Graduate Thesis Or Dissertation
 

Alternative splicing in the obligate biotrophic oomycete pathogen, Pseudoperonospora cubensis

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  • Oomycetes are some of the most devastating pathogens, causing upwards of billions of dollars of damage each year to plants. They also diminish ecological diversity and health through the destruction of trees and shrubs. The genome sequence of Pseudoperonospora cubensis, an obligate plant pathogen and causative agent of downy mildew in cucurbits, was originally generated as a first step for discovering candidate virulence genes. Through these efforts, the novel discovery was made that a multidrug transport encoding gene was alternatively spliced, giving rise to a truncated protein that, unlike the full length form, exhibits characteristics consistent with in planta virulence functions. Alternative splicing can generate different combinations of gene sequences, thereby increasing transcriptome and proteome complexity to influence gene regulation and phenotypic plasticity. Because of the limited number of studies, the impact of alternative splicing on virulence and development of oomycetes is unknown. To address this knowledge gap, we used RNA-Seq to deeply sequence Ps. cubensis transcriptomes to assess the impact of alternative splicing during its infection of the host Cucumis sativus (cucumber). In addition a number of computational and statistical tools will be described that were developed to help improve the draft genome and faciliate the characterization of alternative splicing. We demonstrate that alternative splicing influences at least 26% of the Ps. cubensis genome with potential effects on gene function, thus highlighting its importance in pathogenesis. This work represents the first step towards understanding the role of alternative splicing in an obligate oomycete pathogen and lays the groundwork for further dissecting the role of alternative splicing in pathogenesis.
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