Gene expression and localization of a β-1,3-glucanase of Lotus japonicus
Phytohormone abscisic acid (ABA) inhibits root nodule formation of leguminous plants. LjGlu1, a β-1,3-glucanase gene of Lotus japonicus, has been identified as an ABA responsive gene. RNA interference of LjGlu1 increased nodule number. This suggests that LjGlu1 is involved in the regulation of nodule formation. Host legumes control nodule number by autoregulation of nodulation (AON), in which the presence of existing root nodules inhibits further nodulation. For further characterization of LjGlu1, we focused on the expression of LjGlu1 in relation to AON. In a split-root system, LjGlu1 expression peaked when AON was fully induced. Hairy roots transformed with LjCLE-RS1, a gene that induces AON, were generated. Expression of LjGlu1 was greater in the transgenic roots than in untransformed roots. LjGlu1 was not induced in a hypernodulating mutant inoculated with Mesorhizobium loti. These results suggest that the expression of LjGlu1 is involved in the system of AON. However, neither hypernodulation nor enlarged nodulation zone was observed on the transgenic hairy roots carrying LjGlu1-RNAi, suggesting that LjGlu1 is not a key player of AON. Recombinant LjGlu1 showed endo-β-1,3-glucanase activity. LjGlu1-mOrange fusion protein suggested that LjGlu1 associated with M. loti on the root hairs. Exogenous β-1,3-glucanase inhibited infection thread formation by both the wild type and the mutant, and nodule numbers were reduced. These results suggest that LjGlu1 is expressed in response to M. loti infection and functions outside root tissues, resulting in the inhibition of infection.
Keywordsβ-1,3-glucanase Infection Legume Lotus japonicus Mesorhizobium loti Nodulation
We thank Prof. Kawaguchi (National Institute for Basic Biology) for providing the seeds of har1-7 and the overexpress construct of LjCLE-RS1. We also thank the National BioResource Project Lotus/Glycine for providing the seeds of L. japonicus ‘Miyakojima’ MG-20.
- Gerhard LM, Frederick M Jr (1999) Functions and regulation of plant β-1,3-glucanases (PR-2). In: Datta SK, Muthukrishnan S (eds) Pathogenesis-related proteins in plants, 3rd edn. Florida, Boca Raton, pp 49–76Google Scholar
- Kaneko T, Nakamura Y, Sato S, Asamizu E, Kato T, Sasamoto S, Watanabe A, Idesawa K, Ishikawa A, Kawashima K, Kimura T, Kishida Y, Kiyokawa S, Kohara M, Matsumoto M, Matsuno A, Mochizuki Y, Nakayama S, Nakazaki N, Shimpo S, Sugimoto M, Takeuchi C, Yamada M, Tabata S (2000) Complete genome structure of the nitrogen-fixing symbiotic bacterium Mesorhizobium loti. DNA Res 7:331–338CrossRefPubMedGoogle Scholar
- Kawaharada Y, Kelly S, Nielsen MW, Hjuler CT, Gysel K, Muszyński A, Carlson RW, Thygesen MB, Sandal N, Asmussen MH, Vinther M, Andersen SU, Krusell L, Thirup S, Jensen KJ, Ronson CW, Blaise M, Radutoiu S, Stougaard J (2015) Receptor-mediated exopolysaccharide perception controls bacterial infection. Nature 523:308–312CrossRefPubMedGoogle Scholar
- Nakatsukasa-Akune M, Yamashita K, Shimoda Y, Uchiumi T, Abe M, Aoki T, Kamizawa A, Ayabe S, Higashi S, Suzuki A (2005) Suppression of root nodule formation by artificial expression of the TrEnodDR1 (coat protein of white clover cryptic virus 1) gene in Lotus japonicus. Mol Plant Microbe Interact 18:1069–1080CrossRefPubMedGoogle Scholar
- Offringa IA, Melchers LS, Regensburg-Tuink AJG, Costantino P, Schilperoort RA, Hooykaas PJJ (1986) Complementation of Agrobacterium tumefaciens tumor-inducing aux mutants by genes from the TR-region of the Ri plasmid of Agrobacterium rhizogenes. Proc Natl Acad Sci USA 83:6935–6939CrossRefPubMedPubMedCentralGoogle Scholar
- Takahara M, Magori S, Soyano T, Okamoto S, Yoshida C, Yano K, Sato S, Tabata S, Yamaguchi K, Shigenobu S, Takeda N, Suzaki T, Kawaguchi M (2013) TOO MUCH LOVE, a novel kelch repeat-containing F-box protein, functions in the long-distance regulation of the legume–Rhizobium symbiosis. Plant Cell Physiol 54:433–447CrossRefPubMedGoogle Scholar