30 sep. 2026
Katelynn R. Kazane et al. (BioRxiv)
Keywords
● human NK cells
● viral infection
● epigenetic regulator
● secretory pathway
Main Findings
Natural Killer (NK) cells are a critical component of the immune system controlling anti-viral and anti-tumor immunity. Naturally occurring deficiencies in human NK cells are rare, yet poorly understood. In this preprint article, the authors aimed at investigating the presence and function of gene variants associated with defective NK cell development and function in a human cohort. Through the assessment of NK cell specific tumor cell lysis, the authors identify 2 patients defective in tumor cell lysis, carrying gene variants in the epigenetic regulator KMT2D (Histone H3 Lysine 4 Methyltransferase 2D). The authors observed that PBMCs from these two patients showed a significant decrease in blood NK cells. Analysing another publicly available patient sequencing repository, the authors identify another novel variant in KMT2D. Using a previously developed CRSIPR/Cas9 system on primary human NK cells, the authors introduce the three genetic variants into the KMT2Dgene of healthy human NK cells. A functional characterization of these NK cells revealed a defective production and secretion of Interferon (IFN)-g, Perforin (PRF1) and Granzyme B (GZMB). The mode of simulation of the NK cells appear irrelevant to determine this effect as NK cell receptor stimulation Similarly to their observations, the authors fully recapitulate these findings in mice deficient in Kmt2d either in all cells and selectively in Ncr1-expressing cells (including NK cells and a some subsets of other innate lymphoid cells). Moreover, when challenged with mouse cytomegalovirus (MCMV) animals deficient in Kmt2d were severely impaired in their antiviral defence.
Using RNASeq analysis and Cut+Tag sequencing of H3K4me1 modified regions, the authors first identified a striking downregulation in KMT2D-dependent genes that are associated with the secretory granule and excretion pathway in mice and human NK cells. RAB3D, a critical element in the secretion of granules was significantly decreased in its expression and H3K4me1 methylation marks resulting in a decrease in RAB3D protein in KMT2D-deficient NK cells. Consequently, the author decided to delete RAB3D in human and murine NK cells to determine whether alteration in RAB3D would impact the secretory capacity of NK cells. Measuring the abundance of degranulation markers on the surface NK cells by flow cytometry or assessing the amounts of secreted GZMB and IFN-g via ELISA revealed that similar to a deficiency in KMT2D, an absence of RAB3D – its downstream target – negatively impacts the ability of NK cells to release their effector molecules.
Collectively, these findings identify 3 novel gene variants within KMT2D, an epigenetic regulator of genes controlling the secretory pathway in murine and human NK cells. Deficiency in KMT2D or its downstream targets results in a developmental and functional NK cell deficiency in humans and elevates the susceptibility towards viral infections in mice.
Limitations
- While the authors nicely connect their observations through multiple elegant approaches, one wonders whether the reported defects are fully restrained to NK cells. An assessment of immune cell functions in NCR-expressing ILCs and other secretory cells would be an appreciated addition to the preprint, particularly in the context of Kmt2ddeficient animals and their wild type littermates.
- The authors provide data using the murine MCMV model that NK cells-specific Kmt2d plays an important role. Would Kmt2d-deficiency in NK cells also detrimentally impact the response against tumors? A simple model to explore here would be the manifestation of B16F10 lung metastasis.
- The authors provide data demonstrating changes in the expression of 52 genes that are conserved across murine and human NK cells. Are there uniquely regulated gene expression modules within the species-specifically regulated genes that could indicate distinct modes of action leading to NK cell deficiency in humans?
- The authors report 3 variants of KMT2D. Each variant shares a distinct location on a unique exon. It would be supportive to their work to demonstrate the consequence of these locations on the expression, abundance, enzymatic activity or subcellular localization of KMT2D. This could inform whether a common or variable mode of action promotes the manifestation of KMT2D-deficiency.
Significance/Novelty
The authors provide a compelling report of new genetic variants and knock out systems that reveal a previously underappreciated role of epigenetic regulation in NK cell development and function. These findings are of great importance to better understand and design management strategies for NK cell deficiencies in humans. Moreover, the uncovered mechanism demonstrates that irrespective of the stimulatory input, epigenetic modulation of genes controlling the secretory pathway are essential in control NK cells function. Such insight could be of use to bypass cytokine or receptor-mediated NK cell stimulation and could open doors to promote NK cell function through specific regulation of histone methylation with KMT2D. This could serve helpful in the treatment of cancers.
Credit
Reviewed by Arthur Mortha as part of a cross-institutional journal club between the Icahn School of Medicine at Mount Sinai, the University of Oxford, the Karolinska Institute, the MD Anderson Cancer Center and the University of Toronto.
The author declares no conflict of interests in relation to their involvement in the review.