Fig. 5: DEIP1 is involved in the accumulation of Cytb₆f, but does not contribute to subunit expression or complex stability.

a BN-PAGE of thylakoids from the wild type (WT) and the deip1-1, deip1-2, and deip1-3 mutants grown under moderate light intensity. Thylakoid samples equivalent to 8 µg of chlorophyll were solubilized with the non-ionic detergent β-DDM, and resolved in a 6-12.5% native gradient gel. The main photosynthetic complexes are indicated as PSII megacomplexes (PSII mc), PSII supercomplexes (PSII sc), Cytb₆f, LHCII assembly (LHCII(a)), LHCII trimer (tLHCII) and monomeric LHCII (mLHCII). The molecular weights (in kDa) of the marker protein bands are given at the left of the gels. n = 3 independent biological replicates. b BN-PAGE of thylakoid samples from the wild type (WT) and the deip1-1, deip1-2 and deip1-3 mutants grown under low light. Thylakoid samples equal to 8 µg of chlorophyll were solubilized with β-DDM, and resolved in a 6–12.5% native gradient gel. The main photosynthetic complexes are indicated as PSII megacomplexes (PSII mc), PSII supercomplexes (PSII sc), PSII dimer and PSI, Cytb₆f, LHCII assembly (LHCII(a)), LHCII trimer (tLHCII) and monomeric LHCII (mLHCII). The molecular weight (in kDa) of the marker protein bands are given at the left of the gels. n = 3 independent biological replicates. c Transcript levels (RNA) and d translation output (ribosome footprint abundances) as analyzed by microarray-based ribosome profiling of deip1-1 and wild-type seedlings grown under low light. Average ribosome footprint and transcript abundances in deip1-1 were calculated for each chloroplast reading frame from three biological replicates, and the mean signal intensities were plotted against the wild type. The labeled psbA reading frame represents the only chloroplast-encoded gene with a significant (P < 0.05) and more than 2-fold change in expression. Statistical significance was determined by LIMMA, P values were adjusted by FDR-test for multiple comparisons and compared to the wild type (Supplementary Data 1). e Immunoblot of total protein extracts from the wild type (WT) and the deip1-2 mutant 4 days after infiltration with water (−) or 1 mM lincomycin (Linc; +). The turnover of the Cytb₆f complex was assessed by immunoblotting with an anti-PetB antibody. The anti-PsbD antibody was used as a positive control for the effectiveness of the lincomycin treatment, and the Coomassie-stained PAA gel (CBB) is shown as loading control. n = 3 independent biological replicates.