GATA2-TAL1 Axis Is Essential for Human NK Cell Development and Function

Introduction: Natural killer (NK) cells, as the major subset of innate lymphocytes, produce proinflammatory cytokines and mediate anti-tumor cytotoxicity (1-3). Since NK cells are not limited by clonotypic receptors, they can be utilized in cell therapies against a broad spectrum of malignancies. Irrespective of the clinical potentials, the transcriptional regulation of the development and functions of human NK cells is far from fully understood. A dysregulated NK cell development is a hallmark of GATA2 haploinsufficiency (4)(eg. GATA2^T354M mutation). The specific loss of the CD56^bright NK cell population is a significant striking feature in GATA2-deficient patients (4), with or without reduced total CD56^dim NK cells (4, 12, 13). Consequently, the effector functions of these NK cells are significantly reduced, including inflammatory cytokine production and anti-tumor cytotoxicity (4, 12). However, a mechanistic understanding of GATA2 functions is lacking.

In humans, GATA2 forms a core heptad complex with six other transcription factors (TF), including TAL1, regulating more than 1000 target genes. However, the unique functions of the GATA2-TAL1 axis in immature and mature NK cell has not been established. Based on these, we hypothesize that GATA2-dependent and independent functions of TAL1 play an essential role in the development and functions of human NK cells.

Results: To explore the mechanism of co-transcriptional regulation of GATA2 and TAL1, we transduced NK-92 cells with lentiviral vectors containing GATA2^WT or GATA2^T354M plasmid and quantified TAL1 mRNA levels by qPCR and protein levels by WB. TAL1 is upregulated in GATA2^T354M-expressing NK-92 cells both at mRNA (Fig. 1A) and protein levels (Fig. 1B). TAL1 is a TF activating GTPase of immunity-associated protein (GIMAP) family super-enhancer regions in HSC and T cells (11). As TAL1 target genes, we further quantify GIMAP mRNA levels by qPCR. The result indicates that genes from the GIMAP family were notably upregulated in GATA2^T354M-expressing NK-92 cells (Fig. 1C). Knocking down TAL1 results in the significant downregulation of TAL1 and GIMAPs in GATA2^T354M-expressing cells (Fig. 1D). To further validate the GATA2-TAL1 axis, sorted NK cells from PBMCs of three GATA2^T354M patients and five age-matched healthy controls (HC) were analyzed with single-cell RNA-seq. We identified the genes from GIMAP family were significantly upregulated in all three GATA2^T354M patients. The representative data from one patient is shown in Fig. 1E. To explore how GATA2 and TAL1 mediate transcriptional regulation, we performed GATA2 CUT&Tag experiments with human NK cells, in which TAL1 motif was predominately enriched in 63.2% of GATA2 targets among NK cells through HOMER motif enrichment analyses (Fig. 1F). On the other hand, the TAL1 ChIP-seq data from the K562 cells (download from ENCFF101DBG) reveals that the GATA2 motif is enriched in approximately 44% of TAL1 targets (Fig. 1G). Moreover, both GATA2 and TAL1 expression in NK cells are modified by H3K27Me3 instead of H3K4Me3, suggesting a repressive methylenation signature in their expression (Fig. 1H).

Conclusion: Based on this study, we uncover a novel role for the GATA2-TAL1 axis in human NK cells. we define the transcriptional regulation role of GATA2 and TAL1 complex and provide direct evidence for the mechanism of NK cell deficiency among GATA2^T354M patients. Importantly, through our current study, we will determine the unique role of TAL1 in NK development and function and its transcriptional network.