Small RNA discovery in the interaction between barley and the powdery mildew pathogen

Authors: HUNT, MATT - Iowa State University; BANERJEE, SAGNIK - Iowa State University; SURANA, PRIYANKA - Iowa State University; LIU, MEILING - Iowa State University; Fuerst, Gregory; MATHIONI, SANDRA - Danforth Plant Science Center; MEYERS, BLAKE - Danforth Plant Science Center; NETTLETON, DAN - Iowa State University; Wise, Roger

Submitted to: BMC Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/30/2019
Publication Date: 7/25/2019
Citation: Hunt, M., Banerjee, S., Surana, P., Liu, M., Fuerst, G.S., Mathioni, S., Meyers, B.C., Nettleton, D., Wise, R.P. 2019. Small RNA discovery in the interaction between barley and the powdery mildew pathogen. Biomed Central(BMC)Genomics. 20. https://doi.org/10.1186/s12864-019-5947-z.
DOI: https://doi.org/10.1186/s12864-019-5947-z

Interpretive Summary: Small RNAs (sRNAs) in plants play key roles in regulating development, metabolism, and response to both abiotic and biotic stress. The expression of pathogen response proteins is carefully controlled through sRNAs to allow full growth potential during non-infection conditions, with a switch to defense during pathogen challenge. Plants have evolved an innate immune system that allows them to prevent infection from many potential pathogens. Pathogens in turn, have evolved effector molecules that suppress the immune response. Several sRNA families are involved in regulating plant responses to pathogen infection, including hormone signaling, reactive oxygen species evolution, and callose deposition, among others. The goal of this project was to understand how sRNAs regulate gene expression for both barley and its’ powdery mildew pathogen, as well as trans-kingdom sRNA communication between host and pathogen. To accomplish this goal we sequenced sRNA populations from a time course experiment representing penetration of barley cells by the pathogen, as well as subsequent development of fungal feeding structures. After extensive informatics processing of the data, novel sRNAs were predicted to target protein-encoding transcripts encoding a mix of functional categories including signaling, metabolism, transcriptional regulation, virulence and defense. These findings indicate that sRNAs are integral to the regulation of gene expression during defense of barley to powdery mildew infection, and are also used by the pathogen to colonize the host. Impact: Knowledge from this research will impact how plant breeders select for disease resistance, one of the most important traits that affect crop yield, and thus food security.

Technical Abstract: Plants and their pathogens have constantly co-evolving mechanisms that determine infection success. Small RNAs (sRNAs) of 18-30 nucleotides can have a large effect regulating plant defense responses as well as fungal virulence factors. The goal of this project was to understand how sRNAs regulate gene expression both for species of origin, as well as trans-kingdom sRNA communication between barley and Blumeria graminis f. sp. hordei (Bgh), the causal agent of barley powdery mildew. To accomplish this goal we examined Bgh sRNA expression over a time course representing the key stages of Bgh infection of barley (appressorium formation, penetration of epidermal cells, and development of haustoria) in five barley lines including four fast-neutron derived immune-signaling mutants and their progenitor line CI 16151. The sRNA expression data was complemented by parallel analysis of RNA ends (PARE) analysis that confirms sRNA transcript cut sites with in vivo data. In barley, conserved and novel miRNAs were identified with predicted target transcripts enriched in the transcriptional regulation, signaling, and photosynthesis categories. Phasing siRNAs (phasiRNAs) were also identified in barley overlapping with protein coding genes including receptor-like kinases and resistance genes. Bgh micro RNA-like RNAs (milRNAs) were identified that are predicted to regulate transcripts encoding effectors, metabolic proteins, and translation-related proteins. A subset of effectors homologous to the AVRk1 and AVRa10 (EKA) family may be regulated by a sRNA-encoding hairpin that is overlapping and antiparallel to an EKA gene. These genes are heavily regulated by sRNAs, in contrast to most Bgh protein-coding genes. Potential trans-kingdom functional sRNAs were identified from both barley and Bgh. The predicted Bgh trans-kingdom sRNA are highly enriched in transcripts that function in non-species-specific defenses. The transcript targets encode proteins related to vesicle secretion, cell wall synthesis, protein turnover, transcriptional regulation, ROS response, and fungal cell wall breakdown. The potential barley trans-kingdom sRNAs are predicted to target Bgh transcripts including Bgh-specific effector proteins, ribosome synthesis/function, core transcription factors, and cell cycle regulators. Overall, these findings indicate that sRNAs are integral in regulation of gene expression during Bgh infection of barley leaves.