The barley HvWRKY6 transcription factor is required for resistance against Pyrenophora teres f. teres

Authors: TAMANG, PRABIN - North Dakota State University; RICHARDS, JONATHAN - North Dakota State University; SOLANKI, SHYAM - Washington State University; AMEEN, GAZALA - Washington State University; POUDEL, ROSHAN - North Dakota State University; DEKA, PRIYANKA - North Dakota State University; EFFERTZ, KARL - Washington State University; CLARE, SHAUN - Washington State University; HEGSTAD, JUSTIN - North Dakota State University; BEZBARUAH, ACHINTYA - North Dakota State University; LI, XUEHUI - North Dakota State University; HORSLEY, RICHARD - North Dakota State University; Friesen, Timothy; BRUEGGEMAN, ROBERT - Washington State University

Submitted to: Frontiers in Genetics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/14/2020
Publication Date: 1/15/2021
Citation: Tamang, P., Richards, J.K., Solanki, S., Ameen, G., Poudel, R.S., Deka, P., Effertz, K., Clare, S., Hegstad, J., Bezbaruah, A.N., Li, X., Horsley, R.D., Friesen, T.L., Brueggeman, R.S. 2021. The barley HvWRKY6 transcription factor is required for resistance against Pyrenophora teres f. teres. Frontiers in Genetics. 11:601500. https://doi.org/10.3389/fgene.2020.601500.
DOI: https://doi.org/10.3389/fgene.2020.601500

Interpretive Summary: Net form net blotch (NFNB) is arguably the most economically important leaf disease of barley in the United States and the world. Understanding how barley defends itself against this pathogen is not well understood, especially as it relates to the highly resistant barley line CI5791. In this study, we evaluated irradiated barley seed of the NFNB resistant barley line CI5791 to identify two mutant barley lines that were susceptible to NFNB. The same mutated gene was identified in both mutant barley lines and silencing of this gene in the resistant CI5791 barley line resulted in susceptibility to NFNB. Based on the class of the mutated gene, the gene was hypothesized to be controlling the expression of other genes involved in the defense response. Understanding precisely what this gene does is important to understanding how barley mounts a defense response against the NFNB pathogen. This information will be used by pathologists and geneticists to continue to better understand this important pathosystem.

Technical Abstract: The barley line CI5791 exhibits a high level of resistance to diverse isolates of Pyrenophora teres f. teres 30 (Ptt), causal agent of the disease net form net blotch, which affects barley production world-wide. To identify genes involved in this effective resistance, CI5791 seed were gamma-irradiated and two mutants, designated CI5791- gamma 3 and CI5791- gamma 8, with compromised Ptt resistance were identified at the M2 generation. Phenotyping of CI5791- gamma 3 and gamma 8 × Heartland F2 populations showed 3:1 (resistant: susceptible) segregation ratios and CI5791- gamma 3 × gamma 8 F1 individuals were susceptible, thus, the single mutated genes are allelic. Thirty-four homozygous mutant (susceptible) CI5791- gamma 3 × Heartland F2 individuals, representing 68 recombinant gametes, were genotyped via PCR-genotype-by-sequencing and single marker regression mapping placed the mutation on chromosome 3H within an ~75 cM interval. Sequencing of the mutants and wild-type (wt) CI5791 genomic DNA following exome capture identified independent mutations of the HvWRKY6 transcription factor located on chromosome 3H at ~50.7 cM, within the genetically delimited region. Post transcriptional gene silencing of HvWRKY6 in barley line CI5791 resulted in Ptt susceptibility, confirming that it functions in NFNB resistance, validating it as the gene underlying the mutant phenotypes. Allele analysis and transcript regulation of HvWRKY6 from resistant and susceptible lines revealed sequence identity and up regulation upon pathogen challenge in all genotypes analyzed, suggesting a conserved transcription factor that is involved in defense against the necrotrophic pathogen. We hypothesize that HvWRKY6 functions as a conserved component of a basal defense mechanism that restricts Ptt growth in barley.