Investigating the involvement of cytoskeletal proteins MreB and FtsZ in the origin of legume-rhizobial symbiosis

Rhizobia are rod-shaped bacteria that form nitrogen-fixing root nodules on leguminous plants, but they don't carry MreB, a key determinant of rod-like cell shape. Here, we introduced an actin-like homologue from pseudomonad into 7653R (a microsymbiont of L.) and examined the molecular, cellular and symbiotic phenotypes of the resultant mutant. Exogenous caused an enlarged cell size and slower growth in laboratory medium. However, the mutant formed small ineffective nodules on (Nod Fix ), and rhizobial cells in the infection zone were unable to differentiate into bacteroids. RNA-seq analysis also revealed minor effects of on global gene expression in free-living cells but larger effects for cells grown . Differentially expressed nodule-specific genes include cell cycle regulators such as the tubulin-like and . Unlike the ubiquitous FtsZ , FtsZ was commonly found in , and , but not in closely related non-symbiotic species. Bacterial Two-Hybrid analysis revealed that MreB interacts with FtsZ and FtsZ , which are targeted by the host-derived NCR peptides. Significantly, MreB mutation D283A disrupted the protein-protein interactions and restored the aforementioned phenotypic defects caused by MreB in . Together, our data indicate that MreB is detrimental for modern rhizobia and its interaction with FtsZ /FtsZ cause the symbiotic process to cease at the late stage of bacteroid differentiation. These findings led to a hypothesis that loss of in the common ancestor of and subsequent acquisition of are critical evolutionary steps leading to legume-rhizobial symbiosis.