Biology subjects

Coers, J.

Publications and source records attributed to Coers, J..

15 recordsLinked to original sources

bioRxiv2026

Toxoplasma gondii GRA66 prevents premature egress driven by the host phospholipase RARRES3, independently of RNF213

Toxoplasma gondii replicates inside a host-derived parasitophorous vacuole (PV), and interferon-gamma (IFN{gamma}) induces host restriction factors that target this compartment. Previous CRISPR screens identified the dense granule protein GRA66 and the GRA57/GRA70/GRA71 complex as required for parasite fitness in IFN{gamma}-stimulated human cells, but the host pathways they oppose were unknown. Here, we show that GRA66, a PV membrane (PVM)-associated protein predicted to be an N-acylphosphatidylethanolamine (NAPE)-hydrolyzing phospholipase D, is required to prevent premature egress driven by the host phospholipase and acyltransferase RARRES3. Loss of GRA66 caused premature parasite egress, host cell death, and impaired replication in human cells. These phenotypes persisted in cells lacking RNF213, the E3 ubiquitin ligase that dominates IFN{gamma}-dependent Toxoplasma restriction in human cells, and{Delta} gra66 vacuoles recruited less RNF213 and ubiquitin than wild type, indicating an RNF213-independent mechanism rather than an exaggerated RNF213 response. Complementation with a catalytic-site mutant of the GRA66 zinc-binding motif failed to restore any of these phenotypes, indicating a requirement for its predicted enzymatic activity. Deleting RARRES3 rescued the premature egress of {Delta}gra66 and catalytic-mutant parasites but not of{Delta} gra70 parasites, whereas the replication defect persisted, revealing a second, RARRES3-independent consequence of GRA66 loss. RARRES3 was recruited to the PVM and intravacuolar network after IFN{gamma} stimulation, and structural modeling supported assignment of GRA66 to the NAPE-phospholipase D family with an intact di-zinc active site. These findings define a lipid-centered, RNF213-independent arm of human cell-autonomous immunity and identify the parasite effector required to withstand it.

microbiology↗

bioRxiv2026

Intracellular pathogen targeting by IL32 elicits cell-autonomous immunity

Interferon-{gamma} safeguards humans against intracellular pathogens, yet how most interferon-stimulated genes protect host cells, and how human-adapted pathogens evade these defenses is unclear1,2. Here, we discover a potent immune surveillance and effector circuit executed by an intracellularly acting cytokine, IL32, that targets and restricts phylogenetically distinct vacuolar pathogens, including the bacterium Chlamydia and the microsporidian Encephalitozoon. Quantitative proteomics coupled to a tailored CRISPR screen, uncovered components of the cysteine/Arg N-degron pathway3 that modify IL32 through oxidation-dependent arginylation, thereby enabling the recruitment of the autophagy machinery to pathogen-containing vacuoles. A forward genetics screen in Chlamydia trachomatis, the leading cause of sexually transmitted bacterial infection, identified the secreted virulence factor IncS as an evasion factor that blocks IL32 targeting and shields this human pathogen from xenophagy. These findings establish an IL32-dependent intracellular sensing mechanism linking IFN{gamma} signaling to N-degron-mediated xenophagy, revealing a broadly relevant axis of human host-pathogen conflict.

immunology↗

bioRxiv2025

Guanylate-Binding Proteins Promote Host Defense Against Leishmania major by Balancing iNOS/Arg-1 in Myeloid Cells

