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Genome-wide screen and functional analysis in Xanthomonas reveal a large number of mRNA-derived sRNAs, including the novel RsmA-sequester RsmU

Genome-wide screen and functional analysis in Xanthomonas reveal a large number of mRNA-derived sRNAs, including the novel RsmA-sequester RsmU
Genome-wide screen and functional analysis in Xanthomonas reveal a large number of mRNA-derived sRNAs, including the novel RsmA-sequester RsmU

Although bacterial small noncoding RNAs (sRNAs) are known to play a critical role in various cellular processes, including pathogenesis, the identity and action of such sRNAs are still poorly understood in many organisms. Here we have performed a genome-wide screen and functional analysis of the sRNAs in Xanthomonas campestris pv. campestris (Xcc), an important phytopathogen. The 50-500-nt RNA fragments isolated from the wild-type strain grown in a virulence gene-inducing condition were sequenced and a total of 612 sRNA candidates (SRCs) were identified. The majority (82%) of the SRCs were derived from mRNA, rather than specific sRNA genes. A representative panel of 121 SRCs were analysed by northern blotting; 117 SRCs were detected, supporting the contention that the overwhelming majority of the 612 SRCs identified are indeed sRNAs. Phenotypic analysis of strains overexpressing different candidates showed that a particular sRNA, RsmU, acts as a negative regulator of virulence, the hypersensitive response, and cell motility in Xcc. In vitro electrophoretic mobility shift assay and in vivo coimmunoprecipitation analyses indicated that RsmU interacted with the global posttranscriptional regulator RsmA, although sequence analysis displayed that RsmU is not a member of the sRNAs families known to antagonize RsmA. Northern blotting analyses demonstrated that RsmU has two isoforms that are processed from the 3'-untranslated region of the mRNA of XC1332 predicted to encode ComEA, a periplasmic protein required for DNA uptake in bacteria. This work uncovers an unexpected major sRNA biogenesis strategy in bacteria and a hidden layer of sRNA-mediated virulence regulation in Xcc.

RsmA, RsmU, Xanthomonas, sRNA, small noncoding RNA, virulence
1464-6722
1573-1590
Tang, Dong-Jie
46609bbf-2dd4-4ec8-8a7b-a6d6a3b8983b
Chen, Xiao-Lin
9546927b-2f40-4e20-994f-d5ea49c55136
Jia, Yu
d8ac25b5-adcb-4e93-9d59-ea78ce928584
Liang, Yu-Wei
2f2427c0-b513-4584-9beb-e3dc01d7aeec
He, Yuan-Ping
5a076839-e494-4563-8e39-f2db5d95eaa2
Lu, Ting-Ting
35e96ff6-4cc6-422f-8b59-120ae68c0df8
Zhu, Chuan-Rang
68faec7d-dfe8-42a6-ab1d-4ea51ce050b4
Han, Bin
af3f99ed-8fe1-4bea-ac06-432b5ac17725
An, Shi-Qi
0e05f480-cec1-4c0e-bc1d-359d30ea9a6e
Tang, Ji-Liang
74e46e1a-4f75-4c94-871d-317339f309f2
Tang, Dong-Jie
46609bbf-2dd4-4ec8-8a7b-a6d6a3b8983b
Chen, Xiao-Lin
9546927b-2f40-4e20-994f-d5ea49c55136
Jia, Yu
d8ac25b5-adcb-4e93-9d59-ea78ce928584
Liang, Yu-Wei
2f2427c0-b513-4584-9beb-e3dc01d7aeec
He, Yuan-Ping
5a076839-e494-4563-8e39-f2db5d95eaa2
Lu, Ting-Ting
35e96ff6-4cc6-422f-8b59-120ae68c0df8
Zhu, Chuan-Rang
68faec7d-dfe8-42a6-ab1d-4ea51ce050b4
Han, Bin
af3f99ed-8fe1-4bea-ac06-432b5ac17725
An, Shi-Qi
0e05f480-cec1-4c0e-bc1d-359d30ea9a6e
Tang, Ji-Liang
74e46e1a-4f75-4c94-871d-317339f309f2

Tang, Dong-Jie, Chen, Xiao-Lin, Jia, Yu, Liang, Yu-Wei, He, Yuan-Ping, Lu, Ting-Ting, Zhu, Chuan-Rang, Han, Bin, An, Shi-Qi and Tang, Ji-Liang (2020) Genome-wide screen and functional analysis in Xanthomonas reveal a large number of mRNA-derived sRNAs, including the novel RsmA-sequester RsmU. Molecular Plant Pathology, 21 (12), 1573-1590. (doi:10.1111/mpp.12997).

Record type: Article

Abstract

Although bacterial small noncoding RNAs (sRNAs) are known to play a critical role in various cellular processes, including pathogenesis, the identity and action of such sRNAs are still poorly understood in many organisms. Here we have performed a genome-wide screen and functional analysis of the sRNAs in Xanthomonas campestris pv. campestris (Xcc), an important phytopathogen. The 50-500-nt RNA fragments isolated from the wild-type strain grown in a virulence gene-inducing condition were sequenced and a total of 612 sRNA candidates (SRCs) were identified. The majority (82%) of the SRCs were derived from mRNA, rather than specific sRNA genes. A representative panel of 121 SRCs were analysed by northern blotting; 117 SRCs were detected, supporting the contention that the overwhelming majority of the 612 SRCs identified are indeed sRNAs. Phenotypic analysis of strains overexpressing different candidates showed that a particular sRNA, RsmU, acts as a negative regulator of virulence, the hypersensitive response, and cell motility in Xcc. In vitro electrophoretic mobility shift assay and in vivo coimmunoprecipitation analyses indicated that RsmU interacted with the global posttranscriptional regulator RsmA, although sequence analysis displayed that RsmU is not a member of the sRNAs families known to antagonize RsmA. Northern blotting analyses demonstrated that RsmU has two isoforms that are processed from the 3'-untranslated region of the mRNA of XC1332 predicted to encode ComEA, a periplasmic protein required for DNA uptake in bacteria. This work uncovers an unexpected major sRNA biogenesis strategy in bacteria and a hidden layer of sRNA-mediated virulence regulation in Xcc.

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More information

Accepted/In Press date: 26 August 2020
e-pub ahead of print date: 23 September 2020
Published date: December 2020
Keywords: RsmA, RsmU, Xanthomonas, sRNA, small noncoding RNA, virulence

Identifiers

Local EPrints ID: 444300
URI: http://eprints.soton.ac.uk/id/eprint/444300
ISSN: 1464-6722
PURE UUID: 9ab47a38-4397-4116-bb69-2a822eeebf04
ORCID for Yuan-Ping He: ORCID iD orcid.org/0000-0003-3768-519X

Catalogue record

Date deposited: 09 Oct 2020 16:35
Last modified: 16 Mar 2024 09:28

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Contributors

Author: Dong-Jie Tang
Author: Xiao-Lin Chen
Author: Yu Jia
Author: Yu-Wei Liang
Author: Yuan-Ping He ORCID iD
Author: Ting-Ting Lu
Author: Chuan-Rang Zhu
Author: Bin Han
Author: Shi-Qi An
Author: Ji-Liang Tang

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