NCR343 is required to maintain the viability of differentiated bacteroids in nodule cells in Medicago truncatula

Bacteroid (name for rhizobia inside nodule cells) differentiation is a prerequisite for successful nitrogen-fixing symbiosis. In certain legumes, under the regulation of host proteins, for example, a large group of NCR (nodule cysteine rich) peptides, bacteroids undergo irreversible terminal differentiation. This process causes them to lose the ability to propagate inside nodule cells while boosting their competency for nitrogen fixation. How host cells maintain the viability of differentiated bacteroids while maximizing their nitrogen-reducing activities remains elusive. Here, through mutant screen, map-based cloning, and genetic complementation, we find that NCR343 is required for the viability of differentiated bacteroids. In Medicago truncatula debino1 mutant, differentiated bacteroids decay prematurely, and NCR343 is proved to be the casual gene for debino1. NCR343 is mainly expressed in the nodule fixation zone, where bacteroids are differentiated. In nodule cells, mature NCR343 peptide is secreted into the symbiosomes. RNA-Seq assay shows that many stress-responsive genes are significantly induced in debino1 bacteroids. Additionally, a group of stress response-related rhizobium proteins are identified as putative interacting partners of NCR343. In summary, our findings demonstrate that beyond promoting bacteroid differentiation, NCR peptides are also required in maintaining the viability of differentiated bacteroids.