Cutaneous leishmaniasis (CL) is a debilitating neglected tropical disease characterized by lesions that can range from self-healing to permanent disfigurations. A predominant Th1 response, which stimulates IFN-{gamma} production, is crucial for parasite control during self-healing CL. While IFN-{gamma} primarily activates macrophages to produce nitric oxide via inducible nitric oxide synthase (iNOS) leading to parasite control, IFN-{gamma} also activates other downstream pathways involved in cell autonomous immunity. One such pathway is the activation of guanylate binding proteins (GBPs), a class of interferon inducible GTPases. However, the role of GBPs during CL has been minimally explored. Utilizing RNA-Seq we found that Leishmania major infection leads to the upregulation of several GBPs in C57Bl/6 mice. In vitro studies using GBPChr3 knockout (KO), and C57Bl/6 control mice reveal that bone marrow-derived macrophages (BMDMs) from KO mice exhibit higher parasite burdens following IFN-{gamma} treatment, independent of GBP localization to the parasite. Single-cell RNA-Seq identifies macrophages as the primary expressers of GBPs during L. major infection in vivo. In vivo, GBPChr3 KO mice display increased disease severity and parasite load. GBPChr3 KO macrophages and monocytes show elevated ARG-1 and reduced iNOS expression, indicating a shift toward a parasite-permissive environment that supports parasite growth. These findings highlight a critical role for GBPs in immune-mediated control of CL.

immunology↗

bioRxiv2025

Human giant GTPase GVIN1 forms an antimicrobial coatomer around the intracellular bacterial pathogen Burkholderia thailandensis

Several human pathogens exploit the kinetic forces generated by polymerizing actin to power their intracellular motility. Human cell-autonomous immune responses activated by the cytokine interferon-gamma (IFNg) interfere with such microbial actin-based motility, yet the underlying molecular mechanisms are poorly defined. Here, we identify the IFNg-inducible human giant GTPases GVIN1 as a novel host defense protein that blocks the bacterial pathogen Burkholderia thailandensis from high-jacking the hosts actin polymerization machinery. We found that GVIN1 proteins form a coatomer around cytosolic bacteria and prevent Burkholderia from establishing force-generating actin comet tails. Coatomers formed by a second IFNg-inducible GTPase, human guanylate binding protein 1 (GBP1), constitute a GVIN1-independent but mechanistically related anti-motility pathway. We show that coating with either GVIN1 or GBP1 displaces the Burkholderia outer membrane protein BimA, an actin nucleator that is essential for actin tail formation. Both GVIN1 and GBP1 coatomers require additional IFNg-inducible co-factors to disrupt the membrane localization of BimA, demonstrating the existence of two parallel-acting IFNg-inducible defense modules that evolved to target a virulence trait critical for the pathogenesis of numerous bacterial infectious agents.

immunology↗

bioRxiv2024

The Shigella flexneri effector IpaH1.4 facilitates RNF213 degradation and protects cytosolic bacteria against interferon-induced ubiquitylation

A central signal that marshals host defense against many infections is the lymphocyte-derived cytokine interferon-gamma (IFN{gamma}). The IFN{gamma} receptor is expressed on most human cells, and its activation leads to the expression of antimicrobial proteins that execute diverse cell-autonomous immune programs. One such immune program consists of the sequential detection, ubiquitylation, and destruction of intracellular pathogens. Recently, the IFN{gamma}-inducible ubiquitin E3 ligase RNF213 was identified as a pivotal mediator of such a defense axis. RNF213 provides host protection against viral, bacterial, and protozoan pathogens. To establish infections, potentially susceptible intracellular pathogens must have evolved mechanisms that subdue RNF213-controlled cell-autonomous immunity. In support of this hypothesis, we demonstrate here that a causative agent of bacillary dysentery, Shigella flexneri, uses the type III secretion system (T3SS) effector IpaH1.4 to induce the degradation of RNF213. S. flexneri mutants lacking IpaH1.4 expression are bound and ubiquitylated by RNF213 in the cytosol of IFN{gamma}-primed host cells. Linear (M1-) and lysine-linked ubiquitylation of S. flexneri requires RNF213 but is independent of the linear ubiquitin chain assembly complex (LUBAC). We find that ubiquitylation of S. flexneri is insufficient to kill intracellular bacteria, suggesting that S. flexneri employs additional virulence factors to escape from host defenses that operate downstream from RNF213-driven ubiquitylation. In brief, this study identified the bacterial IpaH1.4 protein as an inhibitor of mammalian RNF213 and highlights evasion of RNF213-driven immunity as a characteristic of the human-tropic pathogen Shigella.

