umu.sePublications
Change search
Link to record
Permanent link

Direct link
BETA
Alternative names
Publications (10 of 53) Show all publications
Gurung, J. M., Amer, A., Francis, M., Costa, T., Chen, S., Zavialov, A. V. & Francis, M. S. (2018). Heterologous complementation studies with the YscX and YscY protein families reveals a specificity for Yersinia pseudotuberculosis type III secretion. Frontiers in Cellular and Infection Microbiology, 8, Article ID 80.
Open this publication in new window or tab >>Heterologous complementation studies with the YscX and YscY protein families reveals a specificity for Yersinia pseudotuberculosis type III secretion
Show others...
2018 (English)In: Frontiers in Cellular and Infection Microbiology, E-ISSN 2235-2988, Vol. 8, article id 80Article in journal (Refereed) Published
Abstract [en]

Type III secretion systems harbored by several Gram-negative bacteria are often used to deliver host-modulating effectors into infected eukaryotic cells. About 20 core proteins are needed for assembly of a secretion apparatus. Several of these proteins are genetically and functionally conserved in type III secretion systems of bacteria associated with invertebrate or vertebrate hosts. In the Ysc family of type III secretion systems are two poorly characterized protein families, the YscX family and the YscY family. In the plasmid-encoded Ysc-Yop type III secretion system of human pathogenic Yersinia species, YscX is a secreted substrate while YscY is its non-secreted cognate chaperone. Critically, neither an yscX nor yscY null mutant of Yersinia is capable of type III secretion. In this study, we show that the genetic equivalents of these proteins produced as components of other type III secretion systems of Pseudomonas aeruginosa (PscX and PscY), Aeromonas species (AscX and AscY), Vibrio species (VscX and VscY), and Photorhabdus luminescens (SctX and SctY) all possess an ability to interact with its native cognate partner and also establish cross-reciprocal binding to non-cognate partners as judged by a yeast two-hybrid assay. Moreover, a yeast three-hybrid assay also revealed that these heterodimeric complexes could maintain an interaction with YscV family members, a core membrane component of all type III secretion systems. Despite maintaining these molecular interactions, only expression of the native yscX in the near full-length yscX deletion and native yscY in the near full-length yscY deletion were able to complement for their general substrate secretion defects. Hence, YscX and YscY must have co-evolved to confer an important function specifically critical for Yersinia type III secretion.

Place, publisher, year, edition, pages
Frontiers Research Foundation, 2018
Keywords
T3S chaperone, secretion hierarchy, substrate sorting, LcrH/SycD, YscV, protein-protein interaction
National Category
Microbiology
Research subject
Microbiology; Molecular Biology; Infectious Diseases
Identifiers
urn:nbn:se:umu:diva-146348 (URN)10.3389/fcimb.2018.00080 (DOI)000427608900001 ()29616194 (PubMedID)
Funder
Swedish Research Council
Available from: 2018-04-05 Created: 2018-04-05 Last updated: 2018-06-09Bibliographically approved
Francis, M. S., Amer, A. A., Milton, D. & Costa, T. R. (2017). Site-directed mutagenesis and its application in studying the interactions of T3S components. In: Matthew L. Nilles and Danielle L. Jessen Condry (Ed.), Type 3 secretion systems: methods and protocols (pp. 11-31). Humana Press
Open this publication in new window or tab >>Site-directed mutagenesis and its application in studying the interactions of T3S components
2017 (English)In: Type 3 secretion systems: methods and protocols / [ed] Matthew L. Nilles and Danielle L. Jessen Condry, Humana Press, 2017, p. 11-31Chapter in book (Refereed)
Abstract [en]

Type III secretion systems are a prolific virulence determinant among Gram-negative bacteria. They are used to paralyze the host cell, which enables bacterial pathogens to establish often fatal infections—unless an effective therapeutic intervention is available. However, as a result of a catastrophic rise in infectious bacteria resistant to conventional antibiotics, these bacteria are again a leading cause of worldwide mortality. Hence, this report describes a pDM4-based site-directed mutagenesis strategy that is assisting in our foremost objective to better understand the fundamental workings of the T3SS, using Yersinia as a model pathogenic bacterium. Examples are given that clearly document how pDM4-mediated site-directed mutagenesis has been used to establish clean point mutations and in-frame deletion mutations that have been instrumental in identifying and understanding the molecular interactions between components of the Yersinia type III secretion system.

