Figure 4.

PGRL1 dimer formation and protein accumulation in the absence of NTRC and PGR5. (A) Total leaf proteins were obtained from Col-0, ntrc, pgr5 and pgr5 ntrc after 6 weeks of growth under short-day conditions, precipitated with TCA and alkylated using NEM at the end of the dark period (dark, D) and after a 30-s exposure to light (L, 100 μmol photons m⁻² s⁻¹). Samples were fractionated by SDS-PAGE and subjected to immunoblotting using PGRL1-specific antibody. MW stands for molecular weight scale (KDa). d, c and m indicate dimer, complex and monomer forms of PGRL1, respectively. A representative blot from three experiments is presented, and the ratio of PGRL1 dimers to monomers (d/m), based on the quantification of band intensities, is shown below the panel. (B) Aliquots of total leaf proteins from Arabidopsis wild type (Col-5 and Col-0), ntrc and pgr5 single mutants and pgr5 ntrc double mutant after 6 weeks of growth under short-day conditions were fractionated by SDS-PAGE and subjected to immunoblotting using NTRC-, PGR5- or PGRL1-specific antibodies. Decreasing amounts of Col-0 were loaded. Representative blots from three experiments are presented, as well as the values corresponding to the quantification of the intensity of each band relative to Col-0 100%. In both (A,B), protein samples were adjusted according to fresh weight, and PVDF membranes were stained with Coomassie brilliant blue (C.B.B.) to show protein loading.