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NK cell defects in X-linked pigmentary reticulate disorder
Petro Starokadomskyy, … , Daniel D. Billadeau, Ezra Burstein
Petro Starokadomskyy, … , Daniel D. Billadeau, Ezra Burstein
Published November 1, 2019
Citation Information: JCI Insight. 2019;4(21):e125688. https://doi.org/10.1172/jci.insight.125688.
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Research Article Infectious disease Inflammation

NK cell defects in X-linked pigmentary reticulate disorder

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Abstract

X-linked reticulate pigmentary disorder (XLPDR, Mendelian Inheritance in Man #301220) is a rare syndrome characterized by recurrent infections and sterile multiorgan inflammation. The syndrome is caused by an intronic mutation in POLA1, the gene encoding the catalytic subunit of DNA polymerase-α (Pol-α), which is responsible for Okazaki fragment synthesis during DNA replication. Reduced POLA1 expression in this condition triggers spontaneous type I interferon expression, which can be linked to the autoinflammatory manifestations of the disease. However, the history of recurrent infections in this syndrome is as yet unexplained. Here we report that patients with XLPDR have reduced NK cell cytotoxic activity and decreased numbers of NK cells, particularly differentiated, stage V, cells (CD3–CD56dim). This phenotype is reminiscent of hypomorphic mutations in MCM4, which encodes a component of the minichromosome maintenance (MCM) helicase complex that is functionally linked to Pol-α during the DNA replication process. We find that POLA1 deficiency leads to MCM4 depletion and that both can impair NK cell natural cytotoxicity and show that this is due to a defect in lytic granule polarization. Altogether, our study provides mechanistic connections between Pol-α and the MCM complex and demonstrates their relevance in NK cell function.

Authors

Petro Starokadomskyy, Katelynn M. Wilton, Konrad Krzewski, Adam Lopez, Luis Sifuentes-Dominguez, Brittany Overlee, Qing Chen, Ann Ray, Aleksandra Gil-Krzewska, Mary Peterson, Lisa N. Kinch, Luis Rohena, Eyal Grunebaum, Andrew R. Zinn, Nick V. Grishin, Daniel D. Billadeau, Ezra Burstein

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Figure 2

Pol-α/primase and the MCM complex are required for optimal NK cell function.

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Pol-α/primase and the MCM complex are required for optimal NK cell funct...
(A) Expression of MCM4 as determined by RNA-Seq analysis in unaffected (UA1) and XLPDR-derived immortalized dermal fibroblasts (XLPDR, P2 and P3). Bars represent the mean; error bars represent the SEM; *P < 0.05 by Student’s 1-tailed t test. Data are the average of 3 independent experiments. RPKM, reads per kilobase of transcript per million mapped reads. (B) Same as A, but comparing MCM4 expression in UA1 fibroblasts treated with siCtrl or siPOLA1. Bars represent the mean; error bars represent the SEM. *P < 0.05 by Student’s 1-tailed t test. Data are the average of 2 independent experiments. (C) Expression of the indicated proteins was determined by immunoblotting in immortalized dermal fibroblasts derived from an XLPDR patient (P3 + EV), isogenic “rescued” control line (P3 + POLA1), and a patient (P6) with a recently reported POLA1 mutation (c.328G>A). The Western blot is representative of 2 independent experiments. (D) HEK293T cell lysate was subjected to POLA1 or PRIM2 immunoprecipitation and then immunoblotted for MCM4. Nonspecific control antibody (IgG) was used as a negative control. The Western blot is representative of 2 independent experiments. (E) NK cell lines NK29mi (left) and YTS (right) were subjected to POLA1 or MCM4 silencing using corresponding siRNA. NK cell direct cytotoxicity against the 721.221 target cell line was determined over the indicated E/T ratios. Data represent the average of 3 experiments. Error bars represent the SEM. *P < 0.0001 by 2-way ANOVA comparing siPOLA1 or siMCM4 against siCtrl in each E/T ratio. Data are from a single experiment.

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