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Translocation of surface-localized effectors in type III secretion.
Proc Natl Acad Sci U S A. 2011 Jan 25; 108(4):1639-44
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20 Jan 2011
DOI: 10.3410/f.7778956.8115054
Type III secretion systems (T3SS) are assumed to deliver substrates directly from the bacterial cytoplasm into target host cells in a single step, with no extracellular intermediate. Surprisingly, Akopyan et al. demonstrate that certain T3SS effectors from Yersinia are exposed on the bacterial surfa and...Continue reading
Type III secretion systems (T3SS) are assumed to deliver substrates directly from the bacterial cytoplasm into target host cells in a single step, with no extracellular intermediate. Surprisingly, Akopyan et al. demonstrate that certain T3SS effectors from Yersinia are exposed on the bacterial surface, and that they can be translocated from here into target eukaryotic cells.
Type III secretion systems are needle-like macromolecular structures that are thought to form a continuous conduit from bacteria to eukaryotic host cells, and to 'inject' bacterial effector proteins directly into target cells. Contrary to this model, Akopyan et al. show that YopH, a T3SS substrate from Yersinia, can be localised on the bacterial surface. They show that this exposed YopH can still be translocated into eukaryotic cells by the T3SS 'translocon' components, proteins which have been proposed to form a pore structure at the tip of the T3SS needle. Exogenous purified YopH could also be translocated into target cells when added to the surface of either a yopH-mutant strain of Yersinia, or Salmonella.
This work suggests that our understanding of type III secretion, and especially the function of the 'translocon', may have to be significantly revised.
Type III secretion systems are needle-like macromolecular structures that are thought to form a continuous conduit from bacteria to eukaryotic host cells, and to 'inject' bacterial effector proteins directly into target cells. Contrary to this model, Akopyan et al. show that YopH, a T3SS substrate from Yersinia, can be localised on the bacterial surface. They show that this exposed YopH can still be translocated into eukaryotic cells by the T3SS 'translocon' components, proteins which have been proposed to form a pore structure at the tip of the T3SS needle. Exogenous purified YopH could also be translocated into target cells when added to the surface of either a yopH-mutant strain of Yersinia, or Salmonella.
This work suggests that our understanding of type III secretion, and especially the function of the 'translocon', may have to be significantly revised.
Disclosures
None declared
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Koronakis V and Hume P: F1000Prime Recommendation of [Akopyan K et al., Proc Natl Acad Sci U S A 2011, 108(4):1639-44]. In F1000Prime, 20 Jan 2011; DOI: 10.3410/f.7778956.8115054. F1000Prime.com/7778956#eval8115054
10 Feb 2011
DOI: 10.3410/f.7778956.8385054
This paper supports a new and distinctive mechanism by which bacterial proteins are translocated through the type III secretion system (T3SS). The novelty lies in the fact that Yersinia pseudotuberculosis T3SS effectors such as YopD, YopE, and YopH are found uniformly distributed on the surface of bacteria...Continue reading
This paper supports a new and distinctive mechanism by which bacterial proteins are translocated through the type III secretion system (T3SS). The novelty lies in the fact that Yersinia pseudotuberculosis T3SS effectors such as YopD, YopE, and YopH are found uniformly distributed on the surface of bacteria and from there they are translocated to the cytoplasm of the host cell in a T3SS-dependent manner. The authors further show that Yersinia strains coated with purified YopH in vitro are also able to translocate this protein through the T3SS.
The results shown in this paper suggest that proteins translocated through the T3SS can transition through a membrane bound state. Indeed, careful electron microscopy and biochemical analyses show that effectors are first addressed to the surface. Interestingly, deletion of different domains reveals that secretion and translocation can be uncoupled. This two-step model of translocation is very different from the textbook view of a one-step direct transfer of T3SS effectors from the cytoplasm of the bacterium to the cytoplasm of the host cell. Importantly though, the authors, who were the first to demonstrate the 1-step transfer of Yersinia proteins {1}, stress that the two models are not mutually exclusive.
