Role of non-programmed cell death inducing effectors in the Parastagonospora nodorum-wheat necrotrophic interaction

Authors: MALVESTITI, MICHELE - North Dakota State University; KARIYAWASAM, GAYAN - North Dakota State University; LI, JINLING - North Dakota State University; Wyatt, Nathan; NELSON, ASHLEY - North Dakota State University; SKIBA, RYAN - Oak Ridge Institute For Science And Education (ORISE); Fiedler, Jason; EFFERTZ, KARL - North Dakota State University; LIU, ZHAOHUI - North Dakota State University; WILLIAMS, SIMON - Australian National University; BRUGGEMAN, ROBERT - Washington State University; Friesen, Timothy

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 12/6/2023
Publication Date: 3/12/2024
Citation: Malvestiti, M., Kariyawasam, G., Li, J., Wyatt, N.A., Nelson, A., Skiba, R., Fiedler, J.D., Effertz, K., Liu, Z., Williams, S., Bruggeman, R., Friesen, T.L. 2024. Role of non-programmed cell death inducing effectors in the Parastagonospora nodorum-wheat necrotrophic interaction. Meeting Abstract.

Interpretive Summary:

Technical Abstract: The necrotrophic Ascomycete Pyrenophora teres f. teres is the causal agent of net form net blotch in barley (Hordeum vulgare). A previous study used a biparental population of P. teres f. teres isolates 15A and 6A to identify two quantitative trait loci (QTL), namely, VR1 and VR2. Each QTL contained a gene which contributed to virulence in Rika barley. In this study, we cloned and functionally validated both VR1 and VR2 and investigated their role in fungal virulence. We used CRISPR-Cas9-based gene disruption and gene editing, QTL analysis, haplotype and isoform diversity analysis, protein structure prediction, quantitative PCR, and laser confocal microscopy to validate and functionally characterize VR1 and VR2 and the corresponding proteins. Both VR1 and VR2 were present in a global P. teres f. teres collection and isolates possessing different VR1 and VR2 protein isoforms quantitatively varied in virulence. Protein structure prediction revealed that VR1 encodes for a secreted prolyl-endopeptidase, whereas VR2 encodes for a small secreted protein with unknown domains. Inoculation of the VR1 and VR2 edited isolates onto the Rika × Kombar barley population showed that both VR1 and VR2 were likely targeting the same susceptibility locus Spt1 in Rika barley chromosome 6H. The VR1 and VR2 gene-edited isolates showed that virulent alleles of VR1 and VR2, derived from P. teres f. teres isolate 6A, were sufficient to cause disease on Rika barley alone. However, increased symptom severity was observed on Rika barley when both VR1 and VR2 virulent alleles were present in the VR1 and VR2 gene-edited isolates. Analogously, confocal microscopy data showed that the fungal isolates possessing the virulent VR1 and VR2 allele displayed more rapid host tissue colonization and increased fungal biomass. Isogenic isolates carrying both VR1 and VR2 showed an additional increase in symptom severity and biomass relative to isolates carrying VR1 or VR2 alone. Taken together, these observations suggest that VR1 and VR2 may act in a synergistic manner. To validate this hypothesis, single and double VR1 and VR2 gene disruption mutants in P. teres f. teres isolate 6A were generated to independently assess the contribution of each effector gene to fungal virulence. Infection assays on Rika barley with the wild type P. teres f. teres isolate 6A and single and double VR1 and VR2 mutant isolates are currently ongoing. We expect to present the outcome of the infection assays at the time of the conference.