Chloroplast proteins involved in drought stress response in selected cultivars of common bean (Phaseolus vulgaris L.)

Abstract

One of the major cell organelles, whose functions are affected during drought stress are chloroplasts. In this study, chloroplast proteome under drought was studied in two cultivars of common bean (Phaseolus vulgaris L), Tiber and more sensitive to drought, Starozagorski čern, which were subjected to drought for 6 and 13 days. A comparative proteomic analysis with 2D-DIGE was performed on the isolated chloroplast proteins from leaves. Together, 44 proteins with changed abundance between control and stressed plants were identified with LC-MS/MS from both cultivars. The majority of the identified proteins were involved in photosynthetic processes. The results showed a decrease in abundance in different structure components of photosystem I and II, and ATP synthase, which may indicate a suppression of light-dependent reactions by drought stress. Similar proteomic response for both cultivars after 6 and 13 days of drought was observed. Proteins with contrasting abundance patterns between the cultivars or proteins specific for only one cultivar, such as ferredoxin-NADP reductase, photosystem II stability/assembly factor HCF136, curvature thylakoid protein 1B, and plastidial membrane protein porin were pointed out as major identified proteins revealing differential abundance between the cultivars. Taken together, our results provide insight into the molecular response of chloroplasts in common bean under drought stress, whereas conclusions about the tolerance mechanisms require further studies.

Keywords: Chloroplast; Drought stress; Phaseolus vulgaris; Proteomics.

Fig. 1

Effect of drought stress on RWC (a) and SWC (b). RWC and SWC data were analyzed using Student t test to determine the significant difference between group means. Values are the means ± SE from eight replicates. One asterisk indicates significant difference (p < 0.05) between control and stress sample for each cultivar (Tiber and Starozagorski), whereas two asterisks point to significant difference between the cultivars of drought-stressed plants

Fig. 2

Immunoblot analysis of chloroplast and total protein fractions from common bean leaves. Isolated protein fractions were separated by SDS–PAGE, transferred to a PVDF membrane and incubated with anti-COX II and anti-Lhcb7 antibodies. The signal was detected by chemiluminescence after incubation with the HRP-conjugated goat anti-rabbit secondary antibodies

Fig. 3

Representative 2-DE gel image of chloroplast proteins extracted from leaves of common bean. Separation of proteins was performed on 24 cm IPG strip (pH 4–10 non-linear gradient) by isoelectric focusing, followed by 12.5% SDS–PAGE. Proteins were visualized by Coomassie Brilliant Blue G-250 staining. The arrows show the position of the identified proteins which were changed in abundance under drought stress along with numbers which represent protein spots as shown in Table 1

Fig. 4

Distribution of functional categories of all identified proteins in chloroplasts of Tiber and Starozagorski in response to drought stress

Fig. 5

Venn diagram showing the number of identified protein spots revealing a significant increase (↑) or a decrease (↓) in abundance in drought-stressed plants with respect to well-watered plants

Fig. 6

Schematic representation of chloroplast proteins with changed abundance under drought in Tiber (left side) and Starozagorski (right side) with regard to protein functional groups. Proteins with changed abundance after 6 and 13 days were considered. ATPase ATP synthase, CURT1B curvature thylakoid 1B protein, FBA fructose-bisphosphate aldolase, FNR ferredoxin–NADP reductase, HCF136 photosystem II stability/assembly factor HCF136, LHCI photosystem I light-harvesting chlorophyll a/b-binding protein, OEE1 oxygen-evolving enhancer proteins 1, OEE2 oxygen-evolving enhancer proteins 2, PG1 Plastoglobulin-1, PORIN outer plastidial membrane protein porin, Psa-D photosystem I reaction center subunit II, PSI-E photosystem I reaction center subunit IV. Arrows: up increased in abundance under drought, down decreased in abundance under drought, single arrow one protein with changed abundance, bold arrow two or more proteins with changed abundance