microbiology↗

bioRxiv2023

An Aging-Susceptible Circadian Rhythm Controls Cutaneous Antiviral Immunity

Aged skin is prone to viral infections, but the mechanisms responsible for this immunosenescent immune risk are unclear. We observed that aged murine and human skin expressed reduced antiviral proteins (AVPs) and circadian regulators including Bmal1 and Clock. Bmal1 and Clock were found to control rhythmic AVP expression in skin and such circadian-control of AVPs was diminished by disruption of immune cell interleukin 27 signaling and deletion of Bmal1/Clock genes in mouse skins, as well as siRNA-mediated knockdown of CLOCK in human primary keratinocytes. We found that treatment of circadian enhancing agents, nobiletin and SR8278, reduced infection of herpes simplex virus 1 (HSV1) in epidermal explants and human keratinocytes in a Bmal1/Clock-dependent manner. Circadian enhancing treatment also reversed susceptibility of aging murine skin and human primary keratinocytes to viral infection. These findings reveal an evolutionarily conserved and age-sensitive circadian regulation of cutaneous antiviral immunity, underscoring circadian restoration as an antiviral strategy in aging populations.

immunology↗

bioRxiv2022

Shigella IpaH9.8 limits GBP1-dependent LPS release from intracytosolic bacteria to suppress caspase-4 activation

Pyroptosis is an inflammatory form of cell death induced upon the recognition of invading microbes. During an infection, pyroptosis is enhanced in interferon-gamma (IFN{gamma}) exposed cells due to the actions of members of the guanylate binding protein (GBP) family. GBPs promote activation of caspase-4 (CASP4) by enhancing its interactions with lipopolysaccharide (LPS), a component of the outer envelope of Gram-negative bacteria. Once activated, CASP4 promotes the formation of non-canonical inflammasomes, signaling platforms that mediate pyroptosis. To establish an infection, intracellular bacterial pathogens, like Shigella species, inhibit pyroptosis. The pathogenesis of Shigella is dependent on its type III secretion system, which injects ~30 effector proteins into host cells. Upon entry into host cells, Shigella are encapsulated by GBP1, followed by GBP2, GBP3, GBP4, and in some cases, CASP4. It has been suggested that the recruitment of CASP4 to bacteria leads to its activation. Here, we demonstrate that two Shigella effectors, OspC3 and IpaH9.8, cooperate to inhibit CASP4-mediated pyroptosis. We show that in the absence of OspC3, an inhibitor of CASP4, IpaH9.8 inhibits pyroptosis via its known degradation of GBPs. We find that, while some LPS is present within the host cell cytosol of epithelial cells infected with wild type Shigella, in the absence of IpaH9.8, increased amounts are shed in a GBP1-dependent manner. Furthermore, that additional IpaH9.8 targets, likely GBPs, promote CASP4 activation, even in the absence of GBP1. These observations suggest that by promoting LPS release, GBP1 provides CASP4 access to LPS, thus promoting host cell death via pyroptosis. Significance StatementNon-canonical inflammasomes mediate the death of intestinal epithelial cells in response to invading Gram-negative bacterial pathogens. CASP4 is activated upon binding to LPS. Recent studies have suggested that GBP1 recruits and enables the activation of CASP4 on the surface of intracytosolic bacteria. This has led to the proposal that non-canonical inflammasomes assemble on the bacterial surface due to GBP1-mediated access of CASP4 to the membrane-embedded lipid A component of LPS. Here, we show that by targeting the degradation of GBPs, Shigella IpaH9.8 limits the release of LPS from intracellular Shigella into the host cell cytosol in a GBP1-dependent manner. Furthermore, we find evidence to suggest that in the absence of GBP1, other members of the GBP family promote CASP4 activation.

