Transcriptome analysis of programmed cell death associated with allorecognition in chestnut blight fungus Cryphonectria parasitica

In this thesis, I analysed transcriptional profiles of the chestnut blight fungus, Cryphonectria parasitica, during vegetative incompatibility (allorecognition) reactions due to differences at the vic3 locus. Out of a total 13944 identified expressed transcripts, including 2334 novel ones, only 1411 were differentially expressed during vic3 incompatibility reaction. Functional enrichment analysis showed increased expression of genes involved in detoxification and stress response (e.g.s Cytochrome p450, Glutathione S-transferase), and toxin biosynthesis. Surprisingly, even though the test strains were both the same mating type (MAT-2), genes involved in sexual reproduction (mf2-1, mf2-2 and mat-2) showed the most dramatic increase in expression during allorecognition response. Further qPCR analysis showed that activation of mating pheromone genes occurs during incompatible reactions involving five of the six known vic incompatibility loci. The only exception was vic4, which elicits a weak incompatibility and showed almost no change in pheromone gene expression. Analysis suggests that mf2-1, mf2-2 and mat-2 expression is triggered by activation of asexual sporulation. Genes encoded at the vic3 locus, vic3a and vic3b, were upregulated in barraging samples along with seven HET-domain genes located at other regions of the genome. Among the seven HET genes activated, one is located at the vic1 locus and previously implicated in vic1 incompatibility. Activation of these same HET domain genes also occurred in other vic incompatible pairings. For example, upregulation of dev3a and vic1a genes occurred during incompatibility reactions associated with each of the six known vic loci. This suggests that some HET genes serve as universal allorecognition factors. Furthermore, this data indicates that each incompatibility locus uses a set of several HET genes to activate Programmed Cell Death (PCD). In addition, I analysed the effects on barrage formation of p29, a protein-coding region from Cryphonectria hypovirus 1 (CHV1). Expression of p29 in C. parasitica was previously shown to delay the onset of vic3-associated PCD. Results of the analysis indicated that ectopically expressed p29 does not have a strong modifying effect on gene expression in barraging strains. This study illustrates that nonself recognition is an active defence mechanism, where stress response and detoxification are combined with mycotoxins production.