Place, publisher, year, edition, pages
Humana Press, 2017
Series
Methods in Molecular Biology, ISSN 1064-3745 ; 1531
Keywords
Site-directed mutagenesis, Type III secretion systems, Suicide vector pDM4, Mutant libraries, Genetic-based screens, Protein-protein interaction assays
National Category
Biochemistry and Molecular Biology
Research subject
Microbiology
Identifiers
urn:nbn:se:umu:diva-128091 (URN)10.1007/978-1-4939-6649-3_2 (DOI)27837478 (PubMedID)978-1-4939-6647-9 (ISBN)978-1-4939-6649-3 (ISBN)
Funder
Swedish Research Council, 2014–2105
Available from: 2016-11-22 Created: 2016-11-22 Last updated: 2018-06-09Bibliographically approved
Chen, S., Thompson, K. & Francis, M. S. (2016). Environmental Regulation of Yersinia Pathophysiology. Frontiers in Cellular and Infection Microbiology, 6, Article ID 25.
Open this publication in new window or tab >>Environmental Regulation of Yersinia Pathophysiology
2016 (English)In: Frontiers in Cellular and Infection Microbiology, E-ISSN 2235-2988, Vol. 6, article id 25Article in journal (Refereed) Published
Abstract [en]

Hallmarks of Yersinia pathogenesis include the ability to form biofilms on surfaces, the ability to establish close contact with eukaryotic target cells and the ability to hijack eukaryotic cell signaling and take over control of strategic cellular processes. Many of these virulence traits are already well-described. However, of equal importance is knowledge of both confined and global regulatory networks that collaborate together to dictate spatial and temporal control of virulence gene expression. This review has the purpose to incorporate historical observations with new discoveries to provide molecular insight into how some of these regulatory mechanisms respond rapidly to environmental flux to govern tight control of virulence gene expression by pathogenic Yersinia.

Place, publisher, year, edition, pages
Frontiers Media S.A, 2016
Keywords
acidity, temperature, metabolism, RovA, c-di-GMP, cAMP, extracytoplasmic stress, transition metals
National Category
Microbiology
Research subject
Microbiology
Identifiers
urn:nbn:se:umu:diva-117568 (URN)10.3389/fcimb.2016.00025 (DOI)000371266600001 ()26973818 (PubMedID)
Funder
Swedish Research Council, 2014-2105Swedish Research Council, 2014-6652
Note

Research in the author's own laboratories has been possible through the generous past or present funding support of the National Science Foundation of China (#31170133 and #31570132; SC), National Institute of General Medical Sciences of the National Institutes of Health (#SC2 GM105419; KT), Howard University Medical Alumni Association (KT), Medical Research Foundation of Umeå University (MF), Swedish Research Council (#2014-2105; MF), and Swedish Research Council framework grant—antibiotics and infection (#2014-6652; SC, KT, and MF). Work in the laboratory of MF is performed within the framework of the Umeå Centre for Microbial Research—Linnaeus Program.

