Isolation of a cDNA clone for the gamma subunit of the chloroplast ATP synthase of Chlamydomonas reinhardtii: import and cleavage of the precursor protein.

Yu, Lloyd M.; Merchant, Sabeeha; Theg, Steven M.; Selman, Bruce R.

doi:10.1073/pnas.85.5.1369

Cited by 21 publications

(9 citation statements)

References 35 publications

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“…Since that time, we have published numerous studies based on work with that strain (Yu et al, 1988;Long et al, 2008;Castruita et al, 2011). In this study, we resequenced this strain, along with two examples of 2137 from the Chlamydomonas collection (CC-3269 and CC-1021) and one from Robert Spreitzer (2137A+).…”

Section: Mislabeled Strains and Inaccurate Histories Were Identifiedmentioning

confidence: 99%

Chlamydomonas Genome Resource for Laboratory Strains Reveals a Mosaic of Sequence Variation, Identifies True Strain Histories, and Enables Strain-Specific Studies

et al. 2015

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Chlamydomonas reinhardtii is a widely used reference organism in studies of photosynthesis, cilia, and biofuels. Most research in this field uses a few dozen standard laboratory strains that are reported to share a common ancestry, but exhibit substantial phenotypic differences. In order to facilitate ongoing Chlamydomonas research and explain the phenotypic variation, we mapped the genetic diversity within these strains using whole-genome resequencing. We identified 524,640 single nucleotide variants and 4812 structural variants among 39 commonly used laboratory strains. Nearly all (98.2%) of the total observed genetic diversity was attributable to the presence of two, previously unrecognized, alternate haplotypes that are distributed in a mosaic pattern among the extant laboratory strains. We propose that these two haplotypes are the remnants of an ancestral cross between two strains with ;2% relative divergence. These haplotype patterns create a fingerprint for each strain that facilitates the positive identification of that strain and reveals its relatedness to other strains. The presence of these alternate haplotype regions affects phenotype scoring and gene expression measurements. Here, we present a rich set of genetic differences as a community resource to allow researchers to more accurately conduct and interpret their experiments with Chlamydomonas.

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Section: Mislabeled Strains and Inaccurate Histories Were Identifiedmentioning

confidence: 99%

Chlamydomonas Genome Resource for Laboratory Strains Reveals a Mosaic of Sequence Variation, Identifies True Strain Histories, and Enables Strain-Specific Studies

et al. 2015

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“…After the nylon membrane dried, it was treated as described previously (29). Immunoblots were then treated with a rabbit anti--y-subunit antiserum (29 genes (strain nitl-305), and one of which was defective in photophosphorylation, the chloroplast mutant strain cc-2045 (27). The latter control was done to test the predication that -^a mutant deficient in one of the subunits of the ATP synthase complex might be deficient in the others as well.…”

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confidence: 99%

“…The aqueous layer was removed, and 20 RI was spotted onto a nylon membrane for a protein immunoblot. After the nylon membrane dried, it was treated as described previously (29). Immunoblots were then treated with a rabbit anti--y-subunit antiserum (29 genes (strain nitl-305), and one of which was defective in photophosphorylation, the chloroplast mutant strain cc-2045 (27).…”

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confidence: 99%

Isolation and characterization of a Chlamydomonas reinhardtii mutant lacking the gamma-subunit of chloroplast coupling factor 1 (CF1).

Smart¹,

Selman²

1991

Mol. Cell. Biol.