microbiology↗

bioRxiv2022

Differential requirement for IRGM proteins during tuberculosis infection in mice

Mycobacterium tuberculosis (Mtb) is a bacterium that exclusively resides in human hosts and remains a dominant cause of morbidity and mortality among infectious diseases worldwide. Host protection against Mtb infection is dependent on the function of immunity-related GTPase clade M (IRGM) proteins. Polymorphisms in human IRGM associate with altered susceptibility to mycobacterial disease, and human IRGM promotes the delivery of Mtb into degradative autolysosomes. Among the three murine IRGM orthologs, Irgm1 has been singled out as essential for host protection during Mtb infections in cultured macrophages and in vivo. However, whether the paralogous murine Irgm genes, Irgm2 and Irgm3, play roles in host defense against Mtb or exhibit functional relationships with Irgm1 during Mtb infection remains undetermined. Here, we report that Irgm1-/- mice are indeed acutely susceptible to aerosol infection with Mtb, yet the additional deletion of the paralogous Irgm3 gene restores protective immunity to Mtb infections in Irgm1-deficient animals. Mice lacking all three Irgm genes (panIrgm-/-) are characterized by shifted lung cytokine profiles at 4 and 24 weeks post infection, but control disease until the very late stages of the infection, when panIrgm-/- mice display increased mortality compared to wild type mice. Collectively, our data demonstrate that disruptions in the balance between Irgm isoforms is more detrimental to the Mtb-infected host than total loss of Irgm-mediated host defense, a concept that also needs to be considered in the context of human Mtb susceptibility linked to IRGM polymorphisms.

microbiology↗

bioRxiv2022

GBP2 aggregates LPS and activates the caspase-4 inflammasome independent of the bacterial encapsulation factor GBP1

Sensing and killing of intracellular bacterial pathogens are important features of cell-autonomous immunity. The cytokine gamma-interferon (IFN{gamma}) enhances cell-autonomous immunity through upregulation of interferon stimulated genes such as guanylate binding proteins (GBPs). GBPs promote defense against Gram-negative cytosolic bacteria in part through the induction of an inflammatory cell death pathway called pyroptosis. To activate pyroptosis, GBPs facilitate caspase-4 sensing of the Gram-negative bacterial outer membrane component lipopolysaccharide (LPS). There are seven human GBP paralogs and it is unclear how each GBP contributes to LPS sensing and pyroptosis induction. GBP1 forms a multimeric microcapsule on the surface of cytosolic bacteria through direct interactions with LPS and recruits caspase-4 to bacteria, a process deemed essential for caspase-4 activation. In contrast to GBP1, closely related paralog GBP2 is unable to bind bacteria on its own but requires GBP1 for direct bacterial binding. Unexpectedly, we find that GBP2 overexpression can restore Gram-negative-induced pyroptosis in GBP1KO cells, without GBP2 binding to the bacterial surface. A mutant of GBP1 that lacks the triple arginine motif required for microcapsule formation also rescues pyroptosis in GBP1KO cells, showing that binding to bacteria is dispensable for GBPs to promote pyroptosis. Instead, we find that GBP2, like GBP1, directly binds and aggregates free LPS through protein polymerization. This provides a novel mechanistic framework for non-canonical inflammasome activation where GBP1 or GBP2 assemble cytosol-contaminating LPS into a protein-LPS interface for caspase-4 activation as part of a coordinated host response to Gram-negative bacterial infections. Significance StatementSensing Gram-negative bacterial lipopolysaccharide by human caspase-4 is critical for host defense to intracellular Gram-negative bacterial pathogens. Human guanylate binding proteins (GBPs) facilitate caspase-4 activation in response to Gram-negative infections by a poorly understood mechanism. The prevailing model suggests GBP1 binding to bacteria and consequential recruitment of caspase-4 to the bacterial surface are essential for triggering this host response. Here, we show GBP1 binding to bacteria is dispensable for caspase-4 activation and identify GBP2 as an additional lipopolysaccharide-binding protein that can functionally replace GBP1. We demonstrate that GBP1 and GBP2 share the ability to form lipopolysaccharide-protein complexes, which, we propose, allow caspase-4 activation. Our study provides a new mechanistic framework for cytosolic LPS sensing.

immunology↗

bioRxiv2022

Irgm proteins attenuate inflammatory disease in mouse models of genital Chlamydia infection

