Quantitative proteomics of a chloroplast SRP54 sorting mutant and its genetic interactions with CLPC1 in Arabidopsis.
cpSRP54 (for chloroplast SIGNAL RECOGNITION PARTICLE54) is involved in cotranslational and posttranslational sorting of thylakoid proteins. The Arabidopsis (Arabidopsis thaliana) cpSRP54 null mutant, ffc1-2, is pale green with delayed development. Western-blot analysis of individual leaves showed that the SRP sorting pathway, but not the SecY/E translocon, was strongly down-regulated with progressive leaf development in both wild-type and ffc1-2 plants. To further understand the impact of cpSRP54 deletion, a quantitative comparison of ffc2-1 was carried out for total leaf proteomes of young seedlings and for chloroplast proteomes of fully developed leaves using stable isotope labeling (isobaric stable isotope labeling and isotope-coded affinity tags) and two-dimensional gels. This showed that cpSRP54 deletion led to a change in light-harvesting complex composition, an increase of PsbS, and a decreased photosystem I/II ratio. Moreover, the cpSRP54 deletion led in young leaves to up-regulation of thylakoid proteases and stromal chaperones, including ClpC. In contrast, the stromal protein homeostasis machinery returned to wild-type levels in mature leaves, consistent with the developmental down-regulation of the SRP pathway. A differential response between young and mature leaves was also found in carbon metabolism, with an up-regulation of the Calvin cycle and the photorespiratory pathway in peroxisomes and mitochondria in young leaves but not in old leaves. The Calvin cycle was down-regulated in mature leaves to adjust to the reduced capacity of the light reaction, while reactive oxygen species defense proteins were up-regulated. The significance of ClpC up-regulation was confirmed through the generation of an ffc2-1 clpc1 double mutant. This mutant was seedling lethal under autotrophic conditions but could be partially rescued under heterotrophic conditions.
Rutschow
Heidi
H
Department of Plant Biology, Cornell University, Ithaca, New York 14853, USA.
Ytterberg
A Jimmy
AJ
Friso
Giulia
G
Nilsson
Robert
R
van Wijk
Klaas J
KJ
D017360:Arabidopsis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D060365:Chloroplast Proteins; D002736:Chloroplasts / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D049071:Endopeptidase Clp / Q000235:genetics / Q000378:metabolism*; D006706:Homeostasis; D007525:Isoelectric Focusing; D007554:Isotopes / Q000378:metabolism; D008027:Light; D045342:Light-Harvesting Protein Complexes / Q000378:metabolism; D013058:Mass Spectrometry; D009154:Mutation; D009584:Nitrogen / Q000378:metabolism; D010788:Photosynthesis; D045332:Photosystem II Protein Complex / Q000378:metabolism; D018515:Plant Leaves / Q000378:metabolism; D010940:Plant Proteins / Q000096:biosynthesis; D021381:Protein Transport; D040901:Proteomics; D036226:Seedlings / Q000254:growth & development / Q000378:metabolism; D018271:Signal Recognition Particle / Q000235:genetics / Q000378:metabolism*; D020524:Thylakoids / Q000378:metabolism
D029681:Arabidopsis Proteins; D060365:Chloroplast Proteins; C083707:FFC protein, Arabidopsis; D007554:Isotopes; D045342:Light-Harvesting Protein Complexes; D045332:Photosystem II Protein Complex; D010940:Plant Proteins; D018271:Signal Recognition Particle; D049071:Endopeptidase Clp; D009584:Nitrogen