After such a study and a novel hypothesis is published, many questions remain, such as how are the proteins secreted? What is the contribution of the one-step versus the two-step model? How widespread is this new mechanism among bacteria that have a T3SS? Etc.
The results shown in this paper suggest that proteins translocated through the T3SS can transition through a membrane bound state. Indeed, careful electron microscopy and biochemical analyses show that effectors are first addressed to the surface. Interestingly, deletion of different domains reveals that secretion and translocation can be uncoupled. This two-step model of translocation is very different from the textbook view of a one-step direct transfer of T3SS effectors from the cytoplasm of the bacterium to the cytoplasm of the host cell. Importantly though, the authors, who were the first to demonstrate the 1-step transfer of Yersinia proteins {1}, stress that the two models are not mutually exclusive.
After such a study and a novel hypothesis is published, many questions remain, such as how are the proteins secreted? What is the contribution of the one-step versus the two-step model? How widespread is this new mechanism among bacteria that have a T3SS? Etc.
References
1.
Target cell contact triggers expression and polarized transfer of Yersinia YopE cytotoxin into mammalian cells.
EMBO J 1994 Feb 15; 4(13):964-72
PMID: 8112310
Disclosures
None declared
Cossart P and Hamon M: F1000Prime Recommendation of [Akopyan K et al., Proc Natl Acad Sci U S A 2011, 108(4):1639-44]. In F1000Prime, 10 Feb 2011; DOI: 10.3410/f.7778956.8385054. F1000Prime.com/7778956#eval8385054
14 Feb 2011
Olivia Steele-Mortimer
F1000 Microbiology
Rocky Mountain Laboratories, National Institute of Allergy and Infectious Disease, NIH,
Hamilton , MT, USA.
F1000 Microbiology
Rocky Mountain Laboratories, National Institute of Allergy and Infectious Disease, NIH,
Hamilton , MT, USA.
Carrie Jolly
F1000 Microbiology
Rocky Mountain Laboratories, National Institute of Allergy and Infectious Disease, NIH,
Hamilton, MT, USA.
F1000 Microbiology
Rocky Mountain Laboratories, National Institute of Allergy and Infectious Disease, NIH,
Hamilton, MT, USA.
DOI: 10.3410/f.7778956.8729055
How do type III secretion systems (T3SSs) deliver effector proteins to the target host cell? T3SSs are typically depicted as static structures with the needle complex 'docked' to the translocation pore (i.e. translocon), thus producing a stationary channel through which the effectors are delivered from...Continue reading
How do type III secretion systems (T3SSs) deliver effector proteins to the target host cell? T3SSs are typically depicted as static structures with the needle complex 'docked' to the translocation pore (i.e. translocon), thus producing a stationary channel through which the effectors are delivered from the bacterial cytosol directly into the host cell cytoplasm. In this report, Akopyan et al. demonstrate the surprising find that some Yersinia T3SS effectors are located on the surface of the bacteria prior to host cell contact and that these effectors can be delivered via the T3SS translocon to the target host cell cytoplasm.
Many Gram negative bacterial pathogens use a conserved injection machine, the type III secretion system (T3SS) to translocate bacterial effector proteins into the cytoplasm of host cells. These effectors manipulate the host cellular or immune processes to allow the bacteria to colonize their host, establish their explicative niche, evade immune detection, etc. Significant progress has been made in our understanding of the regulation, assembly, and structure of the T3SS. However, we still lack a complete understanding of how the injection apparatus functions to deliver the effectors into the target host cell. Current models depict the T3SS as an apparatus that directly injects effectors from the bacterial cytosol to the host cell cytoplasm. In this report, the authors provide evidence for an alternative mechanism that involves transfer of effectors located on the surface of the bacteria through the translocon and into the host cell.
