Abstract
Drought is one of the environmental factors most affecting crop production. Under drought, symbiotic nitrogen fixation is one of the physiological processes to first show stress responses in nodulated legumes. This inhibition process involves a number of factors whose interactions are not yet understood. This work aims to further understand changes occurring in nodules under drought stress from a proteomic perspective. Drought was imposed on Medicago truncatula 'Jemalong A17' plants grown in symbiosis with Sinorhizobium meliloti strain 2011. Changes at the protein level were analyzed using a nongel approach based on liquid chromatography coupled to tandem mass spectrometry. Due to the complexity of nodule tissue, the separation of plant and bacteroid fractions in M. truncatula root nodules was first checked with the aim of minimizing cross contamination between the fractions. Second, the protein plant fraction of M. truncatula nodules was profiled, leading to the identification of 377 plant proteins, the largest description of the plant nodule proteome so far. Third, both symbiotic partners were independently analyzed for quantitative differences at the protein level during drought stress. Multivariate data mining allowed for the classification of proteins sets that were involved in drought stress responses. The isolation of the nodule plant and bacteroid protein fractions enabled the independent analysis of the response of both counterparts, gaining further understanding of how each symbiotic member is distinctly affected at the protein level under a water-deficit situation.
Highlights
Drought is one of the environmental factors most affecting crop production
As the separation of plant and bacteroid fractions in nodules was one of the aims of this study, we initially evaluated the degree of protein cross contamination between the fractions when comparing homogenization using liquid nitrogen to homogenization using ice-cold extraction buffer followed by centrifugation of the extract to pellet bacteroids
Plant fractions were analyzed via LC/ MS/MS and peptide mass spectra were matched against the S. meliloti protein database to detect bacterial proteins present in the putative plant fraction
Summary
Drought is one of the environmental factors most affecting crop production. Under drought, symbiotic nitrogen fixation is one of the physiological processes to first show stress responses in nodulated legumes. The protein plant fraction of M. truncatula nodules was profiled, leading to the identification of 377 plant proteins, the largest description of the plant nodule proteome so far Both symbiotic partners were independently analyzed for quantitative differences at the protein level during drought stress. N2-fixing legumes are especially sensitive to water deficit and other environmental stresses, with drought being one of the major environmental factors affecting plant productivity (Boyer, 1982; Zahran, 1999) Under this stress, symbiotic nitrogen fixation (SNF) is one of the physiological processes to first show stress responses in nodulated legumes, occurring before the decrease in CO2-photosynthetic assimilation rates (Durand et al, 1987). Special care was taken to ensure minimal cross contamination during the separation of the plant and bacteroid fractions This enabled the independent analysis of the response of both symbiotic partners, gaining further understanding of how each symbiotic member is distinctly affected by drought
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