Project ID CM-HD2025_30

ThemeCM-HD

Co Supervisor 1A Dr Vincenzo Torraca Faculty of Life Sciences & Medicine, School of Immunology & Microbial Sciences, Department of Infectious DiseasesEmail

Co Supervisor 1B Dr Luis Apolonia Faculty of Life Sciences & Medicine, School of Immunology & Microbial Sciences, Department of Infectious DiseasesEmail

2025 Project: Use of human induced pluripotent stem cell-derived macrophages (iMACs) to study Shigella persistent infections

Basis of the project and translational impact
Shigella is an antimicrobial resistance priority pathogen, causing 260 million illnesses and 200,000 deaths yearly. The alternative lifestyles of Shigella have not been completely elucidated. While Shigella can be phagocytosed by macrophages and rapidly induce cell death, recent data showed that it can also persist for long time within these cells. How Shigella persists in macrophages is not fully understood and this knowledge is crucial for developing effective treatments that prevent establishment of persistent infections.

Aims
We aim to establish a Shigella persistent infection model in human inducible pluripotent stem cell-derived macrophages (iMACs) allowing to understand the mechanisms leading to persistence. We will perform longitudinal studies, coupled with RNAseq and high-resolution microscopy, to dissect how macrophages adapt during infection and become permissive to intracellular infections. We will also study the role of antibiotics in persistent infections, since these treatments can encourage the development of persistent bacteria. Finally, we will study the host factors involved, by performing infection studies in iMAC cells from diverse host genetic backgrounds and applying CRISPR-Cas9 mutagenesis/overexpression techniques to validate the role of the discovered factors.

Techniques and skills
This project will provide extensive training in tissue culture (iPSCs and other cell lines, induction and maintenance of iMACs), microbiology/infection (bacterial cultures, analysis of infection dynamics), microscopy, flow cytometry, and molecular biology (CRISPR-Cas9, RNAseq) techniques.

Objectives of each year
Year 1 – Learn iPSC and iMAC culture techniques. Establish the iMAC-Shigella infection model and define infection conditions using microscopy and flow cytometry.
Year 2 – Profile the transcriptome of persistently infected iMACs (RNAseq). Dissect the effect of different host-genetic backgrounds.
Year 3 – Use of genetic tools (CRISPR-Cas9 knockouts, overexpression) to study individual host factors based on RNAseq and host genetics.

Representative Publications

Torraca V, Brokatzky D, Miles SL, Chong CE, De Silva PM, Baker S, Jenkins C, Holt KE, Baker KS, Mostowy S. Shigella Serotypes Associated With Carriage in Humans Establish Persistent Infection in Zebrafish. J Infect Dis. 2023 Oct 18;228(8):1108-1118. doi: 10.1093/infdis/jiad326.

Torraca V, Kaforou M, Watson J, Duggan GM, Guerrero-Gutierrez H, Krokowski S, Hollinshead M, Clarke TB, Mostowy RJ, Tomlinson GS, Sancho-Shimizu V, Clements A, Mostowy S. Shigella sonnei Infection of Zebrafish Reveals that O-antigen Mediates Neutrophil Tolerance and Dysentery Incidence. PLoS Pathog. 2019 Dec 12;15(12):e1008006. doi: 10.1371/journal.ppat.1008006.

Torraca V, Holt K, Mostowy S. Shigella sonnei. Trends Microbiol. 2020 Aug;28(8):696-697. doi: 10.1016/j.tim.2020.02.011.

Betancor G, Dicks MDJ, Jimenez-Guardeño JM, Ali NH, Apolonia L, Malim MH. The GTPase Domain of MX2 Interacts with the HIV-1 Capsid, Enabling Its Short Isoform to Moderate Antiviral Restriction. Cell Rep. 2019 Nov 12;29(7):1923-1933.e3. doi: 10.1016/j.celrep.2019.10.009.

Jimenez-Guardeño JM, Apolonia L, Betancor G, Malim MH. Immunoproteasome Activation Enables Human TRIM5? Restriction of HIV-1. Nat Microbiol. 2019 Jun;4(6):933-940. doi: 10.1038/s41564-019-0402-0.

Doyle T, Moncorge O, Bonaventure B, Pollpeter D, Lussignol M, Tauziet M, Apolonia L, Catanese MT, Goujon C, Malim MH. The Interferon-inducible Isoform of NCOA7 Inhibits Endosome-mediated Viral Entry. Nat Microbiol. 2018 Dec; 3(12):1369-1376. doi: 10.1038/s41564-018-0273-9.