Fli-1 Overexpression in Hematopoietic Progenitors Deregulates T Cell Development and Induces Pre-T Cell Lymphoblastic Leukaemia/Lymphoma

Abstract

The Ets transcription factor Fli-1 is preferentially expressed in hematopoietic tissues and cells, including immature T cells, but the role of Fli-1 in T cell development has not been closely examined. To address this we retrovirally overexpressed Fli-1 in various in vitro and in vivo settings and analysed its effect on T cell development. We found that Fli-1 overexpression perturbed the DN to DP transition and inhibited CD4 development whilst enhancing CD8 development both in vitro and in vivo. Surprisingly, Fli-1 overexpression in vivo eventuated in development of pre-T cell lymphoblastic leukaemia/lymphoma (pre-T LBL). Known Fli-1 target genes such as the pro-survival Bcl-2 family members were not found to be upregulated. In contrast, we found increased NOTCH1 expression in all Fli-1 T cells and detected Notch1 mutations in all tumours. These data show a novel function for Fli-1 in T cell development and leukaemogenesis and provide a new mouse model of pre-T LBL to identify treatment options that target the Fli-1 and Notch1 signalling pathways.

DNMT3A Loss Drives Enhancer Hypomethylation in FLT3-ITD-Associated Leukemias

“…The comparison revealed overrepresentation of more than 80 transcription factors at hypomethylated enhancer sites, including master regulators that determine cell fate during stem, myeloid and lymphoid cell differentiation such as FLI1, GFI1B, and PU.1 (Figure S5B). Many of the transcription factors identified have also been implicated in hematopoietic diseases, including FLI1, LMO2, and RUNX1 in T-ALL (Cleveland et al, 2014; Mok et al, 2014; Smeets et al, 2013; Smith et al, 2014), and PU.1 and RUNX1 in AML (Cancer Genome Atlas Research, 2013; Gerloff et al, 2015). These findings suggest that Dnmt3a loss results in the demethylation of previously methylated, functionally relevant enhancers during leukemogenesis, potentially increasing accessibility of these regions to transcription factors.…”

“…To test this, we utilized the T-ALL model because of the homogeneity of this leukemia and the complete absence of Dnmt3a expression. In addition to being implicated in T cell development and T-ALL (Smeets et al, 2013; Smeets et al, 2014), FLI1 exhibited robust gene expression across conditions, was not differentially expressed, and the FLI1 consensus motif was enriched in hypomethylated enhancer sequences (p = 3.59e-15, Bonferroni correction). To test the effect of re-methylation on FLI1 binding, we performed ChIP-seq for FLI1 in 3aKO/FLT3-ITD leukemic cells transduced with either the DNMT3A-expressing retrovirus, or that of GFP alone, identifying 6,492 FLI1 binding sites.…”

The Rothmund-Thomson syndrome helicase RECQL4 is essential for hematopoiesis

“…For T cell cultures, 20,000 OP9-DL1 cells (42) were plated in a 24-well plate 48 hours before infected LKS + cells were added. T cell cultures were performed in α-MEM (Sigma-Aldrich) with 20% FBS (PAA Gold), P/S, GlutaMAX, 5 × 10 -5 M 2-mercaptoethanol (Sigma-Aldrich), 5 ng/ml Flt3L (PeproTech), 0.25 ng/ml mIL-7 (R&D Systems), and 100 nM 4-OH tamoxifen (74,75). T cell development was analyzed by flow cytometry on days 7, 14, and 21 (n = 4 separate donors/genotype).…”

PHF6 regulates phenotypic plasticity through chromatin organization within lineage-specific genes

“…Conversely, regions that became more accessible upon Phf6 loss are enriched for motifs of transcription factors associated with the development of T-ALL and AML ( Fig. 4B, bottom; Eyquem et al 2004;Thoms et al 2011;Kiaii et al 2013;Smeets et al 2013). Thus, genomic regions important for B and T lineage genes as well as their regulatory factor-binding sites undergo significant changes in chromatin accessibility upon complete loss of Phf6 in B-ALL cells.…”