Regulation of nitric oxide by phytoglobins in Lotus japonicus is involved in mycorrhizal symbiosis with Rhizophagus irregularis

Abstract

Various reactive molecular species are generated in plant–microbe interactions, and these species participate in defense and symbiotic responses. Leguminous plants successfully establish symbiosis by maintaining an appropriate level of nitric oxide (NO), which is generated in the roots and nodules during root nodule symbiosis. Phytoglobin (plant hemoglobin) controls NO levels in plants. In this study, we investigated mycorrhizal symbiosis, which occurs in more than 80% of land plants, between Rhizophagus irregularis and Lotus japonicus to clarify the involvement of phytoglobin-mediated NO regulation. The mycorrhizae of L. japonicus exhibited higher NO levels in the presence of R. irregularis than in its absence, especially at the infection site. LjGlb1–1, a phytoglobin that regulates NO level in L. japonicus, was upregulated during symbiosis with R. irregularis. In transformed hairy roots carrying the ProLjGlb1–1:GUS construct, LjGlb1–1 expression was observed at the R. irregularis infection site. We further examined the symbiotic phenotypes of L. japonicus lines with high and low LjGlb1–1 expression with R. irregularis. During mycorrhizal symbiosis, the high LjGlb1–1 expression line exhibited better growth than the wild-type, whereas the low expression line exhibited poor growth. In addition, the expression of LjPT4, a phosphate transporter specific to mycorrhizal symbiosis, was higher in the high LjGlb1–1 expression line, whereas that of the tubulin gene of R. irregularis was lower in the low LjGlb1–1 expression line than in the wild-type. These results confirm that NO regulation by LjGlb1–1 is involved in mycorrhizal symbiosis in L. japonicus, as it is reportedly in nitrogen-fixing symbiosis.