Genetic targeting of Card19 is linked to disrupted NINJ1 expression, impaired cell lysis, and increased susceptibility to Yersinia infection

Journal article

Cell death plays a critical role in inflammatory responses. During pyroptosis, inflammatory caspases cleave Gasdermin D (GSDMD) to release an N-terminal fragment that generates plasma membrane pores that mediate cell lysis and IL-1 cytokine release. Terminal cell lysis and IL-1β release following caspase activation can be uncoupled in certain cell types or in response to particular stimuli, a state termed hyperactivation. However, the factors and mechanisms that regulate terminal cell lysis downstream of GSDMD cleavage remain poorly understood. In the course of studies to define regulation of pyroptosis duringYersiniainfection, we identified a line ofCard19-deficient mice (Card19lxcn)whose macrophages were protected from cell lysis and showed reduced apoptosis and pyroptosis, yet had wild-type levels of caspase activation, IL-1 secretion, and GSDMD cleavage. Unexpectedly, CARD19, a mitochondrial CARD-containing protein, was not directly responsible for this, as an independently-generated CRISPR/Cas9Card19knockout mouse line (Card19Null) showed no defect in macrophage cell lysis. Notably,Card19is located on chromosome 13, immediately adjacent toNinj1, which was recently found to regulate cell lysis downstream of GSDMD activation. RNA-seq and western blotting revealed thatCard19lxcnBMDMs have significantly reduced NINJ1 expression, and reconstitution ofNinj1inCard19lxcnimmortalized BMDMs restored their ability to undergo cell lysis in response to caspase-dependent cell death stimuli.Card19lxcnmice exhibited increased susceptibility toYersiniainfection, whereas independently-generatedCard19Nullmice did not, demonstrating that cell lysis itself plays a key role in protection against bacterial infection, and that the increased infection susceptibility ofCard19lxcnmice is attributable to loss of NINJ1. Our findings identify genetic targeting ofCard19being responsible for off-target effects on the adjacent geneNinj1, disrupting the ability of macrophages to undergo plasma membrane rupture downstream of gasdermin cleavage and impacting host survival and bacterial control duringYersiniainfection.

Authors: Bjanes, E; Sillas, RG; Matsuda, R; Demarco, B; Fettrelet, T

• Publication status: Published
• Peer review status: Peer reviewed
• Publisher: Public Library of Science
• Journal: PLoS Pathogens
• Volume: 17
• Issue: 10
• Pages: e1009967-e1009967
• Publication date: 2021-10-14
• DOI: 10.1371/journal.ppat.1009967
• EISSN: 1553-7374
• ISSN: 1553-7366
• Language: English
• Keywords: Immunology; Biology; Genetics; Microbiology; Lysis; Bacteria; Yersinia