Chlamydiae are obligate intracellular bacterial pathogens that may cause genital pathology via induction of destructive host immune responses. Human-adapted Chlamydia trachomatis causes inflammatory disease in human hosts but is easily cleared in mice, and mouse-adapted Chlamydia muridarum establishes a productive and pathogenic infection in murine hosts. While numerous anti-chlamydial host resistance factors have been discovered in mice and humans alike, little is known about host factors promoting host fitness independent of host resistance. Here, we show that interferon-inducible immunity-related GTPase M (Irgm) proteins function as such host factors ameliorating infection-associated sequalae in the murine female genital tract, thus characterizing Irgm proteins as mediators of disease tolerance. Specifically, we demonstrate that mice deficient for all three murine Irgm paralogs (pan-Irgm-/-) are defective for cell-autonomous immunity to C. trachomatis, which correlates with an early and transient increase in bacterial burden and sustained hyperinflammation in vivo. In contrast, upon infection of pan-Irgm-/- mice with C. muridarum, bacterial burden is unaffected, yet genital inflammation and scarring pathology are nonetheless increased, demonstrating that Irgm proteins can promote host fitness without altering bacterial burden. Additionally, pan-Irgm-/- mice display increased granulomatous inflammation in genital Chlamydia infection, implicating Irgm proteins in the regulation of granuloma formation and maintenance. These findings demonstrate that Irgm proteins regulate pathogenic immune responses to Chlamydia infection in vivo, establishing an effective infection model to examine the immunoregulatory functions and mechanisms of Irgm proteins. ImportanceIn response to genital Chlamydia infection, the immune system mounts a proinflammatory response to resist the pathogen, yet inflammation must be tightly controlled to avoid collateral damage and scarring to host genital tissue. Variation in the human IRGM gene is associated with susceptibility to autoinflammatory diseases but its role in ameliorating inflammatory diseases caused by infections is poorly defined. Here we use mice deficient for all three murine Irgm paralogs to demonstrate that Irgm proteins not only provide host resistance to Chlamydia infections but also limit associated inflammation in the female genital tract. In particular, we find that murine Irgm expression prevents granulomatous inflammation, which parallels inflammatory diseases associated with variants in human IRGM. Our findings therefore establish genital Chlamydia infection as a useful model to study the roles for Irgm proteins in both promoting protective immunity and limiting pathogenic inflammation.

immunology↗

bioRxiv2020

Mycobacterium tuberculosis evasion of Guanylate Binding Protein-mediated host defense in mice requires the ESX1 secretion system.

Cell-intrinsic immune mechanisms control intracellular pathogens that infect eukaryotes. The intracellular pathogen Mycobacterium tuberculosis (Mtb) evolved to withstand cell-autonomous immunity to cause persistent infections and disease. A potent inducer of cell-autonomous immunity is the lymphocyte-derived cytokine IFN{gamma}. While the production of IFN{gamma} by T cells is essential to protect against Mtb, it is not capable of fully eradicating Mtb infection. This suggests that Mtb evades a subset of IFN{gamma}-mediated antimicrobial responses, yet what mechanisms Mtb resists remains unclear. The IFN{gamma}-inducible Guanylate binding proteins (GBPs) are key host defense proteins able to control infections with intracellular pathogens. GBPs were previously shown to directly restrict Mycobacterium bovis BCG yet their role during Mtb infection has remained unknown. Here, we examine the importance of a cluster of five GBPs on mouse chromosome 3 in controlling Mycobacterial infection. While M. bovis BCG is directly restricted by GBPs, we find that the GBPs on chromosome 3 do not contribute to the control of Mtb replication or the associated host response to infection. The differential effects of GBPs during Mtb versus M. bovis BCG infection is at least partially explained by the absence of the ESX1 secretion system from M. bovis BCG, since Mtb mutants lacking the ESX1 secretion system become similarly susceptible to GBP-mediated immune defense. Therefore, this specific genetic interaction between the murine host and Mycobacteria reveals a novel function for the ESX1 virulence system in the evasion of GBP-mediated immunity.

microbiology↗

bioRxiv2020

Dynamin-related Irgm proteins modulate LPS-induced caspase-4 activation and septic shock

