umu.sePublications
Change search
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Stress response and virulence in Vibrio anguillarum
Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). (Debra L. Milton)
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Bacteria use quorum sensing, a cell to cell signaling mechanism mediated by small molecules that are produced by specific signal molecule synthases, to regulate gene expression in response to population density. In Vibrio anguillarum, the quorum-sensing phosphorelay channels information from three hybrid sensor kinases VanN, VanQ, CqsS that sense signal molecules produced by the synthases VanM, VanS and CqsA, onto the phosphotransferase VanU, to regulate activity of the response regulator VanO. VanO activates transcription of quorum-sensing regulatory RNAs (Qrr), which work together with the RNA chaperone Hfq to repress expression of the transcriptional regulator VanT.

The work presented in this thesis characterizes quorum-sensing independent and quorum-sensing dependent mechanisms that regulate VanT expression. Moreover, an in vivo imaging system was established, as a means to study V. anguillarum infections in the rainbow trout infection model.

Two quorum-sensing independent mechanisms regulating VanT expression were identified. First, the sigma factor RpoS indirectly activates VanT expression during transition into stationary growth phase by inhibiting hfq expression. Both, RpoS and VanT are crucial for stress response. Second, a type VI secretion system (T6SS) has a novel function as a signal sensing mechanism to regulate rpoS and vanT expression. Consequently, RpoS, quorum sensing and T6SS form a global network that senses stress and modulates stress response to ensure survival of the bacteria.

Further analysis of the quorum-sensing dependent regulation of VanT expression by the phosphorelay system revealed that four qrr genes are expressed continuously during growth. The phosphotransferase VanU is suggested to activate two response regulators, VanO and a predicted second response regulator. Activated VanO induces expression of the Qrr sRNAs, whereas, the predicted response regulator represses expression of the Qrr sRNAs. Thus, VanU has a pivotal role in the regulation of VanT expression. The signal synthase VanM and VanT form a regulatory loop, in which VanM represses VanT by inducing expression of the Qrr sRNAs and VanT directly activates vanM expression to repress its own expression. Moreover, Hfq destabilizes vanM mRNA, repressing vanM expression. VanT forms another regulatory loop with the transcriptional regulator LuxT, in which LuxT activates vanT expression and VanT directly represses luxT expression.

V. anguillarum is an opportunistic pathogen that causes vibriosis, a terminal hemorrhagic septicemia. The spatial and temporal progression of the infection was analyzed using the whole animal with an in vivo bioluminescent imaging method. Initial studies showed that colonization of the fish skin requires the siderophore, the RNA chaperone Hfq and the exopolysaccharide transport system, which protects against the innate immunity on the skin. Colonization of the fish skin is crucial for disease.

Place, publisher, year, edition, pages
Umeå: Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet) Umeå Universitet , 2010. , 86 p.
Keyword [en]
Vibrio anguillarum, quorum sensing, Type VI secretion, stress response, virulence, skin colonization
National Category
Biochemistry and Molecular Biology
Research subject
Molecular Biology
Identifiers
URN: urn:nbn:se:umu:diva-33269ISBN: 978-91-7264-958-3 (print)OAI: oai:DiVA.org:umu-33269DiVA: diva2:311166
Public defence
2010-05-18, NUS, byggnad 6L, Major Groove, Umeå Universitet, Umeå, 13:00 (English)
Opponent
Supervisors
Available from: 2010-04-27 Created: 2010-04-20 Last updated: 2011-03-14Bibliographically approved
List of papers
1. RpoS induces expression of the Vibrio anguillarum quorum-sensing regulator VanT
Open this publication in new window or tab >>RpoS induces expression of the Vibrio anguillarum quorum-sensing regulator VanT
2008 (English)In: Microbiology, ISSN 1350-0872, E-ISSN 1465-2080, Vol. 154, no 3, 767-780 p.Article in journal (Refereed) Published
Abstract [en]

