Comparative analysis of lifecycle-dependent A to I RNA editing in three members of the Microbotryum violaceum fungal complex

Microbotryum superbum (MvSup), M. intermedium (MI), and M. lychnidis-dioicae (MVLG) are members of the M. violaceum fungal complex. Each species infects specific host plant species, resulting in what is commonly known as anther smut. The lifecycle of these basidiomycete fungi includes the haploid, mating, and infection stages. RNA editing is a post-transcriptional process where adenosine (A) is converted to inosine (I) by adenosine deaminase enzymes (ADARs); such modifications to RNAs may lead to synonymous and nonsynonymous codon changes, thereby altering protein function. We observed that 57% to 77% of total editing sites created nonsynonymous codon changes in both haploid and mating stages of the three species. Moreover, the a2 haploid strain of MI had fewer editing sites compared to other haploid strains. When we compared amino acid substitutions, we found that in both haploids of MvSup and MVLG, Ala was the preferred codon after nonsynonymous codon changes. Among the edited genes, two were edited only at the mating stage in MvSup, undergoing A to I changes within the regions encoding their functional domains. Differential expression analysis revealed that the gene annotated as Apoptosis-inducing factor-1, was upregulated in MvSup at the mating stage, while another gene, for PHB domain-containing protein responsible for cell proliferation, was downregulated compared to the haploid stage. During all stages of the MvSup lifecycle examined, a specific MAPKKK gene was edited in the portion encoding the PKC-like superfamily domain. Also, that gene was edited at a second site during haploid and mating stages but not during the infection stage. Research on RNA editing in basidiomycetes has been limited and is relatively new. RNA editing mechanisms in fungi have been implicated in fungal pathogenesis, although the exact mechanisms and implications remain unclear. Further research is needed to fully understand functional significance of this apparently ubiquitous process in several members of the Microbotryum fungal complex, with possible ramifications more generally in fungi. Author SummaryEditing of mRNAs after transcription provides another mechanism for selective expression of, especially proteins, under different stages of development or environmental conditions. Here we report on the characterization of A-to-I RNA editing in three species of the Microbotryum violaceum fungal complex, members of the Basidiomycota, where such phenomena have so far been unexplored. We find that such editing is prevalent in different stages throughout the lifecycle of this parasite of plant hosts in the Carnation (Pinks) family. We identified preference for edits that lead to specific amino changes, some of which are limited to one or the other haploid mating-type strains, while others are present preferentially in the mated or plant-infection stages of the lifecycle. Some edits occurred in components of conserved signaling pathways, such as the MAPK pathway, or in genes associated with pathogenicity. Taken together, these results suggest additional hypothesis-driven experiments to further investigate the roles of RNA editing in Microbotryum, providing mechanistic insights into the evolution of species in this fungal complex, as well as for those of other pathogenic fungi.