Ascorbic acid promotes KIR demethylation during early NK cell differentiation

Wu Cheng-Ying, Bin Zhang, Hansol Kim, Stephen K. Anderson, Jeffrey S. Miller, Frank Cichocki

Research output: Contribution to journalArticlepeer-review

Abstract

Variegated expression of killer Ig-like receptors (KIR) in human NK cells is a stochastic process exclusive to subsets of mature NK cells and CD8+ T cells. Allele-specific KIR expression is maintained by DNA methylation within the proximal promoter regions. Because KIR genes are densely methylated in NK cell progenitors, there is an implied stage of human NK cell development in which DNA demethylation takes place to allow for active transcription. When and how this process occurs is unknown. In this study, we show that KIR proximal promoters are densely methylated in less mature CD56bright NK cells and are progressively demethylated in CD56dim NK cells as they mature and acquire KIR. We hypothesized that ten-eleven translocation (TET) enzymes, which oxidize 5mC on DNA could mediate KIR promoter demethylation. The catalytic efficiency of TET enzymes is known to be enhanced by ascorbic acid. We found that the addition of ascorbic acid to ex vivo culture of sorted CD56bright NK cells increased the frequency of KIR expression in a dose-dependent manner and facilitated demethylation of proximal promoters. A marked enrichment of the transcription factor Runx3 as well as TET2 and TET3 was observed within proximal KIR promoters in CD56bright NK cells cultured with ascorbic acid. Additionally, overexpression of TET3 and Runx3 promoted KIR expression in CD56bright NK cells and NK-92 cells. Our results show that KIR promoter demethylation can be induced in CD56bright, and this process is facilitated by ascorbic acid. The Journal of Immunology, 2020, 205: 1513–1523.

Original languageEnglish (US)
Pages (from-to)1513-1523
Number of pages11
JournalJournal of Immunology
Volume205
Issue number6
DOIs
StatePublished - Sep 15 2020

Bibliographical note

Funding Information:
This work was supported by National Heart, Lung, and Blood Institute/National Institutes of Health (NIH) K99/R00 HL123638 (to F.C.) and National Cancer Institute/NIH P01 CA111412 (to J.S.M.), P01 CA65493 (to J.S.M.), and R35 CA197292 (to J.S.M.). This project has been funded in whole or in part with federal funds from the Frederick National Laboratory for Cancer Research/NIH, under contract HHSN261200800001E. This research was supported in part by the Intramural Research Program of the NIH/Frederick National Lab/Center for Cancer Research.

Funding Information:
This work was supported by National Heart, Lung, and Blood Institute/National Institutes of Health (NIH) K99/R00 HL123638 (to F.C.) and National Cancer Institute/NIH P01 CA111412 (to J.S.M.), P01 CA65493 (to J.S.M.), and R35 CA197292 (to J.S.M.). This project has been funded in whole or in part with federal funds from the Frederick National Laboratory for Cancer Research/NIH, under contract HHSN261200800001E. This research was supported in part by the Intramural Research Program of the NIH/Frederick National Lab/Center for Cancer Research. We thank the University of Minnesota Genomics Center, the University of Minnesota Informatics Institute, and the University of Minnesota Flow Cytometry Core for their services.

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