Effects of alteration in LPS structure in <i>Azorhizobium caulinodans</i> on nodule development

Abstract

2005). LPS is composed of three parts: lipid A, containing sugar and fatty acid, which forms the outer leaflet of the outer membrane and anchors LPS to the cell envelope; a core oligosaccharide region, a non-repeating oligosaccharide, which links lipid A and O-antigen; and the Oantigen, consisting of repeating oligosaccharide units. The LPS containing or lacking O-antigen is usually called smooth LPS and rough LPS, respectively. In the majority of gram-negative bacteria, the core oligosaccharide can be subdivided into an outer core and an inner core. The outer core region provides an attachment site for O-antigen. The inner core typically contains residues of 3-deoxy-D -manno-2-octulosonic acid (Kdo) and L-glycero-Dmanno-heptose. Kdo connects the inner core to the lipid A (Heinrichs et al., 1998;Raetz and Whitfield, 2002).Azorhizobium caulinodans ORS571 is a microsymbiont of a tropical legume, Sesbania rostrata (Dreyfus and Dommergues, 1981;Dreyfus et al., 1983Dreyfus et al., , 1988. N 2 -fixing nodules are formed by A. caulinodans on the stems as well as on the roots of S. rostrata. Previously, the whole genome sequence of A. caulinodans was determined (Lee et al., 2008), and we performed a concurrent large-scale screening of rhizobial genetic factors involved in nodule development using A. caulinodans mutants created by random Tn5 mutagenesis (Suzuki et al., 2007). As a result of this screening, we isolated three mutants having a Tn5-insertion in the putative LPS biosynthesis genes (Suzuki et al., 2007). These three mutants, Ao13-C11, Ao77-C09 and Ao80-F04, were disrupted in the rfaF, rfaD, and rfaE genes, respectively. These genes are involved in the synthesis of the LPS inner core region. The rfaD and rfaE genes encode ADP-L-glycero-D-manno-heptose-6-epimerase and ADP-L-glycero-D-manno-heptose synthase, respectively. These enzymes are involved in the synthesis of ADP-L-glycero-D-manno-heptose, which is a component of the LPS inner core (Kneidinger et al., 2002; The lipopolysaccharide (LPS) of Azorhizobium caulinodans ORS571, which forms N 2 -fixing nodules on the stems and roots of Sesbania rostrata, is known to be a positive signal required for the progression of nodule formation. In this study, four A. caulinodans mutants producing a variety of defective LPSs were compared. The LPSs of the mutants having Tn5 insertion in the rfaF, rfaD, and rfaE genes were more truncated than the modified LPSs of the oac2 mutants. However, the nodule formation by the rfaF, rfaD, and rfaE mutants was more advanced than that of the oac2 mutant, suggesting that invasion ability depends on the LPS structure. Our hypothesis is that not only the wild-type LPSs but also the altered LPSs of the oac2 mutant may be recognized as signal molecules by plants. The altered LPSs may act as negative signals that halt the symbiotic process, whereas the wild-type LPSs may prevent the halt of the symbiotic process. The more truncated LPSs of the rfaF, rfaD, and rfaE mutants perhaps no longer function as negative signals inducing discontinu...

A Lipopolysaccharide Synthesis Gene rfaD from Mesorhizobium huakuii Is Involved in Nodule Development and Symbiotic Nitrogen Fixation

“…LPS is also considered as one of the essential components in the symbiosis between rhizobia and legumes [ 19 , 49 ]. The mutant of the rfaD gene of Azorhizobium caulinodans ORS571, which forms nodules on the stems and roots of Sesbania rostrata , was more advanced than the wild-type strain, but the mutant nodules were ineffective [ 50 ]. In this work, the inoculated 7653R ΔrfaD into the roots of A. sinicus , the symbiotic phenotype analysis showed that the shoots of the symbiotic plants grew poorly, and the leaves were yellow in color (an indicator of lacking N).…”