Submitted to: PLoS One
Publication Type: Peer reviewed journal
Publication Acceptance Date: 8/11/2012
Publication Date: 9/18/2012
Citation: Wang, D., Korban, S.S., Pusey, P.L., Zhao, Y.F. 2012. AmyR is a novel negative regulator of amylovoran production in Erwinia amylovora. PLoS One. 7(9):e45038 doi:10.1371/journal.pone.0045038. Interpretive Summary: Fire blight, caused by the bacterium Erwinia amylovora, is the most devastating bacterial disease on apples and pears, resulting in annual economic losses of around $100 million in the US alone. E. amylovora produces exopolysaccharide (EPS), a slime-like substance that is related to the ability of the bacterium to cause disease in pome fruit trees. One major EPS compound involved in fire blight development is amylovoran. In the present study, the protein AmyR, encoded by the gene amyR, was found to play an important role in regulating EPS production. Understanding the genetic and chemical aspects of EPS production will likely contribute to the development of effective strategies for controlling the fire blight disease.
Technical Abstract: We have previously reported the characterization of an orphan gene ybjN from Escherichia coli. In this study, we attempted to understand the role of amyR in Erwinia amylovora, a functionally conserved homolog of E. coli ybjN. As reported earlier, amylovoran production in the amyR knockout mutant is about ten-fold higher than that in the wild type (WT) strain. When multicopy plasmid containing the amyR gene was introduced into the amyR mutant or WT strains, amylovoran production was strongly inhibited. Furthermore, amylovoran production was also suppressed in various amylovoran-over-producing mutants such as grrSA containing multicopy of the amyR gene. Consistent with amylovoran production, an inverse correlation was observed between amyR expression and expression of amylovoran biosynthetic genes in vitro. However, the amyR knockout mutant and over-expression strains both showed reduced levan production, another exopolysaccharide produced by E. amylovora. Virulence assays demonstrated that, while the amyR mutant was capable of inducing slightly severe disease than that of the WT strain, strains over-expressing amyR gene were abolished in causing disease on apple shoots or leaves and caused reduced disease on immature pear fruits. Microarray studies revealed that amylovoran biosynthesis and related membrane protein-encoding genes were highly expressed in the amyR mutant, but down-regulated in amyR over-expression strains in vitro. Down-regulation of both amylovoran biosynthesis genes and some type III secretion (T3SS) genes in amyR over-expression strain partially explained why over expression of amyR led to non-pathogenic or reduced virulence in vivo. These results suggest that AmyR plays an important role in regulating exopolysaccharide production, and thus virulence in E. amylovora.