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Although lack of ADAR (adenosine deaminase acting on RNA) orthologs, genome-wide A-to-I editing occurs specifically during sexual reproduction in a number of filamentous ascomycetes, includingFusarium graminearumandNeurospora crassa. Unlike ADAR-mediated editing in animals, fungal A-to-I editing has a strong preference for hairpin loops and U at −1 position, which leads to frequent editing of UAG and UAA stop codons. Majority of RNA editing events in fungi are in the coding region and cause amino acid changes. Some of these editing events have been experimentally characterized for providing heterozygote and adaptive advantages inF.graminearum. Recent studies showed that FgTad2 and FgTad3, 2 ADAT (adenosine deaminase acting on tRNA) enzymes that normally catalyze the editing of A34 in the anticodon of tRNA during vegetative growth mediate A-to-I mRNA editing during sexual reproduction. Stage specificity of RNA editing is conferred by stage-specific expression of short transcript isoforms ofFgTAD2andFgTAD3as well as cofactors such asAME1andFIP5that facilitate the editing of mRNA in perithecia. Taken together, fungal A-to-I RNA editing during sexual reproduction is catalyzed by ADATs and it has the same sequence and structural preferences with editing of A34 in tRNA. 

A-to-I RNA editing catalyzed by adenosine-deaminase-acting-on-RNA (ADARs) was assumed to be unique to metazoans because fungi and plants lack ADAR homologs. However, genome-wide messenger RNA (mRNA) editing was found to occur specifically during sexual reproduction in filamentous ascomycetes. Because systematic characterization of adenosine/cytosine deaminase genes has implicated the involvement ofTAD2andTAD3orthologs in A-to-I editing, in this study, we used genetic and biochemical approaches to characterize the role ofFgTAD2, an essential adenosine-deaminase-acting-on-tRNA (ADAT) gene, in mRNA editing inFusarium graminearum.FgTAD2had a sexual-stage-specific isoform and formed heterodimers with enzymatically inactiveFgTAD3. Using a repeat-induced point (RIP) mutation approach, we identified 17 mutations inFgTAD2that affected mRNA editing during sexual reproduction but had no effect on transfer RNA (tRNA) editing and vegetative growth. The functional importance of the H352Y and Q375*(nonsense) mutations in sexual reproduction and mRNA editing were confirmed by introducing specific point mutations into the endogenousFgTAD2allele in the wild type. An in vitro assay was developed to show that FgTad2-His proteins purified from perithecia, but not from vegetative hyphae, had mRNA editing activities. Moreover, the H352Y mutation affected the enzymatic activity of FgTad2 to edit mRNA but had no effect on its ADAT activity. We also identified proteins co-purified with FgTad2-His by mass spectrometry analysis and found that two of them have the RNA recognition motif. Taken together, genetic and biochemical data from this study demonstrated that FgTad2, an ADAT, catalyzes A-to-I mRNA editing with the stage-specific isoform and cofactors during sexual reproduction in fungi.