Mutation And Symbiotic Phenotype Of Mesorhizobium Huakuii 7653R Cell Division Gene FtsZ1

ftsZ is an important cell-division gene, which ubiquitously exists in eubacteria, also in some Archaeas and eukaryotes. FtsZ protein is crucial for the division processes of prokaryotic cell and plastids, such as chloroplast and mitochondria. Once chromosome replication and separation are completed, FtsZ protein appears at the division site earliest, then participate in the initiation of division ring formation. Normally there is only one ftsZ gene in most species, but some particular ones have more than one. Recent studies reveal that some rhizobia species have two ftsZs:namely ftsZl and ftsZ2, which encoded two FtsZs with significant sequence similarity but difference in ultrastructure.Mesorhizobium huakuii 7653R, which could form indeterminate nodules on legume plant-Astragalus sinicus, has the typical characteristics of bacteroid differentiation in indeterminate nodules and two ftsZs gene in genome. This work aimed at investigating the function of M. huakuii 7653R cell-division gene ftsZl during the process of bacteroid differentiation. Firstly, a replacement vector, with which partial deletion in ftsZl gene, was constructed for the homologous recombination. Followed by tri-parental transconjugation and screening on selective medium, a mutant strain Z5-17 undergone double crossover was obtained. The phenotypes of mutant strain Z5-17 under both free-living and symbiotic conditions were further examined and analyzed in details. The results showed that Z5-17 displayed similar morphology, size and growth speed as compared with wild type strain 7653R under free-living condition. However, when host plant A. sinicus was in symbiosis with Z5-17, it showed obvious nitrogen-deficiency features and formed small-white-nodules with hardly nitrogenase activity, but the nodule number was dramatically increased. Furthermore, paraffin section and ultrastructure observations were carried out for the 25-dpi (day post inoculation) nodules formed by wild type strain 7653R and mutant Z5-17, respectively. It was found that the mutant strain Z5-17 displayed abnormal bacteroid differentiation, and the functional symbiosome development was blocked, consequently resulting in the interruption of rhizobial chronic infection programme and the lost of symbiotic nitrogen fixation ability. Paraffin section of nodules induced by mutant Z5-17 exhibited prematurely senescence features. Finally, by using the real time PCR, it was revealed that the expression level of ftsZl under free-living condition was much higher than that in symbiotic root nodules. In summary, although the mutation of ftsZl had no significant influences on the physiological attributes of free-living M. huakuii 7653R, it led to the impairment of symbiotic nitrogen fixation ability with A. sinicus. The results suggest that the normal expression of ftsZl is indispensable for bacteroid differentiation and functional symbiosome formation.