IN INOCULANTS Nodulaid - 17th International Nitrogen Fixation ...

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17 th International Congress on Nitrogen Fixation Fremantle, Western Australia 27 November – 1 December 2011 Session Details: Wednesday 30 November 2011 Concurrent Session 11 – Nodule Formation 1100 – 1230 Authors: Christian Staehelin, Ling Zhang, Ying-Ying Ge, Da-Wei Xin, Sha Liao and Zhi-Ping Xie State Key Laboratory of Biocontrol, School of Life Sciences, SunYat-Sen (Zhongshan) University, East Campus, Guangzhou 510006, China Presentation Title: Characterization of type 3 effectors in the Rhizobium-legume symbiosis Presentation Time: 1120 - 1140 Pathogenic bacteria use type 3 secretion systems to deliver virulence factors (type 3 effector proteins) into eukaryotic host cells. Similarly, type 3 effectors of certain nitrogen-fixing rhizobial strains affect nodule formation in the interaction with various host legumes. Here, we characterized NopL and NopM, two type 3 effectors of Rhizobium sp. strain NGR234. Nodulation tests and microscopic analysis showed that distinct necrotic areas were rapidly formed in ineffective nodules of Phaseolus vulgaris (cv. Tendergreen) induced by a nopL mutant, indicating that NopL antagonized nodule senescence. Further experiments revealed that NopL interfered with mitogen-activated protein kinase (MAPK) signaling in yeast and plant cells (Nicotiana tabacum). Expression of nopL in yeast disrupted the mating pheromone (α-factor) response pathway, whereas nopL expression in N. tabacum suppressed cell death induced by over-expression of the MAPK gene SIPK (salicylic acid-induced protein kinase). NopL was multiply phosphorylated either in yeast or N. tabacum cells that expressed nopL. Four phosphorylated serines were confirmed by mass spectrometry. All four phosphorylation sites exhibit a Ser-Pro pattern, a typical motif in MAPK substrates. These data suggest that NopL is a suppressor of MAPK signaling. NopM, another type 3 effector of Rhizobium sp. NGR234 displayed E3 ubiquitin ligase activity in vitro, whereas the mutant protein NopM-C338A was inactive. Corresponding mutant analysis indicated that NopM also acts as an E3 ubiquitin ligase during symbiosis with Lablab purpureus, providing evidence that NopM is delivered into legume host cells and targets host proteins. Finally, effects of NopM on eukaryotic cells will be reported. Zhang L, Chen X-J, Lu H-B, Xie Z-P, & Staehelin C (2011). Functional analysis of the type 3 effector NopL from Rhizobium sp. NGR234: symbiotic effects, phosphorylation and interference with MAPK signaling. J. Biol. Chem. in press 72 2011
17 th International Congress on Nitrogen Fixation Fremantle, Western Australia 27 November – 1 December 2011 Session Details: Wednesday 30 November 2011 Concurrent Session 11 – Nodule Formation 1100 – 1230 Authors: Masaki Hanyu 1,2 , Shin Okazaki 1 , Rie Shirai 1 and Kazuhiko Saeki 1 1 Department of Biological Sciences, Faculty of Science, Nara Women’s University, Nara630-8506 2 Department of Biological Sciences, Graduate School of Science, Osaka University, Toyonaka, Osaka 560-0043, Japan Presentation Title: Mesorhizobium loti mutant lacking superoxide dismutase displays Lotus-host accessiondependent nitrogen fixation capacities Presentation Time: 1140 - 1200 Protection against reactive oxygen species (ROS) is important for legume-nodulating rhizobia during the establishment and maintenance of symbiosis as well as under free-living conditions because legume hosts might assail incoming microbes with ROS and because nitrogenase is extremely sensitive to ROS. We generated a deletion mutant of the putative superoxide dismutase (SOD) gene, with locus tag mlr7636, in Mesorhizobium loti MAFF303099 to investigate physiological significance. Under free-living conditions, the mutant displayed a number of properties typical of SOD-lacking strains. Its cell extract showed no general SOD activity under the assay conditions that enabled to detect the activity of the wild-type. It proliferated very slowly and showed elevated sensitivity to exogenous superoxide generators. Its spontaneous mutation rate was approximately 20times of that of wild-type. These defects were complemented by re-introduction of the wild-type gene. Based on the result and sequence similarity with Escherichia coli enzyme, we named the gene sodA. With regard to symbiotic capacities with Lotus japonicus, the sodA mutant showed quasi-wild-type nodulation and nitrogen fixation efficiencies with the host accession Gifu B-129, whereas it showed low nodulation efficiency and poor support of plant growth with the host accession Miyakojima MG-20. 73 2011
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INVENTING THE FUTURE IN INOCULANTS
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IN INOCULANTS Nodulaid - 17th International Nitrogen Fixation ...

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