Analysis of intestinal epithelial cell responses to IFN-γ during Cryptosporidiuminfection

Abstract Cryptosporidium species are intracellular parasites that infect intestinal epithelial cells (IEC) and the rapid cycle of growth, cell lysis and reinfection every 12 hours can cause severe enteric disease in young or immunocompromised patients. Interferon-γ (IFN-γ) has a crucial role in protective immunity and mice with an IEC-specific deletion of the IFN-γ receptor (IFN-γR ΔIEC) are highly susceptible to infection. However, the dynamics of IFN-γ signalling to IEC and how this leads to parasite control remain poorly understood. Based on the use of a reporter mouse for IFN signalling in vivo, while macrophages showed a transient response to treatment with IFN-γ, IEC exhibited a sustained response to IFN-γ that peaks at 24h. Indeed, the treatment of mice with recombinant IFN-γ transiently limits parasite shedding but takes 24h to show protective activity. Single-cell RNA sequencing of enterocytes from naïve and infected mice identified upregulation of several IFN-γ-inducible effectors but highlighted that induction of these genes was comparable in infected and bystander IEC. Together, these results suggest that while Cryptosporidium-infected cells are responsive to IFN-γ signalling, the protective effects of IFN-γ may be mediated through activation of uninfected bystander enterocytes. These studies provide new insights into the dynamics and function of IFN-γ signalling on IEC that are broadly relevant to other IEC-restricted pathogens. Supported by grants from NIH (R01 AI148249) and post-doctoral fellowships from the Canadian Institutes for Health Research and Fonds de Recherche du Québec – Santé.