A Differential Proteomic Study Of Rice Seedlings In Response To Elevated CO₂

Previous investigations of plants'response to elevated CO₂ were mostly based on physiological measurements and biochemical assays. In this study proteomic approach was employed to investigate plants response to higher CO₂ levels using rice as a model. Ten days old seedlings were progressively exposed to 760 ppm, 1140 ppm and 1520 ppm CO₂ concentrations for 24 h each. Net photosynthesis rate (Pn), stomatal conductance (Gs), transpiration rate (E) and intracellular to ambient CO₂ concentration ratio (Ci/Ca) were measured. Pn, Gs and E showed maximum increase at 1140 ppm CO₂ but further exposure to 1520 ppm for 24h resulted in down regulation of these. Proteins extracted from leaves were subjected to 2-DE analysis and 57 spots showing differential expression patterns, as detected by profile analysis, were identified by MALDI-TOF/TOF-MS. Most of the proteins belonged to photosynthesis, carbon-metabolism and energy pathways. Several molecular chaperones and ascorbate peroxidase were also found responding to higher CO₂ levels. Correlations between the protein expression patterns and the photosynthetic measurements at the three CO₂ levels were explored. Concomitant with the down regulation of Pn and Gs, the levels of enzymes of the regeneration phase of the Calvin cycle were decreased. Apoplast, the extracellular matrix, is a seat of important signaling and stress/defense related proteins. Rice apoplast proteins are largely unexplored. In a subsidiary study, apoplast extraction method was optimized using vacuum infiltration-centrifugation technique. More than 400 and 150 protein spots were visualized in pI 3-10 range from roots and leaves extracts. 2-D gels in pI 4-7 range showed high quality and reproducibility. Further improvements in the method for differential protein analysis are suggested.