YopK of <i>Yersinia pseudotuberculosis </i>controls translocation of Yop effectors across the eukaryotic cell membrane

Holmström, Anna; Petterson, Jonas; Rosqvist, Roland; Håkansson, Sebastian; Tafazoli, Farideh; Fällman, Maria; Magnusson, Karl‐Eric; Wolf‐Watz, Hans; Försberg, Åke

doi:10.1046/j.1365-2958.1997.3211681.x

Cited by 127 publications

(172 citation statements)

References 59 publications

(111 reference statements)

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“…Contact-dependent lysis of sheep erythrocytes is used as a measure of translocon pore formation, which is quantified by determining the release of hemoglobin into the cleared supernatant of infected erythrocytes. In Yersinia infections, assay sensitivity is improved by using bacteria devoid of YopK, a regulator of pore formation (48). Therefore, our yopD mutations were first introduced into the (A and B); CD-like, moderate calcium-dependent growth (C); TS, bacteria that are sensitive to elevated temperature regardless of the presence of calcium (E and F); TS-like, minimal growth observed in the presence of calcium (D).…”

Section: Resultsmentioning

confidence: 99%

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YopD Self-assembly and Binding to LcrV Facilitate Type III Secretion Activity by Yersinia pseudotuberculosis

Costa

Edqvist

Bröms

et al. 2010

Journal of Biological Chemistry

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YopD-like translocator proteins encoded by several Gramnegative bacteria are important for type III secretion-dependent delivery of anti-host effectors into eukaryotic cells. This probably depends on their ability to form pores in the infected cell plasma membrane, through which effectors may gain access to the cell interior. In addition, Yersinia YopD is a negative regulator essential for the control of effector synthesis and secretion. As a prerequisite for this functional duality, YopD may need to establish molecular interactions with other key T3S components. A putative coiled-coil domain and an ␣-helical amphipathic domain, both situated in the YopD C terminus, may represent key protein-protein interaction domains. Therefore, residues within the YopD C terminus were systematically mutagenized. All 68 mutant bacteria were first screened in a variety of assays designed to identify individual residues essential for YopD function, possibly by providing the interaction interface for the docking of other T3S proteins. Mirroring the effect of a full-length yopD gene deletion, five mutant bacteria were defective for both yop regulatory control and effector delivery. Interestingly, all mutations clustered to hydrophobic amino acids of the amphipathic domain. Also situated within this domain, two additional mutants rendered YopD primarily defective in the control of Yop synthesis and secretion. Significantly, protein-protein interaction studies revealed that functionally compromised YopD variants were also defective in self-oligomerization and in the ability to engage another translocator protein, LcrV. Thus, the YopD amphipathic domain facilitates the formation of YopD/YopD and YopD/LcrV interactions, two critical events in the type III secretion process.

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Section: Resultsmentioning

confidence: 99%

“…The lower limit was defined by infections with isogenic Yersinia producing a truncated YopD lacking the 278 -292-residue amphipathic domain (YPIII/pIB622). 15, and dextran 4 were performed as described previously (6,11,17,48).…”

Section: Methodsmentioning

confidence: 99%

YopD Self-assembly and Binding to LcrV Facilitate Type III Secretion Activity by Yersinia pseudotuberculosis

Costa

Edqvist

Bröms

et al. 2010

Journal of Biological Chemistry

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“…In Yersinia, two secreted T3SS substrates, YopB and YopD, have been shown to be involved in the translocation of effector proteins into the target cell and these proteins are defined as translocators to distinguish them from the effector class of Yops that have a more direct effect on eukaryotic cell function (3). Both YopB and YopD have hydrophobic domains (10) and can be inserted into the membranes of erythrocytes and nucleated cells, where they form pores (11,12). It is assumed that binding of Yersinia to a target cell triggers secretion and insertion of these proteins into the target-cell plasma membrane, completing the conduit between the bacterium and the target cell through the T3SS needle complex.…”

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confidence: 99%

Translocation of surface-localized effectors in type III secretion

Akopyan

Edgren

Wang-Edgren

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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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.

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“…YopK is a translocated effector that regulates the translocation of other Yersinia Yops (42,84). YopK deficiency results in significantly attenuated infection in vivo, which was initially surprising given that YopK-deficient Yersinia translocate an increased amount of virulence factors into target cells (36,76,85,86).…”

Section: Discussionmentioning

confidence: 99%

Guanylate Binding Proteins Regulate Inflammasome Activation in Response to Hyperinjected Yersinia Translocon Components

et al. 2017

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Gram-negative bacterial pathogens utilize virulence-associated secretion systems to inject, or translocate, effector proteins into host cells to manipulate cellular processes and promote bacterial replication. However, translocated bacterial products are sensed by nucleotide binding domain and leucine-rich repeat-containing proteins (NLRs), which trigger the formation of a multiprotein complex called the inflammasome, leading to secretion of interleukin-1 (IL-1) family cytokines, pyroptosis, and control of pathogen replication. Pathogenic Yersinia bacteria inject effector proteins termed Yops, as well as pore-forming proteins that comprise the translocon itself, into target cells. The Yersinia translocation regulatory protein YopK promotes bacterial virulence by limiting hyperinjection of the translocon proteins YopD and YopB into cells, thereby limiting cellular detection of Yersinia virulence activity. How hyperinjection of translocon proteins leads to inflammasome activation is currently unknown. We found that translocated YopB and YopD colocalized with the late endosomal/lysosomal protein LAMP1 and that the frequency of YopD and LAMP1 association correlated with the level of caspase-1 activation in individual cells. We also observed colocalization between YopD and Galectin-3, an indicator of endosomal membrane damage. Intriguingly, YopK limited the colocalization of Galectin-3 with YopD, suggesting that YopK limits the induction or sensing of endosomal membrane damage by components of the type III secretion system (T3SS) translocon. Furthermore, guanylate binding proteins (GBPs) encoded on chromosome 3 (Gbp Chr3 ), which respond to pathogen-induced damage or alteration of host membranes, were necessary for inflammasome activation in response to hyperinjected YopB/-D. Our findings indicate that lysosomal damage by Yersinia translocon proteins promotes inflammasome activation and implicate GBPs as key regulators of this process.

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YopK of Yersinia pseudotuberculosis controls translocation of Yop effectors across the eukaryotic cell membrane

Cited by 127 publications

References 59 publications

YopD Self-assembly and Binding to LcrV Facilitate Type III Secretion Activity by Yersinia pseudotuberculosis

YopD Self-assembly and Binding to LcrV Facilitate Type III Secretion Activity by Yersinia pseudotuberculosis

Translocation of surface-localized effectors in type III secretion

Guanylate Binding Proteins Regulate Inflammasome Activation in Response to Hyperinjected Yersinia Translocon Components

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