Available from: 2016-03-02 Created: 2016-03-02 Last updated: 2018-06-07Bibliographically approved
Ekestubbe, S., Bröms, J. E., Edgren, T., Fällman, M., Francis, M. S. & Forsberg, Å. (2016). The amino-terminal part of the needle-tip translocator LcrV of Yersinia pseudotuberculosis is required for early targeting of YopH and in vivo virulence. Frontiers in Cellular and Infection Microbiology, 6, Article ID 175.
Open this publication in new window or tab >>The amino-terminal part of the needle-tip translocator LcrV of Yersinia pseudotuberculosis is required for early targeting of YopH and in vivo virulence
Show others...
2016 (English)In: Frontiers in Cellular and Infection Microbiology, E-ISSN 2235-2988, Vol. 6, article id 175Article in journal (Refereed) Published
Abstract [en]

Type III secretion systems (T3SS) are dedicated to targeting anti-host effector proteins into the cytosol of the host cell to promote bacterial infection. Delivery of the effectors requires three specific translocator proteins, of which the hydrophilic translocator, LcrV, is located at the tip of the T3SS needle and is believed to facilitate insertion of the two hydrophobic translocators into the host cell membrane. Here we used Yersinia as a model to study the role of LcrV in T3SS mediated intracellular effector targeting. Intriguingly, we identified N-terminal IcrV mutants that, similar to the wild-type protein, efficiently promoted expression, secretion and intracellular levels of Yop effectors, yet they were impaired in their ability to inhibit phagocytosis by J774 cells. In line with this, the YopH mediated dephosphorylation of Focal Adhesion Kinase early after infection was compromised when compared to the wild type strain. This suggests that the mutants are unable to promote efficient delivery of effectors to their molecular targets inside the host cell upon host cell contact. The significance of this was borne out by the fact that the mutants were highly attenuated for virulence in the systemic mouse infection model. Our study provides both novel and significant findings that establish a role for LcrV in early targeting of effectors in the host cell.

Keywords
LcrV, type III secretion system, YopH, translocation, pore formation, Yersinia pseudotuberculosis, virulence
National Category
Microbiology in the medical area Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:umu:diva-130149 (URN)10.3389/fcimb.2016.00175 (DOI)000389194200001 ()
Available from: 2017-01-12 Created: 2017-01-12 Last updated: 2018-06-09Bibliographically approved
Amer, A., Gurung, J., Costa, T., Ruuth, K., Zavialov, A., Forsberg, Å. & Francis, M. S. (2016). YopN and TyeA Hydrophobic Contacts Required for Regulating Ysc-Yop Type III Secretion Activity by Yersinia pseudotuberculosis. Frontiers in Cellular and Infection Microbiology, 6, Article ID 66.
Open this publication in new window or tab >>YopN and TyeA Hydrophobic Contacts Required for Regulating Ysc-Yop Type III Secretion Activity by Yersinia pseudotuberculosis
Show others...
2016 (English)In: Frontiers in Cellular and Infection Microbiology, E-ISSN 2235-2988, Vol. 6, article id 66Article in journal (Refereed) Published
Abstract [en]

Yersinia bacteria target Yop effector toxins to the interior of host immune cells by the Ysc-Yop type III secretion system. A YopN-TyeA heterodimer is central to controlling Ysc-Yop targeting activity. A + 1 frameshift event in the 3-prime end of yopN can also produce a singular secreted YopN-TyeA polypeptide that retains some regulatory function even though the C-terminal coding sequence of this YopN differs greatly from wild type. Thus, this YopN C-terminal segment was analyzed for its role in type III secretion control. Bacteria producing YopN truncated after residue 278, or with altered sequence between residues 279 and 287, had lost type III secretion control and function. In contrast, YopN variants with manipulated sequence beyond residue 287 maintained full control and function. Scrutiny of the YopN-TyeA complex structure revealed that residue W279 functioned as a likely hydrophobic contact site with TyeA. Indeed, a YopNW279G mutant lost all ability to bind TyeA. The TyeA residue F8 was also critical for reciprocal YopN binding. Thus, we conclude that specific hydrophobic contacts between opposing YopN and TyeA termini establishes a complex needed for regulating Ysc-Yop activity.