Using immunoelectron microscopy, the authors were able to demonstrate that several Yersinia T3SS effectors (Yops) were located on the surface of the bacteria either prior to or during host cell contact. In addition, these effectors located on the surface did not colocalize with components of the T3SS. The surface-localized effector, YopH was shown to be translocated into the host cell cytoplasm via a mechanism that requires the T3SS translocon. Surprisingly, recombinant YopH coated on the surface of the deltayopH strain of Yersinia was also translocated via the T3SS-dependent mechanism. The authors also demonstrate that the T3SS of Salmonella can be used as a surrogate system to translocate YopH coated on the surface of these bacteria.
Future studies should be conducted to further characterize the importance of the translocon in this novel delivery mechanism of surface-localized effectors. For example, could a pore formed from a different source substitute for the translocon and allow for the delivery of these surface-localized effectors to the host cell cytoplasm? Overall, this study represents an intriguing find and is a fantastic illustration of how much more we need to learn regarding the T3SS.
Many Gram negative bacterial pathogens use a conserved injection machine, the type III secretion system (T3SS) to translocate bacterial effector proteins into the cytoplasm of host cells. These effectors manipulate the host cellular or immune processes to allow the bacteria to colonize their host, establish their explicative niche, evade immune detection, etc. Significant progress has been made in our understanding of the regulation, assembly, and structure of the T3SS. However, we still lack a complete understanding of how the injection apparatus functions to deliver the effectors into the target host cell. Current models depict the T3SS as an apparatus that directly injects effectors from the bacterial cytosol to the host cell cytoplasm. In this report, the authors provide evidence for an alternative mechanism that involves transfer of effectors located on the surface of the bacteria through the translocon and into the host cell.
Using immunoelectron microscopy, the authors were able to demonstrate that several Yersinia T3SS effectors (Yops) were located on the surface of the bacteria either prior to or during host cell contact. In addition, these effectors located on the surface did not colocalize with components of the T3SS. The surface-localized effector, YopH was shown to be translocated into the host cell cytoplasm via a mechanism that requires the T3SS translocon. Surprisingly, recombinant YopH coated on the surface of the deltayopH strain of Yersinia was also translocated via the T3SS-dependent mechanism. The authors also demonstrate that the T3SS of Salmonella can be used as a surrogate system to translocate YopH coated on the surface of these bacteria.
Future studies should be conducted to further characterize the importance of the translocon in this novel delivery mechanism of surface-localized effectors. For example, could a pore formed from a different source substitute for the translocon and allow for the delivery of these surface-localized effectors to the host cell cytoplasm? Overall, this study represents an intriguing find and is a fantastic illustration of how much more we need to learn regarding the T3SS.
Disclosures
None declared
Steele-Mortimer O and Jolly C: F1000Prime Recommendation of [Akopyan K et al., Proc Natl Acad Sci U S A 2011, 108(4):1639-44]. In F1000Prime, 14 Feb 2011; DOI: 10.3410/f.7778956.8729055. F1000Prime.com/7778956#eval8729055
18 Feb 2011
DOI: 10.3410/f.7778956.8798056
This controversial paper challenges the long-held belief that type III secretion (T3S) apparatus provides a continuous conduit between the bacterial and host cytoplasm.
In a series of elegant experiments, this group shows that Yersinia effector proteins, present on the cell surface of the bacteria, will...Continue reading
In a series of elegant experiments, this group shows that Yersinia effector proteins, present on the cell surface of the bacteria, will...Continue reading
This controversial paper challenges the long-held belief that type III secretion (T3S) apparatus provides a continuous conduit between the bacterial and host cytoplasm.
