About the Project
The rapid and structured disassembly of plant proteomes during germination, senescence and defence are massive proteolytic events that underpin plant development and immunity. Surprisingly, however, the proteases responsible for these proteolytic events are yet unidentified. Because of their strong upregulation during proteolysis, papain-like cysteine proteases (PLCPs) are strong candidates. Indeed, chemical knock-out experiments indicate crucial roles for PLCPs in proteolysis. However, single and double mutants lacking PLCPs do not display proteolytic phenotypes, suggesting that they act redundantly [1]. The Arabidopsis genome encodes for 31 PLCPs that fall into nine different subfamilies [2]. Members of several subfamilies are transcriptionally coregulated, which predicts a cooperation or redundancy between PLCPs of different subfamilies.
The aim of this project is to unravel functions of the PLCP super family in the degradation of seed storage proteins, leaf senescence and hypersensitive cell death. The approach is to deplete multiple genes encoding PLCPs using CRISPR/Cas9 genome editing combined with classical genetics and investigate the effect of PLCP depletion on protein degradation. We will investigate proteomes of mutant plants using PROTOMAP [3] and TAILS [4] to identify putative substrates and unravel redundant proteolytic cascades. We will compare this with substrate specificities of each PLCP determined by PICS [4]. These techniques will be combined with protease activity profiling [2, 5] to monitor the effects of gene inactivation on the PLCP activity landscape. These activities will be combined with germination, senescence and disease assays to elucidate biological roles of the PLCPs.
Student Profile
This is an exciting, challenging project for a devoted, inquisitive minded student with an appetite for protein biochemistry and genetics. Experience with molecular cloning is essential, and practice working with Arabidopsis and/or proteins is advantageous. The student will be part of a strong, dynamic interdisciplinary research team that exploits similar approaches to study proteolysis in other plants.
Funding Notes
There are two main routes into the Department of Plant Sciences Graduate Programme dictated by different funding mechanisms: If, after discussion with a potential supervisor, you decide that one of these programmes is right for you, you will need to apply directly to the relevant programme or scholarship
Fully funded studentships/scholarships are available via linked Doctoral Training centres/Partnerships, directly via departmental project opportunities, or via competitive scholarships. Please use the University's Fees, Funding and Scholarship search tool to identify the funding options available to you.
References
Lu, H., Chandrasekar, B., Oeljeklaus, J., Misas-Villamil, J. C., Wang, Z., Shindo, T., Bogyo, M., Kaiser, M., and Van der Hoorn, R. A. L. (2015) Subfamily-specific probes for Cys proteases display dynamic protease activities during seed germination. Plant Physiol. 168, 1462-1475.
Richau, K., Kaschani, F., Verdoes, M., Pansuriya, T. C., Niessen, S., Stüber, K., Overkleeft, H. S., Bogyo, M., and Van der Hoorn, R. A. L. (2012) Subclassification and biochemical analysis of plant papain-like cysteine proteases displays subfamily-specific characteristics. Plant Physiol. 158, 1583-1599.
Dix, M. M., Simon, G. M., and Cravatt, B. F. (2008) Global mapping of the topography and magnitude of proteolytic events in apoptosis. Cell 134, 679-691.
Huesgen, P. F., and Overall, C. M. (2012) N- and C-terminal degradomics: new approaches to reveal biological roles for plant proteases from substrate identification. Physiol. Plant. 145, 5-17.
Continue with Facebook
