Fig. 7
Immunodecoration of thylakoid phosphoproteins with α-P-Thr and α-P-Ser antibodies. Isolated thylakoid samples corresponding to approximately 0.5 and 5 μg of Chl were blotted following separation on SDS PAGE and probed with α-P-Thr (panel A) and α-P-Ser antibodies (panel B), respectively. Experiments included all genotypes created in this work treated with PSII-favoring light and a dark-adapted (equivalent to state 1) wild type sample control. The pattern of the α-P-Thr reaction (panel A) revealed the characteristic thylakoid phosphoproteins D1 (PsbA), D2 (PsbD) and LHCII. Consistent with the results presented in figure 2, the LHCII signal was extremely low in the dark-adapted wild type and was entirely missing in the koLhcb1 and kostn7 genotypes. kostn7 (and to lower extent the koLhcb2 and cB2.1T40V lines) exhibited a stronger Thr phosphorylation of the PSII core complex subunits D1 and D2. Enhanced LHCII Thr phosphorylation was observed in the koLhcb2 and cB.1T40V lines because of persistent plastoquinone reduction and the active state of the STN7 kinase towards the Lhcb1 Thr-38 residue. The α-P-Ser reaction (panel B), instead, revealed an equal phosphorylation level of the LHCII band in all genotypes, except for the koLhcb1 sample where the faint reactive band corresponds to the phosphorylated serine(s) belonging to Lhcb2 polypeptides