Mutations in SRSF2 and U2AF35 trigger replication stress and ATR activation via R-loop formation.

  • Major finding: Mutations in SRSF2 and U2AF35 trigger replication stress and ATR activation via R-loop formation.

  • Mechanism: Mutant SRSF2 promotes R-loop formation by impairing transcription pause release.

  • Impact: Resolving R-loops may partially correct splicing factor mutation growth defects in blood progenitors.

Multiple mutations in general splicing factors are associated with myelodysplastic syndrome (MDS) and various solid tumor types, but different mutations cause distinct splicing defects, suggesting that they may cause disease by a common alternative mechanism. Chen, Chen, Huang, and colleagues investigated this possibility via mutations in the splicing factors SRSF2 and U2AF35 that confer high risk of MDS and exhibited little overlap in induced splicing changes. Despite distinct effects on splicing, SRSF2 and U2AF35 mutations each increased γ-H2AX signals, inducing G2/M arrest, thus suggesting the presence of replication stress. Indeed, SRSF2 and U2AF35 mutations produced replication stress and activated the ATR–CHK1 pathway, but not the ATM pathway. SRSF2 and U2AF35 mutants induced the formation of R-loops (RNA:DNA hybrids), whereas the wild-type alleles did not, and excessive R-loops have been shown to trigger DNA damage at the entry to S phase, suggesting a mechanism for the induction of replication stress and ATR activity. The splicing factor mutations induced R-loop formation by preventing SRSF2 from releasing P-TEFb, a PolII Ser2 kinase, from the 7SK complex, thereby impairing transcription pause release at transcription start sites, leading to elevated R-loop formatin. Consistent with these findings, resolving R-loops could partially rescue the cell growth defects induced by SRSF2 or U2AF35 mutations and reduced ATR pathway activation. In vivo, R-loops were detected in the bone marrow of mice with MDS induced by Srsf2 mutation, and resolving the R-loops partially rescued the proliferation defect in blood progenitor cells derived from these mice. Collectively, these findings suggest that splicing factor mutations may promote MDS in part through the induction of R-loops.

Chen L, Chen JY, Huang YJ, Gu Y, Qiu J, Qian H, et al. The augmented R-loop Is a unifying mechanism for myelodysplastic syndromes induced by high-risk splicing factor mutations. Mol Cell 2018;69:412–25.

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