SUMO: a heavyweight post- translational modification wrestling between plant stress responses.

Abstract

Post-translational modifications (PTMs) enable plants to modify proteins in response to environmental stimuli. Better understanding these PTMS is important to help crops adapt to the changing environment caused by climate change. This thesis has studied the role of a PTM called Small Ubiquitin Like Modifier (SUMO) which is stress induced. Conjugation of SUMO to target proteins alters the target protein, some of these proteins have been examined in this thesis.
The research has specifically examined the role of SUMO in the Abscisic Acid (ABA) pathway. The work has deduced the ABA sensitivity of SUMO protease knockout mutants. It has established the SUMOylation sites in the ABA negative regulator Protein Phosphatase 2CA (PP2CA) and generated transgenic Arabidopsis thaliana expressing both a SUMOylatable and non-SUMOylatable form of PP2CA to determine the ABA phenotype. ABA mediated root inhibition and expression of ABA response genes has indicated that SUMOylated PP2CA may aid ABA signalling. It was hypothesised this may be in part due to a SUMO Interacting Motif (SIM) site located in the PYLS, a SIM site was identified in PYL8 and was not present in PYL1. Arabidopsis thaliana transgenics were generated to enable further understanding of the role of SUMO in the ABA pathways.
Additionally the role of SUMO in root hair formation was examined, based on the double SUMO protease knockout mutant ots1 ots2 being found to have longer and fewer root hairs. A transcription factor required for root hair development RHD-Six Like 4 (RSL4) was found to contain three SUMO sites. Transgenic Arabidopsis thaliana was generated to examine the role of SUMOylatable and SUMO site mutated RSL4, however this led to contradictory results. Protein stability indicated that SUMOylatable RSL4 may be more stable and/or transcribe more RSL4 promoted genes than SUMO site mutated RSL4, but has smaller root hairs when grown on ethylene supplemented media. These contradictory results may be due to different roles of the three SUMO sites in RSL4.
Lastly research was continued on a project that had been started by Dr Vivek Verma, who had determined that a SUMO site in the GRAS domain of DELLA protein, Gibberellic Acid Insensitive (GAI) altered interactions with transcription factors. A SIM site was identified in Phytochrome Interacting Factor 4 (PIF4), which may be responsible for increased interaction of PIF4 when GAI is SUMOylated. Wild-type and SIM mutated PIF4 transgenic Arabidopsis thaliana was generated to understand the role of SUMO in GAI PIF4 interactions. The GAI SUMO site mutated transgenic Arabidopsis thaliana which had been generated by Dr Vivek Verma were analysed for Jasmonic acid phenotype and hypocotyl elongation phenotype.