Keywords
protein-protein interaction, molecular modelling, protein secretion, mutagenesis, bacterial pathogenesis, regulation
National Category
Microbiology in the medical area Biochemistry and Molecular Biology
Research subject
Microbiology
Identifiers
urn:nbn:se:umu:diva-122904 (URN)10.3389/fcimb.2016.00066 (DOI)000378543500001 ()
Funder
Swedish Research Council
Available from: 2016-06-23 Created: 2016-06-23 Last updated: 2018-06-07Bibliographically approved
Zwack, E., Snyder, A., Wynosky-Dolfi, M., Ruthel, G., Philip, N., Marketon, M., . . . Brodsky, I. (2015). Inflammasome activation in response to the Yersinia type III secretion system requires hyperinjection of translocon proteins YopB and YopD. mBio, 6(1), Article ID e02095-14.
Open this publication in new window or tab >>Inflammasome activation in response to the Yersinia type III secretion system requires hyperinjection of translocon proteins YopB and YopD
Show others...
2015 (English)In: mBio, ISSN 2161-2129, E-ISSN 2150-7511, Vol. 6, no 1, article id e02095-14Article in journal (Refereed) Published
Abstract [en]

Type III secretion systems (T3SS) translocate effector proteins into target cells in order to disrupt or modulate host cell signaling pathways and establish replicative niches. However, recognition of T3SS activity by cytosolic pattern recognition receptors (PRRs) of the nucleotide-binding domain leucine rich repeat (NLR) family, either through detection of translocated products or membrane disruption, induces assembly of multiprotein complexes known as inflammasomes. Macrophages infected with Yersinia pseudotuberculosis strains lacking all known effectors or lacking the translocation regulator YopK induce rapid activation of both the canonical NLRP3 and noncanonical caspase-11 inflammasomes. While this inflammasome activation requires a functional T3SS, the precise signal that triggers inflammasome activation in response to Yersinia T3SS activity remains unclear. Effectorless strains of Yersinia as well as ΔyopK strains translocate elevated levels of T3SS substrates into infected cells. To dissect the contribution of pore formation and translocation to inflammasome activation, we took advantage of variants of YopD and LcrH that separate these functions of the T3SS. Notably, YopD variants that abrogated translocation but not pore-forming activity failed to induce inflammasome activation. Furthermore, analysis of individual infected cells revealed that inflammasome activation at the single-cell level correlated with translocated levels of YopB and YopD themselves. Intriguingly, LcrH mutants that are fully competent for effector translocation but produce and translocate lower levels of YopB and YopD also fail to trigger inflammasome activation. Our findings therefore suggest that hypertranslocation of YopD and YopB is linked to inflammasome activation in response to the Yersinia T3SS.

National Category
Microbiology
Research subject
Microbiology
Identifiers
urn:nbn:se:umu:diva-101002 (URN)10.1128/mBio.02095-14 (DOI)000350631900065 ()
Funder
Swedish Research Council
Available from: 2015-03-16 Created: 2015-03-16 Last updated: 2018-06-07Bibliographically approved
Li, Y., Hu, Y., Francis, M. S. & Chen, S. (2015). RcsB positively regulates the Yersinia Ysc-Yop type III secretion system by activating expression of the master transcriptional regulator LcrF. Environmental Microbiology, 17(4), 1219-1233
Open this publication in new window or tab >>RcsB positively regulates the Yersinia Ysc-Yop type III secretion system by activating expression of the master transcriptional regulator LcrF
2015 (English)In: Environmental Microbiology, ISSN 1462-2912, E-ISSN 1462-2920, Vol. 17, no 4, p. 1219-1233Article in journal (Refereed) Published
Abstract [en]

