Impact of spliceosome mutations on RNA splicing in myelodysplasia: dysregulated genes/pathways and clinical associations

Abstract

, , and are the most frequently mutated splicing factor genes in the myelodysplastic syndromes (MDS). We have performed a comprehensive and systematic analysis to determine the effect of these commonly mutated splicing factors on pre-mRNA splicing in the bone marrow stem/progenitor cells and in the erythroid and myeloid precursors in splicing factor mutant MDS. Using RNA-seq, we determined the aberrantly spliced genes and dysregulated pathways in CD34 cells of 84 patients with MDS. Splicing factor mutations result in different alterations in splicing and largely affect different genes, but these converge in common dysregulated pathways and cellular processes, focused on RNA splicing, protein synthesis, and mitochondrial dysfunction, suggesting common mechanisms of action in MDS. Many of these dysregulated pathways and cellular processes can be linked to the known disease pathophysiology associated with splicing factor mutations in MDS, whereas several others have not been previously associated with MDS, such as sirtuin signaling. We identified aberrantly spliced events associated with clinical variables, and isoforms that independently predict survival in MDS and implicate dysregulation of focal adhesion and extracellular exosomes as drivers of poor survival. Aberrantly spliced genes and dysregulated pathways were identified in the MDS-affected lineages in splicing factor mutant MDS. Functional studies demonstrated that knockdown of the mitosis regulators and aberrantly spliced target genes of and mutations, respectively, led to impaired erythroid cell growth and differentiation. This study illuminates the effect of the common spliceosome mutations on the MDS phenotype and provides novel insights into disease pathophysiology.

Distinct and convergent consequences of splice factor mutations in myelodysplastic syndromes

“…39 30 Liang et al also observed that the SRSF2 P95H mutation affected splicing of several members of the hnRNP and SR families of proteins. Importantly, EZH2 is located at chromosome 7q36, a region frequently deleted (−7 and 7q-) in MDS.…”

“…30 More recently, splicing changes and transcriptional consequences of abnormal RNA splicing were also investigated in a large cohort of MDS samples. 30 More recently, splicing changes and transcriptional consequences of abnormal RNA splicing were also investigated in a large cohort of MDS samples.…”

Mutations in spliceosome genes and therapeutic opportunities in myeloid malignancies

“…In addition, the mechanistic basis of how splicing factor mutations confer selective advantage during the evolution of MDS pathogenesis remains a mystery. Although recent large‐scale transcriptomic analysis of mutations affecting SF3B1 , SRSF2 , and U2AF1 revealed global splicing dysregulation consistent with their predicted effects, there was minimal overlap between mis‐spliced transcripts altered by each of the mutant proteins . To identify key splicing abnormalities induced by mutant splicing factors, several studies provided functional evidence that expression of the abnormal splice isoforms in normal cells, or reintroduction of the canonical isoforms in spliceosomal mutant cells, can only partially phenocopy or rescue the effects of mutant spliceosome proteins .…”

“…These findings suggest that the pathological effects of spliceosome mutations are imparted through multiple mis‐spliced products simultaneously. In parallel, pathway analyses followed by functional validation revealed that splicing factor mutations distinctly converge on key cellular pathways to drive disease pathogenesis, including defects in DNA damage response and aberrant innate immune signaling . Further studies will be required to systematically address these fundamental questions, and more importantly, to identify the key players and pathways that can potentially be exploited for therapeutic purposes.…”

“…73 Although recent large-scale transcriptomic analysis of mutations affecting SF3B1, SRSF2, and U2AF1 revealed global splicing dysregulation consistent with their predicted effects, there was minimal overlap between mis-spliced transcripts altered by each of the mutant proteins. 93,94 To identify key splicing abnormalities induced by mutant splicing factors, several studies provided functional evidence that expression of the abnormal splice isoforms in normal cells, or reintroduction of the canonical isoforms in spliceosomal mutant cells, can only partially phenocopy or rescue the effects of mutant spliceosome proteins. 72,80,95,96 These findings suggest that the pathological effects of spliceosome mutations are imparted through multiple mis-spliced products simultaneously.…”