SUMOylation is a post-translational modification in which the small ubiquitinlike protein SUMO is attached to lysine residues of the target protein. In plants, wide-spread SUMOylation is observed upon a variety of different stress cues. We tested Arabidopsis SUMOylation machinery knockout mutants for impaired disease resistance against Pseudomonas syringae pv. tomato (Pst) and identified sumo2-1, sae1a-1 and sae2-3 as showing moderate but statistically insignificant resistance. sumo2-1 also exhibited slightly reduced HR compared to the wild-type plants after PstDC3000(avrB) challenge. Change in the cellular redox status is an important outcome of attempted pathogen ingress. Therefore, we also looked at the redox regulation of SUMOylation both in vivo and in vitro. We found a significant increase in SUMO1/2 conjugation and free SUMO1/2 accumulation in atgsnor1-3 plants after PstDC3000(avrB) challenge which was reversed during the establishment of disease in the absence of an AvrB, suggesting an important role of S-nitrosylation in modulating plant SUMOylation. High basal level of high molecular weight (HMW) SUMO1/2 conjugates was also apparent in atgsnor1-3 plants even in the absence of pathogens. The changes in SUMO3 and SUMO5 remained less significant and their regulation was found to be independent of GSNOR. Biotin switch technique was employed to test further if SUMO enzymes are modified by NO. It was found that SCE1 and SAE1a are S-nitrosylated in vitro in a GSNO dose dependent fashion. MS analysis and site directed mutagenesis revealed Cys139 in SCE, and Cys93, Cys158 and Cys231 in SAE1a as the targets of S-nitrosylation. We established that GSNO treatment to SCE1 differentially regulates in vitro SUMOylation of the model substrate ScPCNA. Furthermore, the S-nitrosylation of Cys139 of SCE1 is important in fine-tuning protein SUMOylation under changing cellular redox tone. These data highlight the complexity of cross-communication between two different post-translational modifications (i.e. S-nitrosylation and SUMOylation) in the control of protein function.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:764176 |
| Date | January 2010 |
| Creators | Malik, Saad Imran |
| Contributors | Loake, Gary ; Kidner, Catherine |
| Publisher | University of Edinburgh |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/1842/33316 |
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