The Rcs phosphorelay is a complex signaling pathway used by the family Enterobacteriaceae to sense, respond and adapt to environmental changes during free-living or host-associated lifestyles. In this study, we show that the Rcs phosphorelay pathway positively regulates the virulence plasmid encoded Ysc-Yop type III secretion system (T3SS) in the enteropathogen Yesinia pseudotuberculosis. Both the overexpression of the wild-type Rcs regulator RcsB or the constitutive active RscB(D56E) variant triggered more abundant Ysc-Yop synthesis and secretion, whereas the non-phosphorylatable mutant RcsB(D56Q) negated this. Congruently, enhanced Yops expression and secretion occurred in an in cis rscB(D56E) mutant but not in an isogenic rscB(D56Q) mutant. Screening for regulatory targets of RcsB identified the virG-lcrF operon that encodes for LcrF, the Ysc-Yop T3SS master regulator. Protein-DNA binding assays confirmed that RcsB directly bound to this operon promoter, which subsequently caused stimulated lcrF transcription. Moreover, active RcsB enhanced the ability of bacteria to deliver Yop effectors into immune cells during cell contact, and this promoted an increase in bacterial viability. Taken together, our study demonstrates the role of the Rcs system in regulating the Ysc-Yop T3SS in Yersinia and reports on RcsB being the first transcriptional activator known to directly control lcrF transcription.

National Category
Microbiology in the medical area
Identifiers
urn:nbn:se:umu:diva-103539 (URN)10.1111/1462-2920.12556 (DOI)000352545100023 ()25039908 (PubMedID)
Available from: 2015-05-25 Created: 2015-05-21 Last updated: 2018-06-07Bibliographically approved
Li, Y., Hu, Y., Francis, M. & Chen, S. (2015). RcsB positively regulates the Yersinia Ysc-Yop type III secretion system by activating expression of the master transcriptional regulator LcrF. Environmental Microbiology, 17(4), 1219-1233
Open this publication in new window or tab >>RcsB positively regulates the Yersinia Ysc-Yop type III secretion system by activating expression of the master transcriptional regulator LcrF
2015 (English)In: Environmental Microbiology, ISSN 1462-2912, E-ISSN 1462-2920, Vol. 17, no 4, p. 1219-1233Article in journal (Refereed) Published
Abstract [en]

The Rcs phosphorelay is a complex signaling pathway used by the family Enterobacteriaceae to sense, respond and adapt to environmental changes during free-living or host-associated lifestyles. In this study, we show that the Rcs phosphorelay pathway positively regulates the virulence plasmid encoded Ysc-Yop type III secretion system (T3SS) in the enteropathogen Yesinia pseudotuberculosis. Both the overexpression of the wild-type Rcs regulator RcsB or the constitutive active RscB(D56E) variant triggered more abundant Ysc-Yop synthesis and secretion, whereas the non-phosphorylatable mutant RcsB(D56Q) negated this. Congruently, enhanced Yops expression and secretion occurred in an in cis rscB(D56E) mutant but not in an isogenic rscB(D56Q) mutant. Screening for regulatory targets of RcsB identified the virG-lcrF operon that encodes for LcrF, the Ysc-Yop T3SS master regulator. Protein-DNA binding assays confirmed that RcsB directly bound to this operon promoter, which subsequently caused stimulated lcrF transcription. Moreover, active RcsB enhanced the ability of bacteria to deliver Yop effectors into immune cells during cell contact, and this promoted an increase in bacterial viability. Taken together, our study demonstrates the role of the Rcs system in regulating the Ysc-Yop T3SS in Yersinia and reports on RcsB being the first transcriptional activator known to directly control lcrF transcription.

Place, publisher, year, edition, pages
John Wiley & Sons, 2015
Keywords
Rcs phosphorelay; type III secretion, transcriptional regulator, protein-DNA binding, Yersinia
National Category
Microbiology
Research subject
Molecular Biology
Identifiers
urn:nbn:se:umu:diva-102412 (URN)10.1111/1462-2920.12556 (DOI)
Funder
Swedish Research Council, 2014-6652Swedish Research Council, 2014-2105
Available from: 2015-04-24 Created: 2015-04-24 Last updated: 2018-06-07Bibliographically approved
Lu, P., Zhang, Y., Hu, Y., Francis, M. & Chen, S. (2014). A cis-encoded sRNA controls the expression of fabH2 in Yersinia. FEBS Letters, 588(10), 1961-1966
Open this publication in new window or tab >>A cis-encoded sRNA controls the expression of fabH2 in Yersinia
Show others...
2014 (English)In: FEBS Letters, ISSN 0014-5793, E-ISSN 1873-3468, Vol. 588, no 10, p. 1961-1966Article in journal (Refereed) Published
Abstract [en]

