RNA editing in filamentous fungi

We analyze mRNA editing in fungi using the model organism Sordaria macrospora . This hyphal fungus from the group of ascomycetes has a simple life cycle that is completed within seven days in the laboratory. Only recently, the adenosine (A) to inosine (I) editing of mRNA was detected in filamentously growing ascomycetes. It takes place exclusively during sexual development, more precisely, during the formation of sexual fruiting bodies and spores. How editing is catalyzed enzymatically in ascomycetes is still unknown.

Identification of editing sites
Editing sites can be identified at the RNA level by RNA-Seq. By comparing transcripts from vegetative cells and fruiting body precursors, we were able to show that editing occurs at sexual developmental stages. We refer to the genes whose transcripts are affected by RNA editing as edited in fruiting body development (efd) genes. In cooperation with ISAS e.V. (Dortmund), we were able to show that the edited transcripts are translated and encode proteins with altered amino acid sequences. Both homology-based databases and proteogenomics approaches were used for this purpose.

Impact of RNA editing on protein functions
Editing from A to I in mRNA initially changes the base sequence and thus possibly also the structure of the mRNA. However, changes in the amino acid sequence can also occur at the protein level, as the ribosome interprets I as guanosine (G). Thus, mRNA editing may lead to diversification of the proteome. Editing sites in fungi are mostly located in coding regions. Therefore, they can change amino acid codons or stop codons (Figure 1). We are currently focusing on so-called stop loss editing. Here, a TAG stop codon is edited to a TGG tryptophan codon so that the translated protein becomes longer by as many amino acids as are encoded in the 3'-UTR up to the next stop codon lying in the reading frame.

We use genetic, molecular biological and cell biological methods to address the following questions:
1. What is the function of EFD proteins during fruiting body formation?
2. Is the editing, and thus the elongation of the protein, necessary for its function?
3. What additional properties do EFD proteins acquire through the C-terminal extension?



Figure: Types of mRNA editing and possible connection with the pathogenesis of fungi. Teichert I (2018)

Editing of nuclear mRNA leads to diversification of the proteome, whereas editing of plastid, mitochondrial, and kinetoplast mRNA is mostly restorative. Distinct editing events (marked by blue asterisks) occur in organellar as well as nuclear transcripts. Undulated lines indicate transcripts. In the human parasite Trypanosoma brucei, transcripts derived from maxicircle DNA (black in kDNA) are edited using guide RNA (red) derived from minicircle DNA (red in kDNA). The plant pathogenic fungus Fusarium graminearum shows A-to-I editing of nuclear transcripts during the late sexual phase. Editing leads to diverse changes at the protein level as shown in the blue-outlined box. Editing of distinct transcripts may affect the maturation of fruiting bodies and the formation and discharge of ascospores (blue arrows), which are the primary inoculum of this fungus. A, adenosine; I, inosine, kDNA, kinetoplastid DNA.