Comparative Proteomic Analysis of Wild Type and Mutant Lacking an SCF E3 Ligase F-Box Protein in Magnaporthe oryzae.

Abstract

Magnaporthe oryzae (M. oryzae) is a pathogenic, filamentous fungus that is a primary cause of rice blast disease. The M. oryzae protein MGG_13065, SCF E3 ubiquitin ligase complex F-box protein, has been identified as playing a crucial role in the infection process, specifically, as part of the ubiquitin mediated proteolysis pathway. Proteins targeted by MGG_13065 E3 ligase are first phosphorylated and then ubiquitinated by E3 ligase. In this study, we used a label-free quantitative global proteomics technique to probe the role of ubiquitination and phosphorylation in the mechanism of how E3 ligase regulates change in virulence of M. oryzae. To do this, we compared the WT M. oryzae 70-15 strain with a gene knock out (E3 ligase KO) strain. After applying a ≥ 5 normalized spectral count cutoff, a total of 4432 unique proteins were identified comprised of 4360 and 4372 in the WT and E3 ligase KO samples, respectively. Eighty proteins drastically increased in abundance, while 65 proteins decreased in abundance in the E3 ligase KO strain. Proteins (59) were identified only in the WT strain; 13 of these proteins had both phosphorylation and ubiquitination post-translational modifications. Proteins (71) were revealed to be only in the E3 ligase KO strain; 23 of the proteins have both phosphorylation and ubiquitination post-translational modifications. Several of these proteins were associated with key biological processes. These data greatly assist in the selection of future genes for functional studies and enable mechanistic insight related to virulence.