YsrH is a novel cis-encoded sRNA located on the opposite strand to fabH2, which is essential for fatty acid biosynthesis in bacteria. In this study, YsrH-mediated regulation of fabH2 expression was investigated in Yersinia pseudotuberculosis. Constitutive and inducible over-expression of YsrH decreased the mRNA level of fabH2, while expression of downstream fabD and fabG remained unaffected. Polynucleotide phosphorylase (PNPase) also played an important role in this regulation process by mediating YsrH decay in the exponential phase. Thus, our data defines a cis-encoded sRNA that regulates fatty acid synthesis via a regulatory mechanism also involving PNPase.

Keywords
3-oxoacyl-ACP synthase (fabH2); Yersinia sRNA regulator of fabH2 (YsrH); cis-Encoded sRNA; PNPase; Yersinia
National Category
Microbiology Biochemistry and Molecular Biology
Research subject
Microbiology; Molecular Biology
Identifiers
urn:nbn:se:umu:diva-88605 (URN)10.1016/j.febslet.2014.04.005 (DOI)000336033700016 ()24735725 (PubMedID)
Available from: 2014-05-09 Created: 2014-05-09 Last updated: 2018-06-07Bibliographically approved
Li, Y., Li, L., Huang, L., Francis, M., Hu, Y. & Chen, S. (2014). Yersinia Ysc-Yop type III secretion feedback inhibition is relieved through YscV-dependent recognition and secretion of LcrQ. Molecular Microbiology, 91(3), 494-507
Open this publication in new window or tab >>Yersinia Ysc-Yop type III secretion feedback inhibition is relieved through YscV-dependent recognition and secretion of LcrQ
Show others...
2014 (English)In: Molecular Microbiology, ISSN 0950-382X, E-ISSN 1365-2958, Vol. 91, no 3, p. 494-507Article in journal (Refereed) Published
Abstract [en]

Human pathogenic Yersinia species share a virulence plasmid encoding the Ysc-Yop type III secretion system (T3SS). A plasmid-encoded anti-activator, LcrQ, negatively regulates the expression of this secretion system. Under inducible conditions, LcrQ is secreted outside of bacterial cells and this activates the T3SS, but the mechanism of targeting LcrQ for type III secretion remains largely unknown. In this study, we characterized the regulatory role of the export apparatus component YscV. Depletion or overexpression of YscV compromised Yop synthesis and this primarily prevented secretion of LcrQ. It followed that a lcrQ deletion reversed the repressive effects of excessive YscV. Further characterization demonstrated that the YscV residues 493–511 located within the C-terminal soluble cytoplasmic domain directly bound with LcrQ. Critically, YscV-LcrQ complex formation was a requirement for LcrQ secretion, since YscVΔ493–511 failed to secrete LcrQ. This forced a cytoplasmic accumulation of LcrQ, which predictably caused the feedback inhibition of Yops synthesis. Based on these observations, we proposed a model for the YscV-dependent secretion of LcrQ and its role in regulating Yop synthesis in Yersinia.

Place, publisher, year, edition, pages
John Wiley & Sons, 2014
Keywords
T3SS, YscV, LcrQ, Yersinia, Export apparatus
National Category
Microbiology Microbiology in the medical area Biochemistry and Molecular Biology Cell and Molecular Biology
Research subject
Molecular Biology; Microbiology
Identifiers
urn:nbn:se:umu:diva-85894 (URN)10.1111/mmi.12474 (DOI)000345498000005 ()2-s2.0-84890320543 (Scopus ID)
Available from: 2014-02-12 Created: 2014-02-12 Last updated: 2018-06-08Bibliographically approved
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0001-6817-9535

Search in DiVA

Show all publications