Proteomic Study of SUMOylation During Solanum tuberosum-Phytophthora infestans Interactions.

Invasive plant pathogens have developed the ability to modify the metabolism of their host, promoting metabolic processes that facilitate the growth of the pathogen at the general expense of the host. The particular enzymatic process SUMOylation, which performs posttranslational modification of target proteins, leading to changes in many aspects of protein activity and, hence, metabolism, has been demonstrated to be active in many eukaryotic organisms, both animals and plants. Here, we provide experimental evidence that indicates that, in leaves of Solanum tuberosum that have been infected by Phytophthora infestans, the SUMO (small ubiquitin-like modifier) pathway enzymes of the host are partially under transcriptional control exerted by the oomycete.

Proteomics analysis of the endogenous, constitutive, leaf SUMOylome.

Unlabelled: SUMOylation is a post-translational modification which regulates a number of critical biological processes in, for example mammals, yeast and plants. In order to fully understand the functional effects of SUMOylation an essential first step is the identification of endogenous targets for SUMOylation. Here we report the results of using a recently developed proteomic approach based on the use of 3D gels to identify the endogenous SUMO targets in leaves of Solanum tuberosum.

Three-dimensional electrophoresis for quantitative profiling of complex proteomes.

Quantitative 2D-gel-dependent proteomics became feasible with 2D fluorescence difference gel electrophoresis (2D-DIGE), and this technique has gained wide acceptance because it has eliminated the gel to gel variations and greatly facilitated the quantitative comparisons across gels for many different experimental conditions. However, the co-migration of several proteins in the same spot is still a major limitation which detracts from the accuracy of comparative quantification and prevents unambiguous post-translational modifications (PTMs) detection.A protocol based on traditional polyacrylamide gel IEF sample fractionation, and followed by two consecutive SDS-PAGE electrophoreses alleviates co-migration limitations.

Evaluation of three-dimensional gel electrophoresis to improve quantitative profiling of complex proteomes.

Two-dimensional remains one of the main experimental approaches in proteome analysis. However, comigration of protein leads to several limitations: lack of accuracy in protein identification, impaired comparative quantification, and PTM detection. We have optimized a third additional step of in-gel separation to alleviate comigration associated drawbacks.