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Several nonphotoautotrophic mutants of Chlamydomonas reinhardtii were generated by transforming strain nitl-305 (cw 15) with exogenous DNA. An enrichment for potential photophosphorylation mutants was performed on medium containing arsenate, and acetate-requiring auxotrophs were then identified by replica plating. Strains containing a potential mutation in the nuclear DNA encoding the chloroplast coupling factor 1 (CFI) -y-subunit (the atpC gene) were first identified serologically with a monospecific antiserum directed against the CF1 fy-subunit polypeptide. Of several mutants isolated, one, designated T1-54, was characterized at the protein, DNA, and RNA levels. Mutant strain T1-54 lacks anti-CF1 y-subunit cross-reacting material, exhibits polymorphism at the atpC locus compared with the parental strain, and lacks the mRNA transcript for the CFI y-subunit. The data are consistent with there being an insertion of exogenous DNA, a deletion of DNA, or both at the 5' end of the gene encoding the CF1 y-subunit.The proton-translocating ATP synthase (chloroplast coupling factor CFO-CF,) complex in chloroplast thylakoid membranes couples the chemical potential energy in the proton motive force to the synthesis of ATP. The complex consists of two discrete sectors, CFo and CF1. CFo is an intrinsic thylakoid membrane complex, contains four different types of polypeptides (designated I through IV), and functions as a proton translocase. CF1 is an extrinsic membrane complex, contains five different types of polypeptides (a through E), and has the catalytic sites for ATP synthesis and hydrolysis (15).The molecular mechanism of CF1-catalyzed ATP synthesis is not well understood. However, ATP synthases from a variety of organisms appear to have similar mechanisms. Consistent with this is the observation that the subunit stoichiometry (a3P3Y8E) of CF1 is rigorously conserved and the amino acid sequences of these subunits are highly conserved, especially those involved in catalysis (a, ,B, and -y) (15). Unlike the reaction mechanism, the regulation of ATP synthase activity depends on both the species and subcellular organelle with which they are associated (1). One unique feature of the chloroplast complex is that its catalytic activity is regulated, in part, by the oxidation state of the y-subunit (9,17).Although there is some diversity in the amino acid composition of the -y-subunit among different plant species, one common feature seems to be a pair of cysteine residues that form an intrapeptide disulfide bridge (18). In the absence of a proton motive force, under conditions in which the catalytic activity of the ATP synthase is latent, the y-subunit cystine bridge is inaccessible to hydrophilic thiol reagents. The development of a proton motive force across the thylakoid membrane not only provides the driving force for ATP synthesis but also causes substantial conformational changes in the coupling factor (18). These conformational changes result in the activation of ATP synthase activity and the * Corresponding author. exp...

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“…It is thought that the 7-subunit in Chlamydomonas is also encoded in a single-copy gene [50]. However, preliminary reports [ 15] suggest that, for another nuclear-encoded ATP synthase subunit, the fi-subunit, there are several genes in the spinach chromosome.…”

Section: Discussionmentioning

confidence: 97%

The ?-subunit of spinach chloroplast ATP synthase: isolation and characterisation of cDNA and genomic clones

Mason¹,

Whitfeld

1990

Plant Mol Biol

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cDNA and genomic clones encoding the complete precursor polypeptide of the gamma-subunit of spinach chloroplast ATP synthase have been isolated and characterised. The longest cDNA (1320 bp), selected from a lambda gt11 spinach cDNA library, encoded a 364 amino acid residue protein (Mr 40,028) that included a putative 41 residue N-terminal transit peptide. All the gamma-subunit cDNAs analysed were derived from the same gene and Southern blot analysis of Bam HI restricted spinach DNA showed only one hybridising band (ca. 15 kb). Analysis of the cloned 15 kb genomic fragment, selected from a lambda EMBL4 spinach DNA library with a cDNA probe, confirmed there was a single gene for the gamma-subunit. Sequencing, primer extension and northern blot analysis showed the gene contains two introns, 1066 and 665 bp in length, a 173 bp 5' untranslated region and a 213 bp 3' untranslated region. A 1450 nucleotide gamma-subunit transcript was detected in RNA from light-grown and dark-grown spinach.

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Isolation of a cDNA clone for the gamma subunit of the chloroplast ATP synthase of Chlamydomonas reinhardtii: import and cleavage of the precursor protein.

Cited by 21 publications

References 35 publications

Chlamydomonas Genome Resource for Laboratory Strains Reveals a Mosaic of Sequence Variation, Identifies True Strain Histories, and Enables Strain-Specific Studies

Chlamydomonas Genome Resource for Laboratory Strains Reveals a Mosaic of Sequence Variation, Identifies True Strain Histories, and Enables Strain-Specific Studies

Isolation and characterization of a Chlamydomonas reinhardtii mutant lacking the gamma-subunit of chloroplast coupling factor 1 (CF1).

The ?-subunit of spinach chloroplast ATP synthase: isolation and characterisation of cDNA and genomic clones

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