Mechanisms Of Age-related Changes in Transcriptional Regulation in lympphocytes

赖氨酸特异性甲基转移酶2D （Lysine specific methyltransferase 2D, Kmt2d）催化组蛋白3赖氨酸4 （H3K4me1）的单甲基化，并通过调节Foxp3基因表达在T细胞生成调控中发挥关键作用。在这里，我们报道了Kmt2d在普通CD8+ T细胞存活中的作用。首先，我们发现，与野生型（Kmt2dfl/flCD4cre-， WT）小鼠相比，缺乏Kmt2d （Kmt2dfl/flCD4cre-， KO）小鼠脾脏中CD8+ T细胞的数量，特别是天然CD8+ T细胞（CD62Lhi/CD44lo）的数量大大减少。其次，我们观察到体外刺激后中性CD8+ T细胞的死亡显著增加。第三，我们观察到增强子中H3K4me1水平降低，在缺乏Kmt2d的情况下，活化的中性CD8+ T细胞中凋亡相关基因的表达降低。最后，我们证实了Kmt2d KO小鼠在单核细胞原李斯特菌感染后体内活化诱导抗原特异性中性CD8+ T细胞死亡。这些发现表明，Kmt2d通过调节凋亡增强子中的H3K4me1水平和免疫相关基因表达来调节活化诱导的中性CD8+ T细胞存活。

Lysine specific methyltransferase 2D (Kmt2d) catalyzes the mono-methylation of histone 3 lysine 4 (H3K4me1) and plays a critical role in regulatory T cell generation via modulating Foxp3 gene expression. Here we report a role of Kmt2d in nave CD8+ T cell survival. First, we found that the number of CD8+ T cells, in particular nave CD8+ T cells (CD62Lhi/CD44lo), in spleen was greatly decreased in the absence of Kmt2d (Kmt2dfl/flCD4cre+, KO) compared to wild type (Kmt2dfl/flCD4cre-, WT) mice. Second, we observed significant increase in death of nave CD8+ T cells upon stimulation in vitro. Third, we observed reduced H3K4me1 level in enhancers reduced expressions of apoptosis-related genes in activated nave CD8+ T cells in the absence of Kmt2d. Finally, we confirmed the activation-induced death of antigen specific nave CD8+ T cells in vivo in Kmt2d KO mice upon challenge with Listeria monocytogen infection. These findings reveal that Kmt2d regulates activation induced nave CD8+ T cell survival via modulating H3K4me1 levels in enhancer of those apoptosis and immune-related gene expressions. CD8 T cells, the essential players in cell mediated immunity against infection and cancers, consist of six major subsets (nave, SCM, central and effector memory, EMRA, and effector) in blood defined by their phenotype and function. However, it is currently unknown how heterogeneous these subpopulations are, and what impact age has on the composition of CD8 T cell subpopulations in humans. Here, we used single cell RNA sequencing (scRNAseq) and multicolor immunophenotyping by flow cytometry to analyze the composition of CD8 T cells. From scRNAseq analysis of isolated CD8 T cells from 16 healthy donors, we analyzed 80,402 cells and identified a total of 10 subpopulations of CD8 T cells including nave, central and effector memory cells and terminally differentiated effector memory cells (EMRA). In parallel, our multi-color (16 antibody-panel) immunophenotyping analysis by flow cytometry of CD8 T cells from 165 donors categorized CD8 T cells into 10 subpopulations. Nine subpopulations identified by these two methods were equivalent based on the correlations of mRNA (UMI counts) and protein (MFI levels) expression. Importantly, the percentage of these nine subpopulations and their identified changes with age were comparable. Together, our findings reveal previously unidentified subpopulations of CD8 T cells and their changes with age. The functions of these newly identified CD8 T subpopulations and their role during aging are currently under investigation.