis of S. flavescens,nodulation-related gene mutants of representative rhizobial strains weregenerated. Strains with mutations in common nodulation genes (nodC, nodM, and nodE) failed tonodulate S. flavescens, indicatingthat the promiscuous nodulation of this plant is strictly dependent on thebasic NF structure. Mutations of the NF decoration genes nodH, nodS, nodZ, and noeI did not affect thenodulation of S. flavescens, butthese mutations affected the nitrogen fixation efficiency of the nodules.Wild-type Bradyrhizobiumdiazoefficiens USDA110 cannot nodulate S. flavescens, but we obtained 14 Tn5mutants of B. diazoefficiens thatnodulated S. flavescens. Thissuggested that the mutations had disrupted a negative regulator that preventsnodulation of S. flavescens, leadingto non-specific nodulation. For Ensiferfredii CCBAU 45436 mutants, the minimal NF structure was sufficient fornodulation of soybean and S.flavescens. In summary, the mechanism ofpromiscuous symbiosis of S. flavescenswith rhizobia might be related to its nonspecific recognition of NF structures,and the host specificity of rhizobia may also be controlled by currentlyunknown nodulation-related genes.
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