NKCLR3 impairs natural killer cell cytotoxicity by epigenetically targeting the Natural Cytotoxicity Triggering Receptor 1 (NCR1) [lncRNA-seq]

Natural killer cells (NK cells) play a critical role in the surveillance of tumor immunity. However, NK cell-based immunotherapy, including autologous and allogeneic NK cell reinfusion, has not brought significant clinical benefits to patients. To identify factors that control the intrinsic cytotoxicity of NK cells, we utilized the histone deacetylase inhibitor valproic acid (VPA) to develop a NK cell cytotoxicity suppression model. With RNA-seq and functional assays, we identified a previously uncharacterized lncRNA, NKCLR3 (NK cytotoxicity-associated long noncoding RNA 3), as a negative regulatory factor of NK cell-mediated cytotoxicity. NKCLR3 was significantly upregulated in VPA-treated NK cells and was negatively associated with the cytotoxicity of NK cells. Knockdown of NKCLR3 enhanced antitumor activity in NK-92MI cells. Using reverse transcription-associated capture sequencing (RAT-seq), we found that NKCLR3 functioned by targeting the Natural cytotoxicity triggering receptor 1 (NCR1) gene, which encodes the activating receptor NKp46 involved in the natural cytotoxicity. Mechanistic studies revealed that NKCLR3 interacted with the regulatory elements of NCR1 and blocked the formation of an intrachromosomal interaction that is required for optimal expression of NCR1. In addition, NKCLR3 inhibited the synthesis of NCR1 enhancer RNA. Through these dual mechanisms, NKCLR3 induced a suppressive epigenotype in the NCR1 promoter and suppressed the expression of the NCR1 gene. Thus, the NKCLR3-NCR1 axis identified in this study may serve as a novel target to improve therapeutic intervention of NK cells in tumor immunotherapy. Overall design: To identify factors that control the intrinsic cytotoxicity of NK cells, we utilized the histone deacetylase inhibitor valproic acid (VPA) to develop a NK cell cytotoxicity suppression model. With RNA-seq and functional assays, we identified a previously uncharacterized lncRNA, NKCLR3 (NK cytotoxicity-associated long noncoding RNA 3), as a negative regulatory factor of NK cell-mediated cytotoxicity. NKCLR3 was significantly upregulated in VPA-treated NK cells and was negatively associated with the cytotoxicity of NK cells.