Inflammation associated with gram-negative bacterial infections is often instigated by the bacterial cell wall component lipopolysaccharide (LPS). LPS-induced inflammation and resulting life-threatening sepsis are mediated by the two distinct LPS receptors TLR4 and caspase-4. Whereas the regulation of TLR4 activation by extracellular and phago-endosomal LPS has been studied in great detail, auxiliary host factors that specifically modulate recognition of cytosolic LPS by caspase-4 are largely unknown. This study identifies dynamin-related membrane remodeling proteins belonging to the family of Immunity related GTPases M clade (IRGM) as negative regulators of caspase-4 activation in macrophages. Phagocytes lacking expression of mouse isoform Irgm2 aberrantly activate caspase-4-dependent inflammatory responses when exposed to extracellular LPS, bacterial outer membrane vesicles or gram-negative bacteria. Consequently, Irgm2-deficient mice display increased susceptibility to caspase-4-mediated septic shock in vivo. This Irgm2 phenotype is partly reversed by the simultaneous genetic deletion of the two additional Irgm paralogs Irgm1 and Irgm3, indicating that dysregulated Irgm isoform expression disrupts intracellular LPS processing pathways that limit LPS availability for caspase-4 activation.

immunology↗

bioRxiv2019

IRG1 and iNOS act redundantly with other interferon gamma induced factors to restrict intracellular replication of Legionella pneumophila.

Interferon gamma (IFN{gamma}) restricts the intracellular replication of many pathogens, but how IFN{gamma} confers cell-intrinsic pathogen resistance remains unclear. For example, intracellular replication of the bacterial pathogen Legionella pneumophila in macrophages is potently curtailed by IFN{gamma}, but consistent with prior results, no individual genetic deficiency we tested compromised IFN{gamma}-mediated control. Intriguingly, however, we observed that the glycolysis inhibitor 2-deoxyglucose (2DG) partially rescued L. pneumophila replication in IFN{gamma}-treated macrophages. 2DG inhibits glycolysis and triggers the unfolded protein response, but unexpectedly, it appears these effects are not responsible for perturbing the antimicrobial activity of IFN{gamma}. Instead, we found that 2DG rescues bacterial replication predominantly by inhibiting the induction of two key antimicrobial factors, inducible nitric oxide synthase (iNOS) and immune responsive gene 1 (IRG1). Using immortalized and primary macrophages deficient in iNOS and IRG1, we confirm that loss of both iNOS and IRG1, but not individual deficiency in each gene, partially reduces IFN{gamma}-mediated restriction of L. pneumophila. Further, using a combinatorial CRISPR/Cas9 mutagenesis approach, we find that mutation of iNOS and IRG1 in combination with four other genes (CASP11, IRGM1, IRGM3 and NOX2) results in a total loss of L. pneumophila restriction by IFN{gamma} in primary bone marrow macrophages. There are few, if any, other examples in which the complete set of cell-intrinsic factors required for IFN{gamma}-mediated restriction of an intracellular bacterial pathogen have been genetically identified. Our results highlight the combinatorial strategy used by hosts to block the exploitation of macrophages by pathogens.\n\nImportanceLegionella pneumophila is one example among many species of pathogenic bacteria that replicate within mammalian macrophages during infection. The immune signaling factor interferon gamma (IFN{gamma}) blocks L. pneumophila replication in macrophages and is an essential component of the immune response to L. pneumophila and other intracellular pathogens. However, to date, no study has determined the exact molecular factors induced by IFN{gamma} that are required for its activity. We generated macrophages lacking different combinations of IFN{gamma}-induced genes in an attempt to find a genetic background in which there is a complete loss of IFN{gamma}-mediated restriction of L. pneumophila. We successfully identified six genes that comprise the totality of the IFN{gamma}-dependent restriction of L. pneumophila replication in macrophages. Our results clarify the molecular basis underlying the potent effects of IFN{gamma} and highlight how redundancy downstream of IFN{gamma} is key to prevent exploitation of the macrophage niche by pathogens.

