Phytopathology. Biochemistry. Molecular Biology.
I joined the Plant Chemetics Lab in June 2014 as a DPhil Student. In my research I study the role of apoplastic Cysteine proteases of tomato in molecular recognition and immunity. I focus on the immune proteases RCR3 and PIP1.
RCR3 and PIP1 are homologous Papain-like Cysteine Proteases (PLCPs). Pathogen-derived Protease Inhibitors, which are required for pathogenicity, target RCR3 and PIP1. Furthermore, knock-out and silencing of RCR3 and PIP1 respectively increase host susceptibility to a variety of pathogens. The interaction of RCR3 with several of these protease inhibitors, but not the interaction of these inhibitors with PIP1, is sensed by the Receptor-like protein Cf-2 originating from Currant tomato (Solanum pimpinellifolium). This perception results in a strong immune response leading to resistance against the pathogen, and has therefore been introgressed into commercial tomato varieties.
In the first part of my project I aim to identify the region in RCR3 interacting with Cf-2. For this I have made a library of mutant RCR3 alleles, which I am currently using for functional assays. In the second part of my project I aim to identify the substrates of RCR3 and PIP1. This will involve a variety of proteomic techniques, including PICS and TAILS. Identifying substrates of RCR3 and PIP1 will allow us to understand the way in which these proteases function in immunity, as well as indicate whether they function in similar or distinct pathways. These two approaches will lead to a comprehensive understanding of the roles of RCR3 and PIP1 in immunity.
So far I have successfully established Golden Gate cloning in our lab, and have made Golden Gate compatible E. coli, Pichia Pastoris, and TRV vectors, as well as many modules commonly used in our lab. Furthermore, I have successfully established CRISPR/Cas9-mediated genome engineering of Nicotiana benthamiana and tomato in the lab.