In vibrios, regulation of the Vibrio harveyi-like LuxR transcriptional activators occurs post-transcriptionally via small regulatory RNAs (sRNAs) that destabilize the luxR mRNA at a low cell population, eliminating expression of LuxR. Expression of the sRNAs is modulated by the vibrio quorum-sensing phosphorelay systems. However, vanT mRNA, which encodes a LuxR homologue in Vibrio anguillarum, is abundant at low and high cell density, indicating that VanT expression may be regulated via additional mechanisms. In this study, Western analyses showed that VanT was expressed throughout growth with a peak of expression during late exponential growth. VanO induced partial destabilization of vanT mRNA via activation of at least one Qrr sRNA. Interestingly, the sigma factor RpoS significantly stabilized vanT mRNA and induced VanT expression during late exponential growth. This induction was in part due to RpoS repressing expression of Hfq, an RNA chaperone. RpoS is not part of the quorum-sensing regulatory cascade since RpoS did not regulate expression or activity of VanO, and RpoS was not regulated by VanO or VanT. VanT and RpoS were needed for survival following UV irradiation and for pigment and metalloprotease production, suggesting that RpoS works with the quorum-sensing systems to modulate expression of VanT, which regulates survival and stress responses.

Place, publisher, year, edition, pages
Reading: Society for General Microbiology, 2008
National Category
Microbiology in the medical area
Research subject
Infectious Diseases; Microbiology
Identifiers
urn:nbn:se:umu:diva-31617 (URN)10.1099/mic.0.2007/014167-0 (DOI)18310023 (PubMedID)
Available from: 2010-02-11 Created: 2010-02-11 Last updated: 2017-12-12Bibliographically approved
2. Type VI secretion modulates quorum sensing and stress response in Vibrio anguillarum.
Open this publication in new window or tab >>Type VI secretion modulates quorum sensing and stress response in Vibrio anguillarum.
Show others...
2009 (English)In: Environmental Microbiology, ISSN 1462-2912, E-ISSN 1462-2920, Vol. 11, no 12, 3018-3028 p.Article in journal (Refereed) Published
Abstract [en]

Type VI protein secretion systems (T6SS) are essential for virulence of several Gram-negative bacteria. In this study, we identified a T6SS in Vibrio anguillarum, a marine bacterium that causes a hemorrhagic septicemia in fish. A partial operon vtsA-H (vibrio type six secretion) was sequenced and shown to encode eight proteins. VtsE-H are signature proteins found in other T6SSs, while VtsA-D are not associated with T6SS studied so far. In-frame deletions were made in each gene. Secretion of a haemolysin-co-regulated-like protein (Hcp), a protein secreted by all studied T6SSs, was decreased in VtsE-H. Unexpectedly, VtsA, VtsC and VtsD activated while VtsB and VtsE-H repressed hcp expression. The T6SS proteins also regulated expression of two extracellular proteases, EmpA and PrtV, but inversely to Hcp expression. This regulation was indirect as T6S positively regulated expression of the stress-response regulator RpoS and the quorum-sensing regulator VanT, which positively regulate protease expression. Moreover, VtsA-H proteins were not needed for virulence but did play a role in various stress responses. Thus, these data characterize a new role for T6S in the ecology of bacteria and we hypothesize this role to be a signal sensing mechanism that modulates the expression of regulators of the general stress response.

National Category
Microbiology in the medical area
Identifiers
urn:nbn:se:umu:diva-31616 (URN)10.1111/j.1462-2920.2009.02005.x (DOI)19624706 (PubMedID)
Available from: 2010-02-11 Created: 2010-02-11 Last updated: 2017-12-12Bibliographically approved
3. The transcriptional regulator VanT activates expression of the signal synthase VanM forming a regulatory loop in the Vibrio anguillarum quorum sensing system
Open this publication in new window or tab >>The transcriptional regulator VanT activates expression of the signal synthase VanM forming a regulatory loop in the Vibrio anguillarum quorum sensing system
Show others...
(English)Manuscript (preprint) (Other academic)
Abstract [en]