In a series of elegant experiments, this group shows that Yersinia effector proteins, present on the cell surface of the bacteria, will translocate into the host cytoplasm via the translocator proteins YopB and YopD. The signal for translocation across the host membrane is apparently distinct from the one required for export by the T3S apparatus, or binding to secretion chaperone in the bacterial cytoplasm. This 'translocation signal' is likely conserved among T3S effectors, as Salmonella will translocate recombinant Yersinia effectors that are coated on the surface of Salmonella. These findings may lead us to reassess how we think effectors are translocated into cells, how translocator proteins decipher signals on effectors, and raises the possibility that secreted proteins without classical T3S signals may use the translocator pores to gain access to the host cytoplasm.
In a series of elegant experiments, this group shows that Yersinia effector proteins, present on the cell surface of the bacteria, will translocate into the host cytoplasm via the translocator proteins YopB and YopD. The signal for translocation across the host membrane is apparently distinct from the one required for export by the T3S apparatus, or binding to secretion chaperone in the bacterial cytoplasm. This 'translocation signal' is likely conserved among T3S effectors, as Salmonella will translocate recombinant Yersinia effectors that are coated on the surface of Salmonella. These findings may lead us to reassess how we think effectors are translocated into cells, how translocator proteins decipher signals on effectors, and raises the possibility that secreted proteins without classical T3S signals may use the translocator pores to gain access to the host cytoplasm.
Disclosures
None declared
Valdivia R: F1000Prime Recommendation of [Akopyan K et al., Proc Natl Acad Sci U S A 2011, 108(4):1639-44]. In F1000Prime, 18 Feb 2011; DOI: 10.3410/f.7778956.8798056. F1000Prime.com/7778956#eval8798056
18 Mar 2011
DOI: 10.3410/f.7778956.9639055
This paper shows that effector proteins located on the bacterial surface can be translocated through type III secretion systems into the host cell. These observations challenge the established single-step model, in which the effector molecule goes directly from the bacterial cytosol to the host cell....Continue reading
This paper shows that effector proteins located on the bacterial surface can be translocated through type III secretion systems into the host cell. These observations challenge the established single-step model, in which the effector molecule goes directly from the bacterial cytosol to the host cell. Understanding the functional mechanisms of secretion systems is essential for the development of new drugs and vaccines that target them.
The authors show that translocation of the effector protein YopH through the type III secretion systems of Yersinia pseudotuberculosis and Salmonella typhimurium does not necessarily occur from the bacterial cytosol to the host cell as has always been thought. Yop effectors have been shown to be transferred into the host cell cytoplasm, but exactly how this process happens is not known {1}. The same is true for other secretion systems such as the translocation of the Helicobacter pylori CagA protein through the type IV secretion system {2}. Given the importance of secretion systems in the virulence of several bacterial species, dissection of their functional mechanisms is critical for the understanding of pathogenic mechanisms as well as the development of new therapeutic approaches. Compounds targeting secretion systems have already been investigated and shown to be effective {3}. The authors of the present paper suggest that effector proteins located on the bacterial surface can be successfully translocated by type III apparatuses. These observations may instruct the design of new inhibitory molecules able to block the translocation of the effector molecules as well new vaccines. Indeed, if the effectors molecules reside on the bacterial surface prior to their translocation, they may be exploited as vaccine targets.
The authors show that translocation of the effector protein YopH through the type III secretion systems of Yersinia pseudotuberculosis and Salmonella typhimurium does not necessarily occur from the bacterial cytosol to the host cell as has always been thought. Yop effectors have been shown to be transferred into the host cell cytoplasm, but exactly how this process happens is not known {1}. The same is true for other secretion systems such as the translocation of the Helicobacter pylori CagA protein through the type IV secretion system {2}. Given the importance of secretion systems in the virulence of several bacterial species, dissection of their functional mechanisms is critical for the understanding of pathogenic mechanisms as well as the development of new therapeutic approaches. Compounds targeting secretion systems have already been investigated and shown to be effective {3}. The authors of the present paper suggest that effector proteins located on the bacterial surface can be successfully translocated by type III apparatuses. These observations may instruct the design of new inhibitory molecules able to block the translocation of the effector molecules as well new vaccines. Indeed, if the effectors molecules reside on the bacterial surface prior to their translocation, they may be exploited as vaccine targets.
