Rhizobial infection-specific accumulation of phosphatidylinositol 4,5-bisphosphate inhibits the excessive infection of rhizobia in Lotus japonicus

During the symbiosis of legume with nitrogen-fixing bacteria, collectively called rhizobia, suppression of excessive rhizobial infection by host plants is important to maximize the benefits of symbiotic nitrogen fixation. However, it remains relatively poorly understood the molecular mechanism involved in the suppression.

We performed LC-MS and RNA-seq analysis using rhizobia-infected Lotus japonicus roots and investigated the role of phosphatidylinositol (PI) and phosphatidylinositol phosphates (PIPs) in the symbiosis. Phosphatidylinositol transfer protein ( PITP ) -like proteins 4 ( PLP4 ) , phosphatidylinositol 3-phosphate 5-kinase 4 ( PIP5K4 ) and PIP5K6 mutants, which are involved in the vesicular transport of lipids and phosphorylation of PIPs, were used to show the involvement of the signaling of PI and PIPs. Accumulation of phosphatidylinositol 4,5-bisphosphate [PI(4,5)P ₂ ] during rhizobial infection were examined by a fluorescent marker 1xTUBBY-C (TUBBY).

We found that PI signaling-related genes were upregulated, and the amount of PIP ₂ increased in L. japonicus roots during rhizobial infection. In the PLP4 , PIP5K4 and PIP5K6 mutants, rhizobial infection increased and PIP ₂ accumulation was failed. Furthermore, the observation of PI(4,5)P ₂ in rhizobia-infected roots revealed that the ectopic accumulation was closely related to suppression of rhizobial infection.

Our findings indicate that the accumulation of PI(4,5)P ₂ , which is mediated by PLP and PIP5Ks, suppresses excessive rhizobial infection in the root epidermis and cortex, leading to the optimal number of nodules.