MPEG1/perforin-2 mutations in human pulmonary nontuberculous mycobacterial infections
Ryan M. McCormack, Eva P. Szymanski, Amy P. Hsu, Elena Perez, Kenneth N. Olivier, Eva Fisher, E. Brook Goodhew, Eckhard R. Podack, Steven M. Holland
Ryan M. McCormack, Eva P. Szymanski, Amy P. Hsu, Elena Perez, Kenneth N. Olivier, Eva Fisher, E. Brook Goodhew, Eckhard R. Podack, Steven M. Holland
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Research Article Genetics Infectious disease

MPEG1/perforin-2 mutations in human pulmonary nontuberculous mycobacterial infections

Abstract

Perforin-2 is a highly conserved pore-forming protein encoded by macrophage expressed gene 1 (MPEG1). A number of studies have shown that Perforin-2–deficient mice are unable to survive following a bacterial challenge that is nonlethal in WT mice. There is also recent evidence that Mpeg1+/– heterozygous mice display an intermediate killing ability compared with Mpeg1 WT and Mpeg1–/– mice. Despite these in vivo findings, to date, no perforin-2 deficiencies have been associated with human disease. Here, we report four patients with persistent nontuberculous mycobacterial infection who had heterozygous MPEG1 mutations. In vitro, neutrophils, macrophages, and B cells from these patients were unable to kill Mycobacterium avium as efficiently as normal controls. CRISPR mutagenesis validated the deleterious antibacterial activity of these mutations. These data suggest that perforin-2 haploinsufficiency may contribute to human susceptibility to infections with intracellular bacteria.

Authors

Ryan M. McCormack, Eva P. Szymanski, Amy P. Hsu, Elena Perez, Kenneth N. Olivier, Eva Fisher, E. Brook Goodhew, Eckhard R. Podack, Steven M. Holland

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Figure 3

Cell lines with MPEG1/perforin-2 mutations are unable to control Mycobacterium smegmatis infection.

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Cell lines with MPEG1/perforin-2 mutations are unable to control Mycobac...
Patient MPEG1 mutations were introduced via CRISPR/Cas9 into THP1 cell lines. WT and mutant cell lines were infected with M. smegmatis, and killing ability was assessed. The WT cell line was able to significantly decrease M. smegmatis CFU over time, p.P316S decreased CFU to a lesser extent, and the remaining mutant cell lines were unable to decrease CFU. The experiment was done with five biological replicates and two technical duplicates per replicate, and it was repeated four times. Statistical analysis was conducted by one-way ANOVA with Tukey post-hoc test. *P < 0.05. Detailed statistical breakdown is described in Supplemental Table 6.