Proteomics analysis of a tobacco variety resistant to brown spot disease and functional characterization of NbMLP423 in Nicotiana benthamiana

Background: Tobacco brown spot disease is an important disease caused by Alternaria alternata that affects tobacco production and quality worldwide. Planting resistant varieties is the most economical and effective way to control this disease. However, the lack of understanding of the mechanism of tobacco resistance to tobacco brown spot has hindered progress in the breeding of resistant varieties.

Methods and results: In this study, differentially expressed proteins (DEPs), including 12 up-regulated and 11 down-regulated proteins, were screened using isobaric tags for relative and absolute quantification (iTRAQ) by comparing resistant and susceptible pools and analyzing the associated functions and metabolic pathways. Significantly up-regulated expression of the major latex-like protein gene 423 (MLP 423) was detected in both the resistant parent and the population pool. Bioinformatics analysis showed that the NbMLP423 cloned in Nicotiana benthamiana had a similar structure to the NtMLP423 in Nicotiana tabacum, and that expression of both genes respond rapidly to Alternaria alternata infection. NbMLP423 was then used to study the subcellular localization and expression in different tissues, followed by both silencing and the construction of an overexpression system for NbMLP423. The silenced plants demonstrated inhibited TBS resistance, while the overexpressed plants exhibited significantly enhanced resistance. Exogenous applications of plant hormones, such as salicylic acid, had a significant inducing effect on NbMLP423 expression.

Conclusions: Taken together, our results provide insights into the role of NbMLP423 in plants against tobacco brown spot infection and provide a foundation for obtaining resistant tobacco varieties through the construction of new candidate genes of the MLP subfamily.