References
1.
The YopB protein of Yersinia pseudotuberculosis is essential for the translocation of Yop effector proteins across the target cell plasma membrane and displays a contact-dependent membrane disrupting activity.
EMBO J 1996 Nov 1; 21(15):5812-23
PMID: 8918459
2.
Tyrosine phosphorylation of the Helicobacter pylori CagA antigen after cag-driven host cell translocation.
Proc Natl Acad Sci USA 2000 Feb 1; 3(97):1263-8
PMID: 10655519
3.
Inhibitors of Helicobacter pylori ATPase Cagalpha block CagA transport and cag virulence.
Microbiology (Reading, Engl) 2006 Oct; Pt 10(152):2919-30
PMID: 17005973
Disclosures
None declared
Rappuoli R and Bagnoli F: F1000Prime Recommendation of [Akopyan K et al., Proc Natl Acad Sci U S A 2011, 108(4):1639-44]. In F1000Prime, 18 Mar 2011; DOI: 10.3410/f.7778956.9639055. F1000Prime.com/7778956#eval9639055
25 Mar 2011
DOI: 10.3410/f.7778956.9813069
This paper is exceptional because it challenges the currently accepted and dogmatic 'conduit model', which claims that several pathogenic bacteria use type III secretion systems (T3SS) to translocate effector proteins from the bacterial cytosol into the host cell through a continuous channel built up...Continue reading
This paper is exceptional because it challenges the currently accepted and dogmatic 'conduit model', which claims that several pathogenic bacteria use type III secretion systems (T3SS) to translocate effector proteins from the bacterial cytosol into the host cell through a continuous channel built up by the T3SS upon bacterium-host cell contact.
Many pathogenic but also commensal bacteria use dedicated multi-protein nanomachines, such as type III or type IV secretion systems (T3SS or T4SS), to deliver bacterial effector proteins into the host cell. It is believed that the effectors are delivered in a single step from the bacterial cytosol into the host cell through a continuous channel created by the needle component and basal structure of T3SS/T4SS. This recent paper from the Fallman lab on Yersinia pseudotuberculosis T3SS seriously challenges this view but, at the same time, includes a set of extremely convincing experiments, which have to be taken into account in the field.
The authors demonstrate that the YopH-deletion mutant of Y. pseudotuberculosis is able to translocate recombinant YopH into a target cell in a T3SS-dependent manner in an experimental set-up where bacteria are coated with the recombinant YopH. The authors further report by extensive deletion mapping that the well-described N-terminal secretion signal sequence of YopH is actually composed of two domains: one domain that directs YopH to be secreted into extracellular space and the other domain that is required for T3SS-dependent translocation of the extracellular YopH into the target cell. The authors are also able to replicate their results by using YopH-coated and T3SS-positive Salmonella typhimurium, which implies that the new effector translocation mechanism might be widespread among bacteria that express T3SS.
What is the purpose of the needle component of T3SS? Perhaps it only inserts the hydrophobic translocator proteins, which form the actual pore and entry site for the effector proteins, into the host cell membrane. However, we can also envision a scenario where the dogmatic 'conduit model' would still selectively apply to translocate highly hydrophobic effectors into the host cell. Indeed, the authors carefully argue that their new model of effector translocation does not necessarily exclude the dogmatic 'conduit model'.
Further studies should be conducted to verify and characterize the fascinating findings of the Fallman lab, possibly also in other secretion settings such as in the evolutionary distinct T4SS. Also, the report of the Fallman lab makes us wonder about the implications of this new secretion model in the dynamics of natural/pathogenic microbial flora of the host organism. Could some bacterial species, strains or even sub-populations devote themselves to effector-secretors, exclude themselves from the costly needle and translocase expression and highjack the translocator pores that are formed by neighbouring bacteria?
