Wed Dec 9 // Ryan Kilburn // PhD Candidate, Queen's University

Ryan Kilburn
PhD Candidate, Plaxton & Snedden Labs
Investigations into the multifaceted functions of Ricinus communis calcium-dependent protein kinase-1 (RcCDPK1) 

As the global population grows and demands on agricultural land rise, there is a need to increase crop yields. Castor (Ricinus communis) oil seeds (COS) have become an important model for oil seed metabolic engineering due to their massive accumulation of oil at maturity relative to commercially important oil seeds such as canola. In developing COS, a unique ‘bacterial-type’ phosphoenolpyruvate carboxylase (PEPC) isozyme is highly expressed as a regulatory and catalytic subunit of a novel Class-2 PEPC complex. Class-2 PEPC’s unique kinetic and regulatory properties, and dynamic subcellular targeting to the mitochondrial surface, support the hypothesis that it facilitates rapid refixation of respiratory CO2 while sustaining a large anaplerotic flux to replenish TCA cycle C-skeletons withdrawn in support of storage oil and protein biosynthesis in developing COS. R. communis Ca2+-dependent protein kinase-1 (RcCDPK1) catalyzes in vivo inhibitory phosphorylation of bacterial-type PEPC (BTPC) at Ser451 in developing COS (Ying et al. 2017 Plant Physiol.). This research aims to address how autophosphorylation influences RcCDPK1’s ability to transphosphorylate BTPC at Ser451. Interestingly, RcCPDK1 exhibits high sequence similarity (83%) to its closest ortholog from the model plant Arabidopsis thaliana, AtCPK4, which also catalyzes Ca2+-dependent phosphorylation of COS BTPC at Ser451 in vitro. AtCPK4 also appears to regulate signal transduction of the stress hormone abscisic acid (ABA) by phosphorylating an important ABA-responsive transcription factor (AtABF4), raising questions about further overlap between substrates of these two CDPK orthologs, and ultimately whether RcCDPK1 might also function in castor ABA signaling. Using a mix of biochemistry and genetic approaches with a focus on enzyme kinetics and functional genomics, this project aims to unravel understudied aspects of autophosphorylation in CDPKs, as well as to determine possible links between the control of central carbon metabolism and ABA signaling.