Abstract Detail

Host/Plant Pathogen Interactions and Plant Health Management

Saville, Barry [1], Ostrowski, Lauren [2], Donaldson, Michael [1], Goulet, Kristi [3], Cheung, Hoi Yee Kitty [2].

Functional and transcriptome analysis of antisense RNA in Ustilago maydis.

Biotrophic fungal plant pathogens cause billions of dollars in losses to North American crops annually. The model for functional investigation of these fungi is Ustilago maydis, the causal agent of common smut of corn. Early transcriptome studies of this fungus revealed the presence of 200+ antisense RNAs (asRNAs). Functional analyses revealed that altering expression of three U. maydis asRNAs led to attenuated pathogenesis. We hypothesized that other U. maydis asRNAs had roles in controlling gene expression which influenced pathogenesis. The two approaches used to investigate this were: characterizing the function of an asRNA (as-ssm1) to the mitochondrial seryl-tRNA synthetase (ssm1) gene, and a comprehensive comparative RNA-seq analysis of closely related smuts U. maydis, Sporisorium reilianum (head smut of corn), and Ustilago hordei (covered smut of barley) through which we discovered large numbers of asRNAs and long non-coding RNAs (lncRNAs). These data suggest a major role for non-coding RNAs in controlling gene expression in biotrophic fungal pathogens. The U. maydis as-ssm1 is an asRNA to ssm1 that is expressed in the dormant teliospore, suggesting it may have a role in gene regulation during dormancy. S. reilianum also has an asRNA to its ssm1 ortholog. Functional analyses revealed that blocking ssm1 expression stops cell division in haploid strains. Ectopic expression of as-ssm1 in solopathogenic haploid cells resulted in a significant decline in growth rate in liquid culture, attenuated pathogenesis in seedlings, and a decrease in mitochondrial membrane potential in comparison to wild-type cells. These findings support the hypothesis that as-ssm1 is involved in moderating mitochondrial function during the metabolic transition from the physiologically active dikaryon to dormant teliospore. Expression of as-ssm1 also resulted in an increase in ssm1 mRNA levels and the formation of sense-antisense double-stranded RNA (dsRNA). This suggested that as-ssm1 expression inhibits Ssm1 protein production through stabilization of ssm1 mRNA. Western analysis is underway to assess this. To expand our knowledge of U. maydis as a model and provide insight regarding the impact of the loss of RNA interference machinery on non-coding RNA profiles, comprehensive strand-specific RNA-seq was performed on the related smut fungi U. maydis, S. reilianum, and U. hordei. A relational database was created and analysis will be presented that shows conservation of asRNAs and lncRNAs among the smut fungi. Further we will present expression profiles of these non-coding RNAs among the various U. maydis cell types as an initial step in determining their roles in pathogenic development.

Related Links:
The Saville Laboratory

1 - Trent University, Environmental and Life Sciences Graduate Program; Forensic Science Program, DNA Building, 2140 East Bank Dr, Peterborough, ON, K9J 7B8, Canada
2 - Trent University, Environmental and Life Sciences Graduate Program, DNA Building, 2140 East Bank Dr, Peterborough, ON, K9J 7B8, Canada
3 - Trent University, Environmental and Life Sciences, DNA Building, 2140 East Bank Drive, Peterborough, ON, K9J 7B8, Canada

Keywords:
Smut fungi
Antisense RNA
Functional genomics
Transcriptome
pathogenesis
mitochondria
Dormancy.

Presentation Type: Oral Paper:Papers for Topics
Session: 57
Location: Salon 9/The Shaw Conference Centre
Date: Wednesday, July 29th, 2015
Time: 9:00 AM
Number: 57005
Abstract ID:1210
Candidate for Awards:None