The Deletion of petG in Chlamydomonas reinhardtii Disrupts the Cytochrome bf Complex

The cytochrome-b 6 f complex, a key component of the photosynthetic electron transport chain, contains a number of very small protein subunits whose functions are not well defined. Here we have investigated the function of the 31-amino acid PetL subunit encoded in the chloroplast genome in all higher plants. Chloroplast-transformed petL knock-out tobacco plants display no obvious phenotype, suggesting that PetL is not essential for cytochrome b 6 f complex biogenesis and function (Fiebig, A., Stegemann, S., and Bock, R. (2004) Nucleic Acids Res. 32, 3615-3622). We show here that, whereas young mutant leaves accumulate comparable amounts of cytochrome b 6 f complex and have an identical assimilation capacity as wild type leaves, both cytochrome b 6 f complex contents and assimilation capacities of mature and old leaves are strongly reduced in the mutant, indicating that the cytochrome b 6 f complex is less stable than in the wild type. Reduced complex stability was also confirmed by in vitro treatments of isolated thylakoids with chaotropic reagents. Adaptive responses observed in the knockout mutants, such as delayed down-regulation of plastocyanin contents, indicate that plants can sense the restricted electron flux to photosystem I yet cannot compensate the reduced stability of the cytochrome b 6 f complex by adaptive up-regulation of complex synthesis. We propose that efficient cytochrome b 6 f complex biogenesis occurs only in young leaves and that the capacity for de novo synthesis of the complex is very low in mature and aging leaves. Gene expression analysis indicates that the ontogenetic down-regulation of cytochrome b 6 f complex biogenesis occurs at the post-transcriptional level.The Cyt-bf 2 is the smallest of the three thylakoid membraneintrinsic multiprotein complexes of photosynthetic electron transport. Cyt-bf functions as a dimer with a molecular mass of 220 kDa and consists of nine subunits per monomer (1). It catalyzes the rate-limiting step of linear electron transport, oxidizing PQ generated by PSII and reducing PC. PC then diffuses through the thylakoid lumen toward PSI and reduces the photooxidized chlorophyll a dimer in the PSI reaction center, P 700 . The PQ-PC oxidoreductase activity of the Cyt-bf is coupled to the translocation of at least two protons per transported electron into the thylakoid lumen, thus contributing to the proton motif force required for ATP synthesis (2).The Cyt-bf is the predominant point of flux control of linear electron transport (3-6); its amount varies strongly in response to growth conditions and the developmental state of the plant, thus adjusting photosynthetic electron flux to the metabolic demand for ATP and NADPH. The mechanisms underlying these adjustments of Cyt-bf concentration are currently not understood. There are some indications that Cyt-bf is a relatively stable complex with lifetimes at least in the range of several days (7), but how Cyt-bf biogenesis is regulated during plant development is completely unknown. Cyt-bf biogenesis is a highly com...

supporting

confidence: 58%

Knock-out of the Plastid-encoded PetL Subunit Results in Reduced Stability and Accelerated Leaf Age-dependent Loss of the Cytochrome b6f Complex

Schöttler

Flügel

Thiele

et al. 2007

“…Three of them are encoded by chloroplast genes (5), petA (encoding cytochrome f, a c-type cytochrome), petB (encoding cytochrome b 6 ), and petD (encoding subunit IV); whereas the Fe 2 S 2 Rieske protein is encoded by a nuclear gene, PetC (6). In addition, the cytochrome b 6 f complex contains three transmembrane subunits of low molecular mass (Յ4 kDa), the products of the chloroplast genes petG (7)(8)(9) and petL (10) and the nuclear gene PetM (11,12).…”

mentioning

confidence: 99%

Molecular Genetic Identification of a Pathway for Heme Binding to Cytochrome b 6

Kuras¹,

Vitry²,

Choquet³

et al. 1997

100

Heme binding to cytochrome b 6 is resistant, in part, to denaturing conditions that typically destroy the noncovalent interactions between the b hemes and their apoproteins, suggesting that one of two b hemes of holocytochrome b 6 is tightly bound to the polypeptide. We exploited this property to define a pathway for the conversion of apo-to holocytochrome b 6 , and to identify mutants that are blocked at one step of this pathway.

“…Its compositional simplicity relative to the PS II and PS I complexes makes it an attractive model for studying the assembly of chloroplast energy-transducing complexes. The chloroplast-encoded subunits of the cytochrome b 6 f complex include products of petA (cytochrome f), petB (cytochrome b 6 ), petD (subunit IV), and two 4-kDa proteins encoded by the petG and petL genes (15)(16)(17)(18)(19)(20). The nucleus-encoded components include the Rieske Fe-S protein (product of PetC; Ref.…”

mentioning

confidence: 99%

Ccs1, a Nuclear Gene Required for the Post-translational Assembly of Chloroplast c-Type Cytochromes

Inoué

Dreyfuss

Kindle

et al. 1997

Nuclear genes play important regulatory roles in the biogenesis of the photosynthetic apparatus of eukaryotic cells by encoding factors that control steps ranging from chloroplast gene transcription to post-translational processes. However, the identities of these genes and the mechanisms by which they govern these processes are largely unknown. By using glass bead-mediated transformation to generate insertional mutations in the nuclear genome of Chlamydomonas reinhardtii, we have generated four mutants that are defective in the accumulation of the cytochrome b 6 f complex. One of them, strain abf3, also fails to accumulate holocytochrome c 6 . We have isolated a gene, Ccs1, from a C. reinhardtii genomic library that complements both the cytochrome b 6 f and cytochrome c 6 deficiencies in abf3. The predicted protein product displays significant identity with Ycf44 from the brown alga Odontella sinensis, the red alga Porphyra purpurea, and the cyanobacterium Synechocystis strain PCC 6803 (25-33% identity). In addition, we note limited sequence similarity with ResB of Bacillus subtilis and an open reading frame in a homologous operon in Mycobacterium leprae (11-12% identity). On the basis of the pleiotropic c-type cytochrome deficiency in the ccs1 mutant, the predicted plastid localization of the protein, and its relationship to candidate cytochrome biosynthesis proteins in Gram-positive bacteria, we conclude that Ccs1 encodes a protein that is required for chloroplast c-type holocytochrome formation.Studies in the unicellular green alga Chlamydomonas reinhardtii and vascular plants have demonstrated that the proteins required for photosynthesis are encoded by genes that reside in two distinct cellular compartments: the nucleus and the chloroplast (reviewed in Refs.