Investigating the Involvement of Cytoskeletal Proteins MreB and FtsZ in the Origin of Legume-Rhizobial Symbiosis

Abstract

Rhizobia are rod-shaped bacteria that form nitrogen-fixing root nodules on leguminous plants; however, they don’t carry MreB, a key determinant of rod-like cell shape. Here, we introduced an actin-like mreB homolog from a pseudomonad into Mesorhizobium huakuii 7653R (a microsymbiont of Astragalus sinicus L.) and examined the molecular, cellular, and symbiotic phenotypes of the resultant mutant. Exogenous mreB caused an enlarged cell size and slower growth in laboratory medium. However, the mutant formed small, ineffective nodules on A. sinicus (Nod + Fix − ), and rhizobial cells in the infection zone were unable to differentiate into bacteroids. RNA sequencing analysis also revealed minor effects of mreB on global gene expression in free-living cells but larger effects for cells grown in planta. Differentially expressed nodule-specific genes include cell cycle regulators such as the tubulin-like ftsZ 1 and ftsZ 2 . Unlike the ubiquitous FtsZ 1 , an FtsZ 2 homolog was commonly found in Rhizobium, Sinorhizobium, and Mesorhizobium spp. but not in closely related nonsymbiotic species. Bacterial two-hybrid analysis revealed that MreB interacts with FtsZ 1 and FtsZ 2 , which are targeted by the host-derived nodule-specific cysteine-rich peptides. Significantly, MreB mutation D283A disrupted the protein–protein interactions and restored the aforementioned phenotypic defects caused by MreB in M. huakuii. Together, our data indicate that MreB is detrimental for modern rhizobia and its interaction with FtsZ 1 and FtsZ 2 causes the symbiotic process to cease at the late stage of bacteroid differentiation. These findings led to a hypothesis that loss of mreB in the common ancestor of members of Rhizobiales and subsequent acquisition of ftsZ 2 are critical evolutionary steps leading to legume-rhizobial symbiosis. [Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .