Imperial College London

ProfessorGadFrankel

Faculty of Natural SciencesDepartment of Life Sciences

Professor of Molecular Pathogenesis
 
 
 
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Contact

 

+44 (0)20 7594 5253g.frankel

 
 
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Location

 

1.46Flowers buildingSouth Kensington Campus

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Summary

 

Publications

Publication Type
Year
to

277 results found

Vieira MFM, Hernandez G, Zhong Q, Arbesú M, Veloso T, Gomes T, Martins ML, Monteiro H, Frazão C, Frankel G, Zanzoni A, Cordeiro TNet al., 2024, The pathogen-encoded signalling receptor Tir exploits host-like intrinsic disorder for infection., Commun Biol, Vol: 7

The translocated intimin receptor (Tir) is an essential type III secretion system (T3SS) effector of attaching and effacing pathogens contributing to the global foodborne disease burden. Tir acts as a cell-surface receptor in host cells, rewiring intracellular processes by targeting multiple host proteins. We investigated the molecular basis for Tir's binding diversity in signalling, finding that Tir is a disordered protein with host-like binding motifs. Unexpectedly, also are several other T3SS effectors. By an integrative approach, we reveal that Tir dimerises via an antiparallel OB-fold within a highly disordered N-terminal cytosolic domain. Also, it has a long disordered C-terminal cytosolic domain partially structured at host-like motifs that bind lipids. Membrane affinity depends on lipid composition and phosphorylation, highlighting a previously unrecognised host interaction impacting Tir-induced actin polymerisation and cell death. Furthermore, multi-site tyrosine phosphorylation enables Tir to engage host SH2 domains in a multivalent fuzzy complex, consistent with Tir's scaffolding role and binding promiscuity. Our findings provide insights into the intracellular Tir domains, highlighting the ability of T3SS effectors to exploit host-like protein disorder as a strategy for host evasion.

Journal article

Seddon C, Frankel G, Beis K, 2024, Structure of the outer membrane porin OmpW from the pervasive pathogen Klebsiella pneumoniae, Acta Crystallographica Section F: Structural Biology and Crystallization Communications Online, Vol: 80, Pages: 22-27, ISSN: 1744-3091

Conjugation is the process by which plasmids, including those that carry antibiotic-resistance genes, are mobilized from one bacterium (the donor) to another (the recipient). The conjugation efficiency of IncF-like plasmids relies on the formation of mating-pair stabilization via intimate interactions between outer membrane proteins on the donor (a plasmid-encoded TraN isoform) and recipient bacteria. Conjugation of the R100-1 plasmid into Escherichia coli and Klebsiella pneumoniae (KP) recipients relies on pairing between the plasmid-encoded TraNα in the donor and OmpW in the recipient. Here, the crystal structure of K. pneumoniae OmpW (OmpWKP) is reported at 3.2 Å resolution. OmpWKP forms an eight-stranded β-barrel flanked by extracellular loops. The structures of E. coli OmpW (OmpWEC) and OmpWKP show high conservation despite sequence variability in the extracellular loops.

Journal article

Di Pilato V, Codda G, Niccolai C, Willison E, Wong JLC, Coppo E, Frankel G, Marchese A, Rossolini GMet al., 2024, Functional features of KPC-109, a novel 270-loop KPC-3 mutant mediating resistance to avibactam-based β-lactamase inhibitor combinations and cefiderocol., Int J Antimicrob Agents, Vol: 63

OBJECTIVES: To investigate a ceftazidime/avibactam (CZA)-resistant Klebsiella pneumoniae (NE368), isolated from a patient exposed to CZA, expressing a novel K. pneumoniae carbapenemase (KPC)-3 variant (KPC-109). METHODS: Antimicrobial susceptibility testing was performed by reference broth microdilution. Whole-genome sequencing (WGS) analysis of NE368 was performed combining a short- and long-reads approach (Illumina and Oxford Nanopore Technologies). Functional characterization of KPC-109 was performed to investigate the impact of KPC-109 production on the β-lactam resistance phenotype of various Escherichia coli and Klebsiella pneumoniae strains, including derivatives of K. pneumoniae with OmpK35 and OmpK36 porin alterations. Horizontal transfer of the KPC-109-encoding plasmid was investigated by conjugation and transformation experiments. RESULTS: K. pneumoniae NE368 was isolated from a patient after repeated CZA exposure, and showed resistance to CZA, fluoroquinolones, piperacillin/tazobactam, expanded-spectrum cephalosporins, amikacin, carbapenems and cefiderocol. WGS revealed the presence of a large chimeric plasmid of original structure (pKPN-NE368), encoding a novel 270-loop mutated KPC-3 variant (KPC-109; ins_270_KYNKDD). KPC-109 production mediated resistance/decreased susceptibility to avibactam-based combinations (with ceftazidime, cefepime and aztreonam) and cefiderocol, with a trade-off on carbapenem resistance. However, in the presence of porin alterations commonly encountered in high-risk clonal lineages of K. pneumoniae, KPC-109 was also able to confer clinical-level resistance to carbapenems. Resistance of NE368 to cefiderocol was likely contributed by KPC-109 production acting in concert with a mutated EnvZ sensor kinase. The KPC-109-encoding plasmid did not appear to be conjugative. CONCLUSIONS: These findings expand current knowledge about the diversity of emerging KPC enzyme variants with 270-loop alterations that can be encountered in the

Journal article

Herzog MK-M, Cazzaniga M, Peters A, Shayya N, Beldi L, Hapfelmeier S, Heimesaat MMM, Bereswill S, Frankel G, Gahan CGM, Hardt W-Det al., 2023, Mouse models for bacterial enteropathogen infections: insights into the role of colonization resistance, GUT MICROBES, Vol: 15, ISSN: 1949-0976

Journal article

Wiggins BG, Wang Y-F, Burke A, Grunberg N, Vlachaki Walker JM, Dore M, Chahrour C, Pennycook BR, Sanchez-Garrido J, Vernia S, Barr AR, Frankel G, Birdsey GM, Randi AM, Schiering Cet al., 2023, Endothelial sensing of AHR ligands regulates intestinal homeostasis, Nature, Vol: 621, Pages: 821-829, ISSN: 0028-0836

Endothelial cells (ECs) line the blood and lymphatic vasculature, and act as an essential physical barrier, control nutrient transport, facilitate tissue immunosurveillance, and coordinate angiogenesis/ lymphangiogenesis1,2. In the intestine, dietary and microbial cues are particularly important in the regulation of organ homeostasis. However, whether enteric ECs actively sense and integrate such signals is currently unknown. Here, we show that the aryl hydrocarbon receptor (AHR) acts as a critical node for EC-sensing of dietary metabolites in adult mice and human primary ECs. We first established a comprehensive single-cell endothelial atlas of the mouse small intestine, uncovering the cellular complexity and functional heterogeneity of blood and lymphatic ECs. Analyses of AHR mediated responses at single-cell resolution identified tissue-protective transcriptional signatures and regulatory networks promoting cellular quiescence and vascular normalcy at steady state. Endothelial AHR-deficiency in adult mice resulted in dysregulated inflammatory responses, and the initiation of proliferative pathways. Furthermore, endothelial sensing of dietary AHR ligands was required for optimal protection against enteric infection. In human ECs, AHR signalling promoted quiescence and restrained activation by inflammatory mediators. Together, our data provide a comprehensive dissection of the impact of environmental sensing across the spectrum of enteric endothelia, demonstrating that endothelial AHR signalling integrates dietary cues to maintain tissue homeostasis by promoting EC quiescence and vascular normalcy.

Journal article

Frankel G, David S, Low WW, Seddon C, Wong JLC, Beis Ket al., 2023, Plasmids pick a bacterial partner before committing to conjugation, NUCLEIC ACIDS RESEARCH, Vol: 51, Pages: 8925-8933, ISSN: 0305-1048

Journal article

Low WW, Seddon C, Beis K, Frankel Get al., 2023, The Interaction of the F-Like Plasmid-Encoded TraN Isoforms with Their Cognate Outer Membrane Receptors, JOURNAL OF BACTERIOLOGY, Vol: 205, ISSN: 0021-9193

Journal article

Wong JLC, Romano M, Kerry LE, Kwong H-S, Low W-W, Brett SJ, Clements A, Beis K, Frankel Get al., 2023, Author Correction: OmpK36-mediated Carbapenem resistance attenuates ST258 Klebsiella pneumoniae in vivo., Nat Commun, Vol: 14

Journal article

Wong J, David S, Sanchez Garrido J, Woo J, Low WW, Morecchiato F, Giani T, Rossolini GM, Beis K, Brett S, Clements A, Aaenensen D, Rouskin S, Frankel Get al., 2022, Recurrent emergence of Klebsiella pneumoniae carbapenem resistance mediated by an inhibitory ompK36 mRNA secondary structure, Proceedings of the National Academy of Sciences of USA, Vol: 119, Pages: 1-12, ISSN: 0027-8424

Outer membrane porins in Gram-negative bacteria facilitate antibiotic influx. In Klebsiella pneumoniae (KP), modifications in the porin OmpK36 are implicated in increasing resistance to carbapenems. Analysis of large KP genome collections, encompassing major healthcare-associated clones, revealed the recurrent emergence of a synonymous cytosine to thymine transition at position 25 (25c>t) in ompK36. We show that the 25c>t transition increases carbapenem resistance through depletion of OmpK36 from the outer membrane. The mutation attenuates KP in a murine pneumonia model, which accounts for its limited clonal expansion observed by phylogenetic analysis. However, in the context of carbapenem treatment, the 25c>t transition tips the balance towards treatment failure, thus accounting for its recurrent emergence. Mechanistically, the 25c>t transition mediates an intramolecular mRNA interaction between a uracil encoded by 25t and the first adenine within the Shine-Dalgarno sequence. This specific interaction leads to the formation of an RNA stem structure, which obscures the ribosomal binding site thus disrupting translation. While mutations reducing OmpK36 expression via transcriptional silencing are known, we uniquely demonstrate the repeated selection of a synonymous ompK36 mutation mediating translational suppression in response to antibiotic pressure.

Journal article

Chaukimath P, Frankel G, Visweswariah SS, 2022, The metabolic impact of bacterial infection in the gut, FEBS JOURNAL, ISSN: 1742-464X

Journal article

David S, Wong JLC, Sanchez-Garrido J, Kwong H-S, Low WW, Morecchiato F, Giani T, Rossolini GM, Brett SJ, Clements A, Beis K, Aanensen DM, Frankel Get al., 2022, Widespread emergence of OmpK36 loop 3 insertions among multidrug-resistant clones of Klebsiella pneumoniae., PLoS Pathogens, Vol: 18, Pages: 1-23, ISSN: 1553-7366

Mutations in outer membrane porins act in synergy with carbapenemase enzymes to increase carbapenem resistance in the important nosocomial pathogen, Klebsiella pneumoniae (KP). A key example is a di-amino acid insertion, Glycine-Aspartate (GD), in the extracellular loop 3 (L3) region of OmpK36 which constricts the pore and restricts entry of carbapenems into the bacterial cell. Here we combined genomic and experimental approaches to characterise the diversity, spread and impact of different L3 insertion types in OmpK36. We identified L3 insertions in 3588 (24.1%) of 14,888 KP genomes with an intact ompK36 gene from a global collection. GD insertions were most common, with a high concentration in the ST258/512 clone that has spread widely in Europe and the Americas. Aspartate (D) and Threonine-Aspartate (TD) insertions were prevalent in genomes from Asia, due in part to acquisitions by KP sequence types ST16 and ST231 and subsequent clonal expansions. By solving the crystal structures of novel OmpK36 variants, we found that the TD insertion causes a pore constriction of 41%, significantly greater than that achieved by GD (10%) or D (8%), resulting in the highest levels of resistance to selected antibiotics. We show that in the absence of antibiotics KP mutants harbouring these L3 insertions exhibit both an in vitro and in vivo competitive disadvantage relative to the isogenic parental strain expressing wild type OmpK36. We propose that this explains the reversion of GD and TD insertions observed at low frequency among KP genomes. Finally, we demonstrate that strains expressing L3 insertions remain susceptible to drugs targeting carbapenemase-producing KP, including novel beta lactam-beta lactamase inhibitor combinations. This study provides a contemporary global view of OmpK36-mediated resistance mechanisms in KP, integrating surveillance and experimental data to guide treatment and drug development strategies.

Journal article

Low WW, Wong J, Beltran L, Seddon C, David S, Kwong H-S, Bizeau T, Wang F, Pena A, Costa TRD, Pham B, Chen M, Egelman E, Beis K, Frankel Get al., 2022, Mating pair stabilization mediates bacterial conjugation species specificity, Nature Microbiology, Vol: 7, Pages: 1016-1027, ISSN: 2058-5276

Bacterial conjugation mediates contact-dependent transfer of DNA from donor to recipient bacteria, thus facilitating thespread of virulence and resistance plasmids. Here we describe how variants of the plasmid-encoded donor outer membrane(OM) protein TraN cooperate with distinct OM receptors in recipients to mediate mating pair stabilization and efficient DNAtransfer. We show that TraN from the plasmids pKpQIL (Klebsiella pneumoniae), R100-1 (Shigella flexneri) and pSLT (SalmonellaTyphimurium), and the prototypical F plasmid (Escherichia coli) interact with OmpK36, OmpW and OmpA, respectively.Cryo-EM analysis revealed that TraN pKpQIL interacts with OmpK36 through the insertion of a β-hairpin in the tip of TraN intoa monomer of the OmpK36 trimer. Combining bioinformatic analysis with AlphaFold structural predictions, we identified afourth TraN structural variant that mediates mating pair stabilization by binding OmpF. Accordingly, we devised a classifica-tion scheme for TraN homologues on the basis of structural similarity and their associated receptors: TraNα (OmpW), TraNβ(OmpK36), TraNγ (OmpA), TraNδ (OmpF). These TraN-OM receptor pairings have real-world implications as they reflect thedistribution of resistance plasmids within clinical Enterobacteriaceae isolates, demonstrating the importance of mating pairstabilization in mediating conjugation species specificity. These findings will allow us to predict the distribution of emergingresistance plasmids in high-risk bacterial pathogens.

Journal article

Sanchez Garrido J, Ruano-Gallego D, Choudhary JS, Frankel Get al., 2022, The type III secretion system effector network hypothesis, Trends in Microbiology, Vol: 30, Pages: 524-533, ISSN: 0966-842X

Type III secretion system (T3SS) effectors are key virulence factors that underpin the infection strategy of many clinically important Gram-negative pathogens, including Salmonella enterica, Shigella spp, enteropathogenic and enterohaemorrhagic Escherichia coli and their murine equivalent, Citrobacter rodentium. The cellular processes or proteins targeted by the effectors can be common to multiple pathogens or pathogen-specific. The main approach to understanding T3SS-mediated pathogenesis has been to determine the contribution of one effector at a time, with the aim to piece together individual functions and unveil infection mechanisms. However, in contrast to this prevailing approach, simultaneous deletion of multiple effectors revealed that they function as an interconnected network in vivo, uncoveringeffector co-dependency and context-dependent effector essentiality. This paradigm shift in T3SS biology is at the heart of this opinion.

Journal article

Higginson EE, Nkeze J, Permala-Booth J, Kasumba IN, Lagos R, Hormazabal JC, Byrne A, Frankel G, Levine MM, Tennant SMet al., 2022, Detection of Salmonella Typhi in Bile by Quantitative Real-Time PCR, MICROBIOLOGY SPECTRUM, Vol: 10, ISSN: 2165-0497

Journal article

Kreutzberger MAB, Sobe RC, Sauder AB, Chatterjee S, Peña A, Wang F, Giron JA, Kiessling V, Costa TRD, Conticello VP, Frankel G, Kendall MM, Scharf BE, Egelman EHet al., 2022, Flagellin outer domain dimerization modulates motility in pathogenic and soil bacteria from viscous environments., Nature Communications, Vol: 13, Pages: 1-13, ISSN: 2041-1723

Flagellar filaments function as the propellers of the bacterial flagellum and their supercoiling is key to motility. The outer domains on the surface of the filament are non-critical for motility in many bacteria and their structures and functions are not conserved. Here, we show the atomic cryo-electron microscopy structures for flagellar filaments from enterohemorrhagic Escherichia coli O157:H7, enteropathogenic E. coli O127:H6, Achromobacter, and Sinorhizobium meliloti, where the outer domains dimerize or tetramerize to form either a sheath or a screw-like surface. These dimers are formed by 180° rotations of half of the outer domains. The outer domain sheath (ODS) plays a role in bacterial motility by stabilizing an intermediate waveform and prolonging the tumbling of E. coli cells. Bacteria with these ODS and screw-like flagellar filaments are commonly found in soil and human intestinal environments of relatively high viscosity suggesting a role for the dimerization in these environments.

Journal article

Serafini N, Jarade A, Surace L, Goncalves P, Sismeiro O, Varet H, Legendre R, Coppee J-Y, Disson O, Durum SK, Frankel G, Di Santo JPet al., 2022, Trained ILC3 responses promote intestinal defense, SCIENCE, Vol: 375, Pages: 859-+, ISSN: 0036-8075

Journal article

Kreutzberger MA, Sobe R, Sauder AB, Chatterjee S, Wang F, Kiessling V, Conticello V, Frankel G, Kendall M, Scharf B, Egelman EHet al., 2022, Cryo-EM of bacterial flagellar filaments with screw-like surfaces and outer domain sheaths, Publisher: CELL PRESS, Pages: 131-131, ISSN: 0006-3495

Conference paper

Mullineaux Sanders C, Kozik Z, Sanchez Garrido J, Hopkins EGD, Choudhary JS, Frankel Get al., 2022, Citrobacter rodentium infection induces persistent molecular changes and interferon gamma-dependent major histocompatibility complex class II expression in the colonic epithelium, mBio, Vol: 13, Pages: 1-18, ISSN: 2150-7511

Most studies of infections at mucosal surfaces have focused on the acute phase of the disease. Consequently, little is known about the molecular processes that underpin tissue recovery and the long-term consequences postinfection. Here, we conducted temporal deep quantitative proteomic analysis of colonic intestinal epithelial cells (cIECs) from mice infected with the natural mouse pathogen Citrobacter rodentium over time points corresponding to the late steady-state phase (10 days postinfection [DPI]), the clearance phase (13 to 20 DPI), and 4 weeks after the pathogen has been cleared (48 DPI). C. rodentium, which relies on a type III secretion system to infect, is used to model infections with enteropathogenic and enterohemorrhagic Escherichia coli. We observe a strong upregulation of inflammatory signaling and nutritional immunity responses during the clearance phase of the infection. Despite morphological tissue recovery, chromogranin B (ChgB)-positive endocrine cells remained significantly below baseline levels at 48 DPI. In contrast, we observed an increased abundance of proteins involved in antigen processing and presentation 4 weeks after pathogen clearance. In particular, long-term changes were characterized by a persistent interferon gamma (IFN-γ) response and the expression of major histocompatibility complex class II (MHCII) molecules in 60% of the EpCAM+ cIECs, which were not seen in Ifnγ−/− mice. Nonetheless, both wild-type and Ifnγ−/− mice mounted similar systemic and colonic IgG responses to C. rodentium and were equally protected from rechallenge, suggesting that cIEC MHCII is not necessary for protective immunity against C. rodentium.

Journal article

Sperandio V, Frankel G, 2022, Editorial overview: Host-microbe interactions: friends, foes and frenemies, CURRENT OPINION IN MICROBIOLOGY, Vol: 65, Pages: VIII-X, ISSN: 1369-5274

Journal article

Zhong Q, Chatterjee S, Choudhary JS, Frankel Get al., 2022, EPEC-induced activation of the Ca<SUP>2+</SUP> transporter TRPV2 leads to pyroptotic cell death, MOLECULAR MICROBIOLOGY, Vol: 117, Pages: 480-492, ISSN: 0950-382X

Journal article

Sanchez Garrido J, Alberdi L, Chatterjee S, Frankel G, Mullineaux-Sanders Cet al., 2021, Type III secretion system effector subnetworks elicit distinct host immune responses to infection, Current Opinion in Microbiology, Vol: 64, Pages: 19-26, ISSN: 1369-5274

Citrobacter rodentium, a natural mouse pathogen which colonises the colon of immuno-competent mice, provides a robust model for interrogating host-pathogen-microbiota interactions in vivo. This model has been key to providing new insights into local host responses to enteric infection, including changes inintestinal epithelial cell immuno metabolism and mucosal immunity. C. rodent iuminjects 31 bacterial effectors into epithelial cells via a type III secretion system (T3SS). Recently, these effectors were shown to be able to form multiple intracellular subnetworks which can withstand significant contractions whilst maintaining virulence. Here we highlight recent advances in understanding gut mucosal responses to infection and effector biology, as well as potential uses for artificial intelligence (AI) in understanding infectious diseaseand speculate on the role of T3SS effector networks in host adaption.

Journal article

Mullineaux Sanders C, Carson D, Hopkins E, Glegola-Madejska I, Escobar-Zepeda A, Browne H, Lawley T, Frankel Get al., 2021, Citrobacter amalonaticus inhibits the growth of Citrobacter rodentium in the gut lumen, mBio, Vol: 5, Pages: 1-19, ISSN: 2150-7511

The gut microbiota plays a crucial role in susceptibility to enteric pathogens, including Citrobacter rodentium, a model extracellular mouse pathogen that colonizes the colonic mucosa. C. rodentium infection outcomes vary between mouse strains, with C57BL/6 and C3H/HeN mice clearing or succumbing to the infection respectively. Kanamycin (Kan) treatment at the peak of C57BL/6 mouse infection with Kan-resistant C. rodentium resulted in re-localisation of the pathogen from the colonic mucosa and cecum to solely the cecal luminal contents; cessation of the Kan treatment resulted in rapid clearance of the pathogen. We now show that in C3H/HeN mice, following Kan-induced displacement of C. rodentium to the cecum, the pathogen stably colonizes the cecal lumen of 65% of the mice in the absence of continued antibiotic treatment, a phenomenon we term antibiotic-induced bacterial commensalisation (AIBC). AIBC C. rodentium was well-tolerated by the host, which showed little signs of inflammation; passaged AIBC C. rodentium robustly infected naïve C3H/HeN mice suggesting that the AIBC state is transient and did not select for genetically avirulent C. rodentium mutants. Following withdrawal of antibiotic treatment, 35% of C3H/HeN mice were able to prevent C. rodentium commensalisation in the gut lumen. These mice presented a bloom of a commensal species, Citrobacter amalonaticus, which inhibited the growth of C. rodentium in vitro in a contact-dependant manner, and luminal growth of AIBC C. rodentium in vivo. Overall our data suggest that commensal species can confer colonization resistance against closely-related pathogenic species.

Journal article

Chatterjee S, Choi A, Frankel G, 2021, A systematic review of Sec24 cargo interactome, Traffic, Vol: 22, Pages: 412-424, ISSN: 1398-9219

Endoplasmic reticulum (ER)-to-Golgi trafficking is an essential and highly conserved cellular process. The coat protein complex-II (COPII) arm of the trafficking machinery incorporates a wide array of cargo proteins into vesicles through direct or indirect interactions with Sec24, the principal subunit of the COPII coat. Approximately one-third of all mammalian proteins rely on the COPII mediated secretory pathway for membrane insertion or secretion. There are four mammalian Sec24 paralogs and three yeast Sec24 paralogs with emerging evidence of paralog-specific cargo interaction motifs. Furthermore, individual paralogs also differ in their affinity for a subset of sorting motifs present on cargo proteins. As with many aspects of protein trafficking, we lack a systematic and thorough understanding of the interaction of Sec24 with cargoes. This systematic review focuses on the current knowledge of cargo binding to both yeast and mammalian Sec24 paralogs and their ER export motifs. The analyses show that Sec24 paralog specificity of cargo (and cargo receptors) range from exclusive paralog dependence or preference to partial redundancy. Wealso discuss how the Sec24 secretion system is hijacked by viral (e.g., VSV-G, Hepatitis B envelope protein) and bacterial (e.g., the enteropathogenic E. coli type III secretion system effector NleA/EspI) pathogens.

Journal article

Chatterjee S, Lekmeechai S, Constantinou N, Grzybowska EA, Kozik Z, Choudhary JS, Berger CN, Frankel G, Clements Aet al., 2021, The type III secretion system effector EspO of enterohaemorrhagic <i>Escherichia coli</i> inhibits apoptosis through an interaction with HAX-1, CELLULAR MICROBIOLOGY, Vol: 23, ISSN: 1462-5814

Journal article

Chisenga CC, Bosomprah S, Simuyandi M, Mwila-Kazimbaya K, Chilyabanyama ON, Laban NM, Bialik A, Asato V, Meron-Sudai S, Frankel G, Cohen D, Chilengi Ret al., 2021, Shigella-specific antibodies in the first year of life among Zambian infants: A longitudinal cohort study, PLOS ONE, Vol: 16, ISSN: 1932-6203

Journal article

Mylona E, Sanchez Garrido J, Nguyen Hoang Thu T, Dongol S, Karkey A, Baker S, Shenoy AR, Frankel Get al., 2021, Very long O-antigen chains of Salmonella Paratyphi A inhibit inflammasome activation and pyroptotic cell death, Cellular Microbiology, Vol: 23, Pages: 1-14, ISSN: 1462-5814

Salmonella Paratyphi A (SPtA) remains one of the leading causes of enteric (typhoid) fever. Yet, despite the recent increased rate of isolation from patients in Asia, our understanding of its pathogenesis is incomplete. Here we investigated inflammasome activation in human macrophages infected with SPtA. We found that SPtA induces GSDMD‐mediated pyroptosis via activation of caspase‐1, caspase‐4 and caspase‐8. Although we observed no cell death in the absence of a functional Salmonella pathogenicity island‐1 (SPI‐1) injectisome, HilA‐mediated overexpression of the SPI‐1 regulon enhances pyroptosis. SPtA expresses FepE, an LPS O‐antigen length regulator, which induces the production of very long O‐antigen chains. Using a ΔfepE mutant we established that the very long O‐antigen chains interfere with bacterial interactions with epithelial cells and impair inflammasome‐mediated macrophage cell death. Salmonella Typhimurium (STm) serovar has a lower FepE expression than SPtA, and triggers higher pyroptosis, conversely, increasing FepE expression in STm reduced pyroptosis. These results suggest that differential expression of FepE results in serovar‐specific inflammasome modulation, which mirrors the pro‐ and anti‐inflammatory strategies employed by STm and SPtA, respectively. Our studies point towards distinct mechanisms of virulence of SPtA, whereby it attenuates inflammasome‐mediated detection through the elaboration of very long LPS O‐polysaccharides.

Journal article

Ruano-Gallego D, Sanchez-Garrido J, Kozik Z, Nunez-Berrueco E, Cepeda-Molero M, Mullineaux-Sanders C, Naemi-Baghshomali Clark J, Slater SL, Wagner N, Glegola-Madejska I, Roumeliotis T, Pupko T, Angel Fernandez L, Rodriguez-Paton A, Choudhary JS, Frankel Get al., 2021, Type III secretion system effectors form robust and flexible intracellular virulence networks, Science, Vol: 371, Pages: 1-21, ISSN: 0036-8075

Infections with many Gram-negative pathogens, including Escherichia coli, Salmonella, Shigella, and Yersinia, rely on type III secretion system (T3SS) effectors. We hypothesized that while hijacking processes within mammalian cells, the effectors operate as a robust network that can tolerate substantial contractions. This was tested in vivo using the mouse pathogen Citrobacter rodentium (encoding 31 effectors). Sequential gene deletions showed that effector essentiality for infection was context dependent and that the network could tolerate 60% contraction while maintaining pathogenicity. Despite inducing very different colonic cytokine profiles (e.g., interleukin-22, interleukin-17, interferon-γ, or granulocyte-macrophage colony-stimulating factor), different networks induced protective immunity. Using data from >100 distinct mutant combinations, we built and trained a machine learning model able to predict colonization outcomes, which were confirmed experimentally. Furthermore, reproducing the human-restricted enteropathogenic E. coli effector repertoire in C. rodentium was not sufficient for efficient colonization, which implicates effector networks in host adaptation. These results unveil the extreme robustness of both T3SS effector networks and host responses.

Journal article

Zheng W, Peña A, Ilangovan A, Clark JN-B, Frankel G, Egelman EH, Costa TRDet al., 2021, Cryoelectron-microscopy structure of the enteropathogenic Escherichia coli type III secretion system EspA filament, Proceedings of the National Academy of Sciences of USA, Vol: 118, ISSN: 0027-8424

Enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic Escherichia coli (EHEC) utilize a macromolecular type III secretion system (T3SS) to inject effector proteins into eukaryotic cells. This apparatus spans the inner and outer bacterial membranes and includes a helical needle protruding into the extracellular space. Thus far observed only in EPEC and EHEC and not found in other pathogenic Gram-negative bacteria that have a T3SS is an additional helical filament made by the EspA protein that forms a long extension to the needle, mediating both attachment to eukaryotic cells and transport of effector proteins through the intestinal mucus layer. Here, we present the structure of the EspA filament from EPEC at 3.4 Å resolution. The structure reveals that the EspA filament is a right-handed 1-start helical assembly with a conserved lumen architecture with respect to the needle to ensure the seamless transport of unfolded cargos en route to the target cell. This functional conservation is despite the fact that there is little apparent overall conservation at the level of sequence or structure with the needle. We also unveil the molecular details of the immunodominant EspA epitope that can now be exploited for the rational design of epitope display systems.

Journal article

Hopkins EGD, Frankel G, 2021, Overview of the Effect of <i>Citrobacter rodentium</i> Infection on Host Metabolism and the Microbiota, SHIGA TOXIN-PRODUCING E. COLI, Vol: 2291, Pages: 399-418, ISSN: 1064-3745

Journal article

Zheng W, Pena A, Low WW, Wong JLC, Frankel G, Egelman EHet al., 2020, Cryoelectron-Microscopic Structure of the pKpQIL Conjugative Pili from Carbapenem-Resistant <i>Klebsiella pneumoniae</i>, STRUCTURE, Vol: 28, Pages: 1321-+, ISSN: 0969-2126

Journal article

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