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• W1973492092 abstract "Three reading frames called ccmFN1, ccmFN2, and ccmFc are found in the mitochondrial genome of Arabidopsis. These sequences are similar to regions of the bacterial gene ccmF involved in cytochrome c maturation. ccmF genes are always absent from animal and fungi genomes but are found in mitochondrial genomes of land plant and several evolutionary distant eukaryotes. In Arabidopsis, ccmFN2 despite the absence of a classical initiation codon is not a pseudo gene. The 3 ccmF genes of Arabidopsis are expressed at the protein level. Their products are integral proteins of the mitochondrial inner membrane with in total 11 to 13 predicted transmembrane helices. The conserved WWD domain of CcmFN2 is localized in the inter membrane space. The 3 CcmF proteins are all detected in a high molecular mass complex of 500 kDa by Blue Native PAGE. Direct interaction between CcmFN2 and both CcmFN1 and CcmFC is shown with the yeast two-hybrid split ubiquitin system, but no interaction is observed between CcmFN1 and CcmFC. Similarly, interaction is detected between CcmFN2 and apocytochrome c but also with apocytochrome c1. Finally, CcmFN1 and CcmFN2 both interact with CCMH previously shown to interact as well with cytochrome c. This strengthens the hypothesis that CcmF and CCMH make a complex that performs the assembly of heme with c-type apocytochromes in plant mitochondria. Three reading frames called ccmFN1, ccmFN2, and ccmFc are found in the mitochondrial genome of Arabidopsis. These sequences are similar to regions of the bacterial gene ccmF involved in cytochrome c maturation. ccmF genes are always absent from animal and fungi genomes but are found in mitochondrial genomes of land plant and several evolutionary distant eukaryotes. In Arabidopsis, ccmFN2 despite the absence of a classical initiation codon is not a pseudo gene. The 3 ccmF genes of Arabidopsis are expressed at the protein level. Their products are integral proteins of the mitochondrial inner membrane with in total 11 to 13 predicted transmembrane helices. The conserved WWD domain of CcmFN2 is localized in the inter membrane space. The 3 CcmF proteins are all detected in a high molecular mass complex of 500 kDa by Blue Native PAGE. Direct interaction between CcmFN2 and both CcmFN1 and CcmFC is shown with the yeast two-hybrid split ubiquitin system, but no interaction is observed between CcmFN1 and CcmFC. Similarly, interaction is detected between CcmFN2 and apocytochrome c but also with apocytochrome c1. Finally, CcmFN1 and CcmFN2 both interact with CCMH previously shown to interact as well with cytochrome c. This strengthens the hypothesis that CcmF and CCMH make a complex that performs the assembly of heme with c-type apocytochromes in plant mitochondria. Plant mitochondrial genomes differ from other mitochondrial genomes by their larger size (1Ward B.L. Anderson R.S. Bendich A.J. Cell. 1981; 26: 793-803Abstract Full Text PDF Scopus (292) Google Scholar). However, even if plant mitochondrial genomes are at least 15 times larger than their animal counterparts, they do not encode many more genes, e.g. 31 protein coding genes in Arabidopsis (2Unseld M. Marienfeld J.R. Brandt P. Brennicke A. Nat. Genet. 1997; 15: 57-61Crossref PubMed Scopus (721) Google Scholar) and 13 in human (3Anderson S. Bankier A.T. Barrell B.G. de Bruijn M.H. Coulson A.R. Drouin J. Eperon I.C. Nierlich D.P. Roe B.A. Sanger F. Schreier P.H. Smith A.J. Staden R. Young I.G. Nature. 