Abstract
Erwinia amylovora, causative agent of fire blight disease of apple and pear trees, has evolved to use small RNAs for post-transcriptional regulation of virulence traits important for disease development. The sRNA ArcZ regulates several virulence traits, and to better understand its roles, we conducted a transcriptomic comparison of wild-type and ΔarcZ mutant E. amylovora. We found that ArcZ regulates multiple cellular processes including genes encoding enzymes involved in mitigating the threat of reactive oxygen species (katA, tpx, osmC), and that the ΔarcZ mutant has reduced catalase activity and is more susceptible to exogenous hydrogen peroxide. We quantified hydrogen peroxide production by apple leaves inoculated with E. amylovora and found that the while wild-type E. amylovora cells produce enough catalase to cope with defense peroxide, the ΔarcZ mutant is likely limited in virulence because of inability to cope with peroxide levels in host leaves. We further found that the ArcZ regulon overlaps significantly with the regulons of transcription factors involved in oxidative sensing including Fnr and ArcA. In addition, we show that ArcZ regulates arcA at the post-transcriptional level suggesting a role for this system in mediating adaptations to oxidative state, especially during disease development.
Highlights
When pathogenic microbes arrive on a host plant, the plant perceives the arrival of a threat through recognition of pathogen associated molecular patterns (PAMPs) (Jones and Dangl, 2006)
The several genes and pathways involved in amino acid biosynthesis and metabolism are likely targets of the transcription factor Lrp, which is known to be regulated by ArcZ (Schachterle and Sundin, 2019) and which we found to be differentially regulated in the arcZ mutant in our transcriptomic analysis
In our search of differentially expressed genes that have links to the oxidative stress response, we found katA, encoding a catalase, tpx, encoding a thiol-peroxidase, and osmC, encoding an osmotically inducible peroxiredoxin. katA and osmC were both down-regulated in the arcZ mutant, and tpx mRNA was more abundant (Figure 3A)
Summary
When pathogenic microbes arrive on a host plant, the plant perceives the arrival of a threat through recognition of pathogen associated molecular patterns (PAMPs) (Jones and Dangl, 2006). In E. amylovora, 42 Hfq-dependent sRNAs have been identified, and the Hfq-dependent sRNA ArcZ in particular is critical for virulence and several virulence-associated traits including production of the exopolysaccharides levan and amylovoran, normal biofilm formation, flagellar motility and translocation of type III effectors to plant cells (Zeng and Sundin, 2014). We have recently shown that ArcZ regulates flagellar motility in E. amylovora through a direct interaction with the flagellar master regulator FlhD (Schachterle et al, 2019) and that ArcZ impacts exopolysaccharide production and biofilm formation through the leucine responsive regulator protein Lrp (Schachterle and Sundin, 2019) It is not known if there are further virulence-associated traits being regulated by ArcZ, nor is it known how ArcZ regulates type III secretion. We further present evidence that ArcZ regulates arcA post-transcriptionally, indicating that ArcZ plays a major role in the oxidative status responsive regulatory pathways
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