Loss of ZRSR2, frequently mutated in blood cancer, enhanced hematopoietic stem cell self-renewal.
Major Finding: Loss of ZRSR2, frequently mutated in blood cancer, enhanced hematopoietic stem cell self-renewal.
Concept: ZRSR2 loss led to impaired minor intron excision in genes such as LZTR1, promoting transformation.
Impact: This study shows that minor intron retention via mutation or dysregulated splicing can drive cancer.
Mutations in RNA splicing factors are prevalent in myelodysplastic syndromes (MDS), and patients with leukemia often harbor mutations in RNA splicing factor genes, including SF3B1, SRSF2, U2AF1, and ZRSR2, of which only ZRSR2 functions primarily in the minor spliceosome. Although the major spliceosome is responsible for the removal of more than 99% of introns, the minor spliceosome splices a small but evolutionarily conserved subset of introns. To investigate the role of ZRSR2 and minor intron splicing in the hematopoietic system, Inoue, Polaski, Taylor, and colleagues engineered mice with hematopoietic cell–specific Zrsr2 deletion and found that loss of Zrsr2 enhanced self-renewal of hematopoietic stem cells (HSC). RNA-sequencing analyses of bone marrow samples from two independent cohorts of patients with MDS (n = 18; n = 12) revealed that ZRSR2-mutant samples displayed impaired removal of minor introns. ZRSR2 was shown to bind minor intron–containing genes, whereas ZRSR2 loss led to intron retention, specifically of a subset of minor introns harboring branchpoints that were proximal to the 3′ splice site; had sequences similar to that of the minor U12 snRNA consensus sequence; and had a weak or absent polypyrimidine tract. To explore whether ZRSR2-mutant disease phenotypes could be attributed to dysregulated splicing of target genes, a CRISPR–Cas9-based knockout screen, encompassing genes that were differently spliced in ZRSR2-mutant MDS patient samples, was performed in hematopoietic cell lines. Genetic knockout was designed to mimic the predicted effect of nonsense-mediated decay upon intron retention, and cells were screened for transformation via cytokine-independent enrichment following cytokine depletion. LZTR1, a gene encoding a cullin-3 adaptor known to suppress RAS-related GTPases, was a hit in all cell lines, and restoration of LZTR1 in Zrsr2-knockout mice decreased HSC self-renewal. In humans, LZTR1 minor intron retention was observed in diverse disease contexts, including Noonan syndrome, schwannomatosis, and many cancer types. In summary, this work shows the importance of minor intron splicing in HSC self-renewal and reveals minor intron retention as a potential cancer driver.
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