1981; 290: 457-465Crossref PubMed Scopus (7644) Google Scholar). Mitochondrial gene contents are highly conserved. The genes encoded include components of essential mitochondrial functions, i.e. subunits of respiratory complexes and components of the mitochondrial translation apparatus. A major difference in gene content between plant and animal mitochondria resides in the occurrence in plants of ribosomal protein genes (2Unseld M. Marienfeld J.R. Brandt P. Brennicke A. Nat. Genet. 1997; 15: 57-61Crossref PubMed Scopus (721) Google Scholar) and of ccm genes (for cytochrome c maturation) (4Gonzalez D.H. Bonnard G. Grienenberger J.M. Curr. Genet. 1993; 21: 248-255Crossref Scopus (61) Google Scholar, 5Schuster W. Combettes B. Flieger K. Brennicke A. Mol. Gen. Genet. 1993; 239: 49-57Crossref PubMed Scopus (47) Google Scholar). These genes are similar to bacterial sequences found to be involved in the biogenesis of c-type cytochromes by genetic studies (6Kranz R.G. J. Bacteriol. 1989; 171: 456-464Crossref PubMed Google Scholar). In Escherichia coli, 8 ccm genes (ccmA–H) are encoded at 1 locus in a single operon (7Beckman D.L. Trawick D.R. Kranz R.G. Genes Dev. 1992; 6: 268-283Crossref PubMed Scopus (147) Google Scholar). On the contrary, in plants, ccm genes are encoded by both the nucleus and mitochondria. In Arabidopsis, CCMA, CCME, and CCMH genes are encoded in the nucleus (8Rayapuram N. Hagenmuller J. Grienenberger J.M. Giege P. Bonnard G. J. Biol. Chem. 2007; 282: 21015-21023Abstract Full Text Full Text PDF PubMed Scopus (37) Google Scholar, 9Meyer E.H. Giegé P. Gelhaye E. Rayapuram N. Ahuja U. Thony-Meyer L. Grienenberger J.M. Bonnard G. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 16113-16118Crossref PubMed Scopus (70) Google Scholar), whereas ccmB, ccmC, ccmFN1, ccmFN2, and ccmFc are encoded in the mitochondrial genome, at 5 different loci (2Unseld M. Marienfeld J.R. Brandt P. Brennicke A. Nat. Genet. 1997; 15: 57-61Crossref PubMed Scopus (721) Google Scholar). The latter 3 sequences are similar to different domains of the bacterial ccmF. ccmFN2 is not an “open reading frame” because it lacks a classical ATG initiation codon. However, transcription has been established for all the ccm sequences encoded in Arabidopsis mitochondria (10Giegé P. Brennicke A. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 15324-15329Crossref PubMed Scopus (319) Google Scholar, 11Raczynska K.D. Le Ret M. Rurek M. Bonnard G. Augustyniak H. Gualberto J.M. FEBS Lett. 2006; 580: 5641-5646Crossref PubMed Scopus (41) Google Scholar). Furthermore, ccm transcripts have the highest RNA editing rate within the Arabidopsis mitochondrial transcriptome, with 26 editing sites/kb on average (10Giegé P. Brennicke A. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 15324-15329Crossref PubMed Scopus (319) Google Scholar). Notable features are that RNA editing shortens the reading frame of ccmFC by introducing a termination codon (12Giegé P. Rayapuram N. Meyer E.H. Grienenberger J.M. Bonnard G. FEBS Lett. 2004; 563: 165-169Crossref PubMed Scopus (23) Google Scholar) and that RNA editing is necessary for the conservation of the WWD motif of CcmFN (4Gonzalez D.H. Bonnard G. Grienenberger J.M. Curr. Genet. 1993; 21: 248-255Crossref Scopus (61) Google Scholar, 5Schuster W. Combettes B. Flieger K. Brennicke A. Mol. Gen. Genet. 1993; 239: 49-57Crossref PubMed Scopus (47) Google Scholar). Cytochromes of c-type are ubiquitous electron transporters. They are defined by the covalent attachment of heme C prosthetic groups to apocytochromes via two thioether bonds between vinyl groups on the heme and conserved cysteines on the protein. In mitochondria, these proteins are essential components of the respiratory chain involved in the generation of cellular energy. Two forms of these key proteins co-exist. They are present as soluble proteins, i.e. cytochrome c in the mitochondrial inter-membrane space and also as membrane proteins, i.e. cytochrome c1 integrated into respiratory complex III, the cytochrome c reductase in the mitochondrial inner membrane. The term “cytochrome c maturation” refers to the processes leading to the covalent attachment of heme to the apocytochromes. Different pathways have been described for this maturation (13Kranz R. Lill R. Goldman B. Bonnard G. Merchant S. Mol. Microbiol. 