immunology↗

bioRxiv2019

A rapidly evolving polybasic motif modulates bacterial detection by guanylate binding proteins

Cell-autonomous immunity relies on the rapid detection of invasive pathogens by host proteins. Guanylate binding proteins (GBPs) have emerged as key mediators of vertebrate immune defense through their ability to recognize a diverse array of intracellular pathogens and pathogen-containing cellular compartments. Human and mouse GBPs have been shown to target distinct groups of microbes, although the molecular determinants of pathogen specificity remain unclear. We show that rapid diversification of a C-terminal polybasic motif (PBM) in primate GBPs controls recognition of the model cytosolic bacterial pathogen Shigella flexneri. By swapping this membrane-binding motif between primate GBP orthologs, we find that the ability to target S. flexneri has been enhanced and lost in specific lineages of New World monkeys. Single substitutions in rapidly evolving sites of the GBP1 PBM are sufficient to abolish or restore bacterial detection abilities, illustrating a role for epistasis in the evolution of pathogen recognition. We further demonstrate that the squirrel monkey GBP2 C-terminal domain recently gained the ability to target S. flexneri through a stepwise process of convergent evolution. These findings reveal a mechanism by which accelerated evolution of a PBM shifts GBP target specificity and aid in resolving the molecular basis of GBP function in cell-autonomous immune defense.

microbiology↗

bioRxiv2017

Detection of cytosolic Shigella flexneri via a C-terminal triple-arginine motif of GBP1 inhibits actin-based motility

Dynamin-like guanylate binding proteins (GBPs) are gamma interferon (IFN{gamma})-inducible host defense proteins that can associate with cytosol-invading bacterial pathogens. Mouse GBPs promote the lytic destruction of targeted bacteria in the host cell cytosol but the antimicrobial function of human GBPs and the mechanism by which these proteins associate with cytosolic bacteria are poorly understood. Here, we demonstrate that human GBP1 is unique amongst the seven human GBP paralogs in its ability to associate with at least two cytosolic Gram-negative bacteria, Burkholderia thailandensis and Shigella flexneri. Rough lipopolysaccharide (LPS) mutants of S. flexneri co-localize with GBP1 less frequently than wildtype S. flexneri, suggesting that host recognition of O-antigen promotes GBP1 targeting to Gram-negative bacteria. The targeting of GBP1 to cytosolic bacteria, via a unique triple-arginine motif present in its C-terminus, promotes the co-recruitment of four additional GBP paralogs (GBP2, GBP3, GBP4 and GBP6). GBP1-decorated Shigella replicate but fail to form actin tails leading to their intracellular aggregation. Consequentially, wildtype but not the triple-arginine GBP1 mutant restricts S. flexneri cell-to-cell spread. Furthermore, human-adapted S. flexneri, through the action of one its secreted effectors, IpaH9.8, is more resistant to GBP1 targeting than the non-human-adapted bacillus B. thailandensis. These studies reveal that human GBP1 uniquely functions as an intracellular glue trap inhibiting the cytosolic movement of normally actin-propelled Gram-negative bacteria. In response to this powerful human defense program S. flexneri has evolved an effective counter-defense to restrict GBP1 recruitment.\n\nImportanceSeveral pathogenic bacterial species evolved to invade, reside and replicate inside the cytosol of their host cells. One adaptation common to most cytosolic bacterial pathogens is the ability to co-opt the hosts actin polymerization machinery, in order to generate force for intracellular movement. This actin-based motility enables Gram-negative bacteria such as Shigella to propel themselves into neighboring cells thereby spreading from host cell to host cell without exiting the intracellular environment. Here, we show that the human protein GBP1 acts as a cytosolic glue trap capturing cytosolic Gram-negative bacteria through a unique protein motif and preventing disseminated infections in cell culture models. To escape from this GBP1-mediated host defense, Shigella employs a virulence factor that prevents or dislodges the association of GBP1 with cytosolic bacteria. Thus, therapeutic strategies to restore GBP1 binding to Shigella may lead to novel treatment options for shigellosis in the future.

microbiology↗