Many pathogenic but also commensal bacteria use dedicated multi-protein nanomachines, such as type III or type IV secretion systems (T3SS or T4SS), to deliver bacterial effector proteins into the host cell. It is believed that the effectors are delivered in a single step from the bacterial cytosol into the host cell through a continuous channel created by the needle component and basal structure of T3SS/T4SS. This recent paper from the Fallman lab on Yersinia pseudotuberculosis T3SS seriously challenges this view but, at the same time, includes a set of extremely convincing experiments, which have to be taken into account in the field.
The authors demonstrate that the YopH-deletion mutant of Y. pseudotuberculosis is able to translocate recombinant YopH into a target cell in a T3SS-dependent manner in an experimental set-up where bacteria are coated with the recombinant YopH. The authors further report by extensive deletion mapping that the well-described N-terminal secretion signal sequence of YopH is actually composed of two domains: one domain that directs YopH to be secreted into extracellular space and the other domain that is required for T3SS-dependent translocation of the extracellular YopH into the target cell. The authors are also able to replicate their results by using YopH-coated and T3SS-positive Salmonella typhimurium, which implies that the new effector translocation mechanism might be widespread among bacteria that express T3SS.
What is the purpose of the needle component of T3SS? Perhaps it only inserts the hydrophobic translocator proteins, which form the actual pore and entry site for the effector proteins, into the host cell membrane. However, we can also envision a scenario where the dogmatic 'conduit model' would still selectively apply to translocate highly hydrophobic effectors into the host cell. Indeed, the authors carefully argue that their new model of effector translocation does not necessarily exclude the dogmatic 'conduit model'.
Further studies should be conducted to verify and characterize the fascinating findings of the Fallman lab, possibly also in other secretion settings such as in the evolutionary distinct T4SS. Also, the report of the Fallman lab makes us wonder about the implications of this new secretion model in the dynamics of natural/pathogenic microbial flora of the host organism. Could some bacterial species, strains or even sub-populations devote themselves to effector-secretors, exclude themselves from the costly needle and translocase expression and highjack the translocator pores that are formed by neighbouring bacteria?
Disclosures
None declared
Dehio C and Pulliainen A: F1000Prime Recommendation of [Akopyan K et al., Proc Natl Acad Sci U S A 2011, 108(4):1639-44]. In F1000Prime, 25 Mar 2011; DOI: 10.3410/f.7778956.9813069. F1000Prime.com/7778956#eval9813069
F1000Prime Recommendations, Dissents and Comments for [Akopyan K et al., Proc Natl Acad Sci U S A 2011, 108(4):1639-44]. In F1000Prime, 24 May 2013; F1000Prime.com/7778956
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Pathogenic Yersinia species suppress the host immune response by using a plasmid-encoded type III secretion system (T3SS) to translocate virulence proteins into the cytosol of the target cells. T3SS-dependent protein translocation is believed to occur in one step from the bacterial cytosol to the target-cell cytoplasm through a conduit created by the T3SS upon target cell contact. Here, we report that T3SS substrates on the surface of Yersinia pseudotuberculosis are translocated into target cells. Upon host cell contact, purified YopH coated on Y. pseudotuberculosis was specifically and rapidly translocated across the target-cell membrane, which led to a physiological response in the infected cell. In addition, translocation of externally added YopH required a functional T3SS and a specific translocation domain in the effector protein. Efficient, T3SS-dependent translocation of purified YopH added in vitro was also observed when using coated Salmonella typhimurium strains, which implies that T3SS-mediated translocation of extracellular effector proteins is conserved among T3SS-dependent pathogens. Our results demonstrate that polarized T3SS-dependent translocation of proteins can be achieved through an intermediate extracellular step that can be reconstituted in vitro. These results indicate that translocation can occur by a different mechanism from the assumed single-step conduit model.
PMID: 21220342
