Mutations affecting spliceosomal proteins are frequently found in hematological malignancies, including myelodysplastic syndromes and acute myeloid leukemia (AML). DDX41/Abstrakt is a metazoan-specific spliceosomal DEAD-box RNA helicase that is recurrently mutated in inherited myelodysplastic syndromes and in relapsing cases of AML. The genetic properties and genomic impacts of disease-causing missense mutations in DDX41 and other spliceosomal proteins have been uncertain. Here, we conduct a comprehensive analysis of the Caenorhabditis elegans DDX41 ortholog, SACY-1. Biochemical analyses defined SACY-1 as a component of the C. elegans spliceosome, and genetic analyses revealed synthetic lethal interactions with spliceosomal components. We used the auxin-inducible degradation system to analyze the consequence of SACY-1 depletion on the transcriptome using RNA sequencing. SACY-1 depletion impacts the transcriptome through splicing-dependent and splicing-independent mechanisms. Altered 39 splice site usage represents the predominant splicing defect observed upon SACY-1 depletion, consistent with a role for SACY-1 in the second step of splicing. Missplicing events appear more prevalent in the soma than the germline, suggesting that surveillance mechanisms protect the germline from aberrant splicing. The transcriptome changes observed after SACY-1 depletion suggest that disruption of the spliceosome induces a stress response, which could contribute to the cellular phenotypes conferred by sacy-1 mutant alleles. Multiple sacy-1/ddx41 missense mutations, including the R525H human oncogenic variant, confer antimorphic activity, suggesting that their incorporation into the spliceosome is detrimental. Antagonistic variants that perturb the function of the spliceosome may be relevant to the disease-causing mutations, including DDX41, affecting highly conserved components of the spliceosome in humans.
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This paper is dedicated to the memory of Michael A. Miller, our colleague and friend whose scientific contributions will not be forgotten. We thank Donna Coetzee for technical assistance. We are grateful to Joshua Arribere, Daniel Dickinson, Barth Grant, Tim Schedl, and Jordan Ward for providing strains or reagents. Some strains were provided by the Caenorhabditis Genetics Center, which is funded by grant P40OD010440 from the National Institutes of Health (NIH) Office of Research Infrastructure Programs. We thank Gabriela Huelgas-Morales, Zohar Sachs, and Todd Starich for their helpful suggestions for the manuscript. This work was supported by NIH grant GM57173 to D.G.
© 2020 by the Genetics Society of America
- C. elegans
- Myelodysplastic disorders