Transcriptome Analysis Revealed Overlapping and Special Regulatory Roles of RpoN1 and RpoN2 in Motility, Virulence, and Growth of Xanthomonas oryzae pv. oryzae

Abstract

σ54 factor (RpoN) plays a crucial role in bacterial motility, virulence, growth, and other biological functions. In our previous study, two homologous σ54 factors, RpoN1 and RpoN2, were identified in Xanthomonas oryzae pv. oryzae (Xoo), the causative agent of bacterial leaf blight in rice. However, their functional roles, i.e., whether they exert combined or independent effects, remain unknown. In the current study, rpoN1 or rpoN2 deletion in Xoo significantly disrupted bacterial swimming motility, flagellar assembly, and virulence. Transcriptome analysis led to the identification of 127 overlapping differentially expressed genes (DEGs) regulated by both RpoN1 and RpoN2. Furthermore, GO and KEGG classification demonstrated that these DEGs were highly enriched in flagellar assembly, chemotaxis, and c-di-GMP pathways. Interestingly, ropN1 deletion decreased ropN2 transcription, while rpoN2 deletion did not affect ropN1 transcription. No interaction between the rpoN2 promoter and RpoN1 was detected, suggesting that RpoN1 indirectly regulates rpoN2 transcription. In addition, RpoN1-regulated DEGs were specially enriched in ribosome, carbon, and nitrogen metabolism pathways. Besides, bacterial growth was remarkably repressed in ΔrpoN1 but not in ΔrpoN2. Taken together, this study demonstrates the overlapping and unique regulatory roles of RpoN1 and RpoN2 in motility, virulence, growth and provides new insights into the regulatory mechanism of σ54 factors in Xoo.

WITHDRAWN: Sucrose-dependence of sugar uptake, quorum sensing and virulence of the rice blight pathogenXanthomonas oryzaepv.oryzae

“…Notably, transcripts of the key sigma factor σ 54 , RpoN1 and RpoN2, and its downstream Gum gene targets, which are responsible for EPS production, were also sucrose inducible as shown here. Since RpoN1 and RpoN2 do not appear to be involved in rpfC and rpfG regulation, the two pathways may operate independently (40). Taken together we propose a model in which pioneering Xoo manages enter the xylem, attach to the xylem parenchyma cells and inject TAL effectors into the host cells via a T3SS.…”

The Regulatory Functions of σ54 Factor in Phytopathogenic Bacteria

“…Two copies of rpoN , namely, rpoN1 and rpoN2 , were identified in the Xanthomonas genome. rpoN1 and rpoN2 are primarily located in a phosphotransferase system and a large flagellar gene cluster, respectively [ 57 , 58 , 59 , 60 ]. The nucleotide sequences of rpoN1 and rpoN2 are not identical, and their protein sequences share 39% identity [ 61 ].…”

“…Furthermore, FliA controls the expression of flagellin protein FliC, flagellar cap protein FliD, flagellar chaperone proteins FliS and FliTX, and anti-σ 28 factor FlgM [ 61 ]. Moreover, RpoN2/FleQ regulates the transcription of flagellin glycosylation-related genes ( gigx1 – gigx10 ), chemotaxis-associated genes ( cheYZA ), and c-di-GMP-related genes ( PXO 06199 , PXO 06201 , and PXO 06202 ) [ 59 , 77 ]. Deletions in rpoN2 , fleQ , fliC, and fliA resulted in the loss of flagella and swimming motility in Xoo .…”

“…Another σ 54 factor, RpoN1, was also identified in Xoo . Transcriptome analysis showed that RpoN1 and RpoN2 regulate more than 30 genes in flagellar regulon [ 59 ]. Interestingly, the rpoN2 expression level was decreased in the rpoN1 mutant; moreover, abnormal flagellum and decreased swimming motility were also reported in the rpoN1 mutant [ 59 ].…”

“…Transcriptome analysis showed that RpoN1 and RpoN2 regulate more than 30 genes in flagellar regulon [ 59 ]. Interestingly, the rpoN2 expression level was decreased in the rpoN1 mutant; moreover, abnormal flagellum and decreased swimming motility were also reported in the rpoN1 mutant [ 59 ]. As per a previous study, RpoN1 indirectly regulates the RpoN2 transcription in Xoo [ 59 ].…”