Functional studies of Ralstonia solanacearum novel pathogenicity determinants RSc0411 and murI
Date Issued
2009
Date
2009
Author(s)
Yang, Wen-Chien
Abstract
Ralstonia solanacearum is a soil-borne bacterium infecting vascular system, causing lethal wilting symptoms on many economically important crops and resulting severe crop losses. Previously, we carried out transposon (Tn5) insertional mutant screens to identify genes involved in pathogenesis of R. solanacearum. Two mutants containing a transposon insertion in RSc0411 or RSc1956 (murI) were found to be avirulent both on tomato and Arabidopsis. The aims of this study are to further elucidate roles of these genes in bacterial pathogenesis and the molecular mechanisms involved. Firstly, R. solanacearum RSc0411 is a homologue of E. coli lptC. LptC, a member of the novel protein family DUF1239, is suggested to be involved in E. coli lipopolysaccharides (LPS) transport into the outer membrane. Here we showed that RSc0411 mutant was defective in cell integrity and in rough LPS production. Notably, this mutant displayed defects in various pathogenesis-related properties and the induction of the type III secretion system was attenuated. The organization of DUF1239-related gene cluster is conserved among gram-negative bacteria, while sequence homology among orthologous genes in the cluster and those involved in LPS biogenesis varied according to phylogenetic relationships. Complementation tests revealed that only DUF1239 members in bacteria phylogenetically related to R. solanacearum were functional to rescue the mutant’s defects, further suggesting certain specificity in the RSc0411-involved pathogenesis mechanism and LPS biogenesis machineries may have evolved. Collectively these results imply a novel and crucial role of RSc0411 in early pathogenesis and LPS biogenesis of R. solanacearum. Secondly, R. solanacearum RSc1956 encodes glutamate racemase (MurI) protein. MurI catalyzes the conversion of L-glutamate to D-glutamate which is an essential component of peptidoglycans in bacterial cell walls. Here we showed that, although RSc1956 mutant behaved similar to the wild-type strain in biofilm formation, root attachment and production of plant cell wall degrading enzymes, it was defective in swimming motility and avirulent in tomato plants. Complementation tests further confirmed the key role of RSc1956 in R. solanacearum pathogenesis and revealed the importance of its promoter in precisely expressing MurI function. Taken together, these detailed studies on RSc0411 and RSc1956 are expected to pave the way not only for elucidating mechanisms and determinants involved in R. solanacearum pathogenesis but also potentially establishing useful disease control means.
Subjects
Ralstonia solanacearum
DUF1239
lipopolysaccharide
type III secretion system
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