Proteomic Analysis of Multiprotein Complexes in the Thylakoid Membrane upon Cadmium Treatment

Abstract

The time course of the thylakoid membranes proteomic profile changes upon cadmium (Cd) addition to hydroponic Spinacia oleracea L. plants has been investigated. Two different proteomic approaches have been used: blue native gel electrophoresis followed by SDS-PAGE (2D BN-SDS-PAGE) and sucrose density gradient ultracentrifugation followed by RP-HPLC. Chlorophyll (Chl) and xanthophylls concentrations, together with ESR and real time PCR measurements, were also performed to get a complete overview of all photosystem changes. Cd only accumulated in basal leaves, that therefore were prevalently investigated for assessment of Cd induced changes. Here, Cd strongly reduced Chl concentration, especially Chl a. During the first 15 days of treatment, native electrophoresis system revealed high sensitivity of PSI to Cd, while minor effects on PSII were observed. Cytochrome b(6)/f and the ATP-synthase complex did not change following the Cd treatment. A significant reduction of antenna proteins of PSI was observed, while PSII antennae were affected to a minor extent, with exception of the isomeric Lhcb1.1 which decreased significantly already at the onset of the treatment. Some PSII core proteins were overexpressed, but showed reduced activity. No new protein was formed and no specific protein disappeared in the photosynthetic apparatus of Cd-treated leaves. Upon removal of Cd, a rapid resynthesis of total Chl and a significant resynthesis of Lhcb1.1 antenna were observed, suggesting that Cd affects specifically the photosynthetic apparatus of spinach basal leaves, replacing other metal ions inside proteins.