Vir1p, the Yeast Homolog of Virilizer, is Required for mRNA m6A Methylation and Meiosis

Abstract

AbstractN6-Methyladenosine (m6A) is one of the most abundant modifications found on eukaryotic mRNAs. mRNA methylation regulates a host of biological processes including meiosis, a specialized diploid cell division program that results in the formation of haploid cells (gametes). During budding yeast meiosis, m6A levels peak early, before the initiation of the meiotic divisions. High-throughput studies and work from our lab showed that Ygl036wp, a previously uncharacterized protein interacts with Kar4p, a meiotic protein required for mRNA m6A-methylation. Ygl036wp has no discernable domains except for several intrinsically disordered regions. However, protein folding prediction tools showed that Ygl036wp folds like VIRMA/Virilizer/VIR, which is involved in mRNA m6A-methylation in higher eukaryotes. In addition, Ygl036wp has several conserved motifs shared with VIRMA/Virilizer/VIR proteins. Accordingly, we propose to call the geneVIR1forbudding yeast ortholog ofVIRMA/Virilizer/VIR1. In support, Vir1p interacts with all other members of the yeast methyltransferase complex and is required for mRNA m6A methylation and meiosis. Vir1p is required for the stability of proteins comprising the methyltransferase complex, suggesting that Vir1p acts as a scaffold to stabilize the complex. Thevir1Δ/Δ mutant is defective for premeiotic S-phase, which is suppressed by overexpression of the early meiotic transcription factorIME1;additional overexpression of the translational regulatorRIM4is required for sporulation.Consistent withIME1suppression,vir1Δ/Δ exhibits a defect in the abundance ofIME1mRNA, as well as transcripts within Ime1p’s regulon. Suppression byIME1revealed a defect in the expression of the middle meiotic transcription factor, Ndt80p (and genes in its regulon), which is rescued by additional overexpression ofRIM4. Together, these data suggest that Vir1p is required for cells to initiate the meiotic program and for progression through the meiotic divisions and spore formation.Author SummaryYgl036wp is a previously uncharacterized protein that we propose to name Vir1p (budding yeast ortholog ofVIRMA/Virilizer/VIR1). Work from our lab and others initially found an interaction between Vir1p and members of the yeast mRNA methyltransferase complex (Kar4p and Mum2p). We found that Vir1p interacts with all known members of the methyltransferase complex and is required for mRNA methylation. Vir1p is required early in meiosis;vir1Δ/Δ mutants arrest due to the reduced expression of Ime1p. Lower levels of Ime1p cause severe disruption to the meiotic transcriptome invir1Δ/Δ. Thevir1Δ/Δ meiotic defect can be partially suppressed by the overexpression ofIME1; full suppression requires overexpression of bothIME1andRIM4. Using recent advances in protein folding predictions, we found that Vir1p is a remote homolog of VIRMA/Virilizer/VIR and shares conserved motifs with the protein from other organisms. Vir1p, like VIRMA/Virilizer/VIR, stabilizes the methyltransferase complex.