In Vibrio anguillarum, one quorum-sensing regulatory sRNA, Qrr1, was previously shown to destabilize vanT mRNA repressing expression of the transcriptional regulator VanT. In this study, three additional Qrr sRNAs that destabilize vanT mRNA were identified. The expression of the Qrr sRNA depends on the active (phosphorylated) form of the σ54-dependent response regulator VanO and the sigma factor RpoN (σ54). The phosphotransferase VanU is predicted to activate VanO, but also a putative second response regulator that may repress qrr expression activating VanT expression. Signal production by the homoserine lactone synthase VanM inhibits the phosphorelay determining VanU activity. Therefore, qrr expression and VanT were analyzed in the ΔvanM mutant. Interestingly, VanM activated qrr expression and repressed VanT expression. Moreover, vanM expression is tightly regulated and peaks early in growth. Several putative Hfq binding sites are present in the vanM 5´-untranslated region. Thus, vanM expression and mRNA stability were measured in the Δhfq mutant. Hfq destabilizes vanM mRNA, but also regulates vanM transcription. The transcriptional effect occurs indirectly through VanT, which is derepressed in the Δhfq mutant. Additionally, VanT directly binds to a conserved binding motif in the vanM promoter to positively regulate vanM expression. In summary, VanM and VanT generate a regulatory loop, in which VanM induces qrr expression to repress VanT expression and VanT negatively regulates its own expression by activating vanM expression.

National Category
Biochemistry and Molecular Biology
Research subject
Molecular Biology
Identifiers
urn:nbn:se:umu:diva-33256 (URN)
Available from: 2010-04-20 Created: 2010-04-20 Last updated: 2010-04-29Bibliographically approved
4. Colonization of fish skin is vital for Vibrio anguillarum to cause disease
Open this publication in new window or tab >>Colonization of fish skin is vital for Vibrio anguillarum to cause disease
2010 (English)In: Environmental Microbiology Reports, ISSN 1758-2229, Vol. 2, no 1, 133-139 p.Article in journal (Refereed) Published
Abstract [en]

Vibrio anguillarum causes a fatal haemorrhagic septicaemia in marine fish. During initial stages of infection, host surfaces are colonized; however, few virulence factors required for colonization of the host are identified. In this study, in vivo bioluminescent imaging was used to analyse directly the colonization of the whole rainbow trout animal by V. anguillarum. The wild type rapidly colonized both the skin and the intestines by 24 h; however, the bacterial numbers on the skin were significantly higher than in the intestines indicating that skin colonization may be important for disease to occur. Mutants defective for the anguibactin iron uptake system, exopolysaccharide transport, or Hfq, an RNA chaperone, were attenuated for virulence, did not colonize the skin, and penetrated skin mucus less efficiently than the wild type.These mutants, however, did colonize the intestines and were as resistant to 2% bile salts as is the wildtype. Moreover, exopolysaccharide mutants were significantly more sensitive to lysozyme and antimicrobial peptides, while the Hfq and anguibactin mutants were sensitive to lysozyme compared with the wildtype. Vibrio anguillarum encodes several mechanisms to protect against antimicrobial components of skin mucus enabling an amazingly abundant growth on the skin enhancing its disease opportunities.

Place, publisher, year, edition, pages
Society for Applied Microbiology and Blackwell Publishing Ltd., 2010
Keyword
Vibrio anguillarum, skin colonization, vibriosis
National Category
Microbiology in the medical area
Research subject
Microbiology
Identifiers
urn:nbn:se:umu:diva-33156 (URN)10.1111/j.1758-2229.2009.00120.x (DOI)000279431900019 ()
Available from: 2010-04-13 Created: 2010-04-13 Last updated: 2010-04-27Bibliographically approved

Open Access in DiVA

fulltext(1726 kB)826 downloads
File information
File name FULLTEXT01.pdfFile size 1726 kBChecksum SHA-512
e477195c009bf48aa53ef0b23ec0f1962918051b6dbc0d2e862ab195a57f257cd0ca05f24b30bd1a1d806487ad7156a0450d184754ea3dbdc1a6ad2985aa0481
Type fulltextMimetype application/pdf

Search in DiVA

By author/editor
Weber, Barbara
By organisation
Department of Molecular Biology (Faculty of Science and Technology)
Biochemistry and Molecular Biology

Search outside of DiVA

GoogleGoogle Scholar
Total: 826 downloads
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

isbn
urn-nbn

Altmetric score

isbn
urn-nbn
Total: 773 hits
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf