Current Projects
Development of the P. capsici-N. benthamiana interaction model system
The broad host range pathogen P. capsici infects N. benthamiana, a Solanaceous model plant for which an impressive functional genomics toolbox is available. Importantly, P. capsici in genetically amenable to transformation, a feature that allows unparalleled efficiency for studying Phytophthora biology. As a pathogen of many crops, P. capsici is also of strategic and scientific importance as it shares tomato as a host with the economically important pathogen P. infestans. These features allows comparative functional studies between these pathogens which should inform us on basic mechanisms underpinning pathogenesis. We will aim to characterise and employ P. capsici-N. benthamiana interactions. qPCR analysis, gene silencing and over-expression studies of both P. capsici and N. benthamiana genes will be initiated to explore and unveil new processes that underpin host-pathogen interactions.
Studies of CRN effector translocation in P. capsici
Recent multi-faceted investigations of Phytophthora secreted proteins have identified and implicated Crinkler genes (CRN for Crinkling and Necrosis) as another family of cytoplasmic effectors. Similar to the RxLRs, CRN proteins have modular structures, harboring predicted secretion signal sequences followed by conserved N-terminal, and diverse C-terminal, domains. This project aims to study the pathways that mediate CRN effector delivery during infection. We hypothesize that CRN delivery requires plant and/or Phytophthora proteins that interact with CRN-N-termini. To identify such components, we employ Yeast two Hybrid (Y2H) analyses as well as biochemical approaches. Candidate interactors will be further studied using the P. capsici-N. benthamiana interaction system.
Implementation of visual fluorescent markers for studying parasite effector trafficking during infection
Phytophthora effector repertoires have emerged as principal determinants of epidemics. From the effectors identified, two (RxLR and CRN) protein classes are delivered inside host plant cells, whereas others act outside of host cells. A lack of robust visualization tools hampers efforts towards answering the most pressing questions: how, when and where do plant pathogens deliver their effectors? To allow critical studies of parasite effector targeting and delivery in hosts we will (1) Test diverse fluorescent proteins in P. capsici (2) Evaluate effector-reporter fusions in vivo (3) Optimize reporters to study effector translocation processes effector translocation processes
Identification and Functional characterization of the P. capsici Crinkler effector repertoire
Phytophthora spp continue to threaten major crop growing regions around the world. With the availability of genome sequences from some of these pathogens, computational and functional analyses have generated a blueprintof effector repertoires that are thought to be required for parasitism. Amongst the effector repertoires present in Phytophthora, a large family of host?translocated proteins was identified. The Crinkler (CRN) gene familyencodes a novel class of effector proteins present in all plant pathogenic oomycetes examined to date. The CRN protein family is a novel class of translocated proteins whose functions and roles in parasitism are completely unknown. This work aims to clone and validate genes identified in P. capsici
sequencing projects and deduce their localisation and function in vivo. These analyses will provide an overview of the Phytophthora effector blueprint and allow comparative analyses between the effector repertoires of economically important pathogens.