Minisymposium 10: Photosynthesis: Light Adaptation

Abs # 22002: Functional analysis of CtpA in transgenic arabidopsis plants

Presenter:	Chen, Yun-Chia S, sophia.y.chen@monsanto.com
Authors	Chen, Yun-Chia S (A)   Duff, Stephen M (A)   Fabbri, Bradon J (A)   Anderson, John C (A)   Oliver, Robyn  (B)   CaJacob, Claire A (A)
Affiliations:	(A): Monsanto Company (B): Mallinckrodt Pharmaceuticals

Photosystem II (PSII) is an essential part of the photosynthetic apparatus in plants that uses light energy to split water, releasing oxygen, protons and electrons. The carboxyl-terminal processing protease (CtpA) is an enzyme located in the thylakoids of higher plants that catalyses the conversion of pre D1 protein to the active form of D1, which is an integral part of PSII, by cleaving a small number of C-terminal residues (9 from spinach CtpA). This process is essential to the formation of active D1 protein. Since the D1 protein is continuously removed from the PSII and replaced, the active CtpA is constantly required under light conditions. To elucidate the biological function of CtpA and to validate CtpA as a target protein for rational herbicide discovery, we over-expressed CtpA sense and antisense mRNAs in Arabidopsis plants to alter the internal level of CtpA protein in the plants. Using antibodies raised against recombinant CtpA protein, we demonstrated that we have generated transgenic Arabidopsis plants with reduced level of CtpA protein or with elevated level of CtpA protein by the over-expression of CtpA sense or antisense mRNAs. Transgenic plants with severely reduced levels of CtpA protein exhibit a bleached/chlorotic phenotype and stunted growth. The mutant phenotypes can be further enhanced by bright illumination. Supplement of sugar can partially rescue the lethal phenotypes. However, plants with a slight reduction of CtpA protein do not exhibit the mutant phenotype and cannot be distinguished from wild-type plants under normal growth conditions. Rational herbicide discovery consists of identifying and validating an essential biological process, which is then targeted for disruption by small molecules. CtpA is an excellent example of a rational herbicide discovery target due to its essential role in plant growth and development. Several CtpA enzyme inhibitors were developed and were shown to have herbicidal activity in planta. The process of development of small molecules targeting CtpA will be discussed with emphasis on the in vivo biological activity of these compounds and validation that their mode of action (MOA) is in fact CtpA. Interestingly, plants that produce excessive amount of CtpA protein were shown to be resistant to these inhibitors. Our results suggest that CtpA is essential for plant growth and development due to its critical role in photosynthesis.