Spliceosomal helicases DDX41/SACY-1 and PRP22/MOG-5 both contribute to proofreading against proximal 3′ splice site usage

Abstract

RNA helicases drive necessary rearrangements and ensure fidelity during the pre-mRNA splicing cycle. DEAD-box helicase DDX41 has been linked to human disease and has recently been shown to interact with DEAH-box helicase PRP22 in the spliceosomal C* complex, yet its function in splicing remains unknown. Depletion of DDX41 homolog SACY-1 from somatic cells has been previously shown to lead to changes in alternative 3′ splice site (3′ss) usage. Here, we show by transcriptomic analysis of published and novel data sets that SACY-1 perturbation causes a previously unreported pattern in alternative 3′ splicing in introns with pairs of 3′ splice sites ≤18 nt away from each other. We find that both SACY-1 depletion and the allele sacy-1(G533R) lead to a striking unidirectional increase in the usage of the proximal (upstream) 3′ss. We previously discovered a similar alternative splicing pattern between germline tissue and somatic tissue, in which there is a unidirectional increase in proximal 3′ss usage in the germline for ∼200 events; many of the somatic SACY-1 alternative 3′ splicing events overlap with these developmentally regulated events. We generated targeted mutant alleles of the Caenorhabditis elegans homolog of PRP22, mog-5, in the region of MOG-5 that is predicted to interact with SACY-1 based on the human C* structure. These viable alleles, and a mimic of the myelodysplastic syndrome-associated allele DDX41(R525H), all promote the usage of proximal alternative adjacent 3′ splice sites. We show that PRP22/MOG-5 and DDX41/SACY-1 have overlapping roles in proofreading the 3′ss.