Single-cell CRISPR screening characterizes transcriptional deregulation in T-cell acute lymphoblastic leukemia

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive leukemia caused by accumulation of genetic alterations in T-cell progenitors. For many genes it remains unknown how loss-of-function mutations contribute to disease development. Single-cell CRISPR screening in ex vivo pro-T cells allowed us to study the transcriptomic impact of such alterations. A first CRISPR screen targeted 17 well-studied regulators and defined the core transcriptional signatures, such as NOTCH1, MYC, STAT5 and E2F. Additionally, Spi1 was identified as an essential gene associated with the MYC regulon. A second screen targeted 42 poorly characterized genes and identified gene clusters with E2F/MYC and STAT/NOTCH1 signatures having opposing roles. Bcl11b inactivation conferred the strongest growth advantage and was associated with JAK/STAT upregulation, corresponding with publicly available T-ALL patient data. Bcl11b inactivation cooperated with mutant JAK3 in transforming pro-T cells to cytokine-independent growth. With this data, we characterized tumor suppressors and oncogenes in T-ALL, thereby providing insight in the mechanisms of leukemia development. ex-vivo murine ProT cells were transduced with a CRISPR library and analysed using the 10x genomics feature-barcoding technology (capture of both mRNA and CRISPR guides)