1998; 29: 383-396Crossref PubMed Scopus (237) Google Scholar). In yeast and animal mitochondria, a maturation process called “system III” has evolved. It involves one or two cytochrome c heme lyases (CCHL proteins) (14Dumont M.E. Ernst J.F. Hampsey D.M. Sherman F. EMBO J. 1987; 6: 235-241Crossref PubMed Scopus (165) Google Scholar). “System I” is found in α, in most γ proteobacteria, and in some β proteobacteria, in deinococci and archaea (6Kranz R.G. J. Bacteriol. 1989; 171: 456-464Crossref PubMed Google Scholar, 15Page M.D. Ferguson S.J. Mol. Microbiol. 1995; 15: 307-318Crossref PubMed Scopus (47) Google Scholar), and in mitochondria of land plants and of some protists and algae (16Lang B.F. Burger G. O'Kelly C.J. Cedergren R. Golding G.B. Lemieux C. Sankoff D. Turmel M. Gray M.W. Nature. 1997; 387: 493-496Crossref PubMed Scopus (487) Google Scholar, 17Giegé P. Grienenberger J.M. Bonnard G. Mitochondrion. 2008; 8: 61-73Crossref PubMed Scopus (82) Google Scholar). The proteins used by cytochrome c maturation system I are termed Ccm proteins. Among them, in plants, CCMA and CcmB form an ABC transporter (8Rayapuram N. Hagenmuller J. Grienenberger J.M. Giege P. Bonnard G. J. Biol. Chem. 2007; 282: 21015-21023Abstract Full Text Full Text PDF PubMed Scopus (37) Google Scholar), CCME is a heme chaperone that was proposed to bind heme covalently in the intermembrane space where its covalent attachment with apocytochromes takes place (18Spielewoy N. Schulz H. Grienenberger J.M. Thony-Meyer L. Bonnard G. J. Biol. Chem. 2001; 276: 5491-5497Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar). Finally, CCMH interacts with apocytochrome c and can reduce cysteines of apocytochrome c that are required to be reduced for the ligation with heme to occur (9Meyer E.H. Giegé P. Gelhaye E. Rayapuram N. Ahuja U. Thony-Meyer L. Grienenberger J.M. Bonnard G. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 16113-16118Crossref PubMed Scopus (70) Google Scholar). In bacteria, CcmABCDE are involved in the delivery of heme to its site of assembly (19Goldman B.S. Beck D.L. Monika E.M. Kranz R.G. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 5003-5008Crossref PubMed Scopus (93) Google Scholar, 20Thöny-Meyer L. Biochim. Biophys. Acta. 2000; 1459: 316-324Crossref PubMed Scopus (76) Google Scholar, 21Ren Q. Thony-Meyer L. J. Biol. Chem. 2001; 276: 32591-32596Abstract Full Text Full Text PDF PubMed Scopus (55) Google Scholar). Co-immunoprecipitation experiments show direct associations between CcmC and CcmE and between CcmF and both CcmE and CcmH (21Ren Q. Thony-Meyer L. J. Biol. Chem. 2001; 276: 32591-32596Abstract Full Text Full Text PDF PubMed Scopus (55) Google Scholar, 22Ren Q. Ahuja U. Thony-Meyer L. J. Biol. Chem. 2002; 277: 7657-7663Abstract Full Text Full Text PDF PubMed Scopus (88) Google Scholar). Interestingly, a tryptophan-rich domain, called the WWD domain is found in both CcmC and CcmF (CcmC and CcmFN2 in Arabidopsis). This motif has been proposed to serve as a hydrophobic platform for heme binding (19Goldman B.S. Beck D.L. Monika E.M. Kranz R.G. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 5003-5008Crossref PubMed Scopus (93) Google Scholar) but other studies in bacteria suggest that this motif is needed for the interaction between CcmC and CcmE (22Ren Q. Ahuja U. Thony-Meyer L. J. Biol. Chem. 2002; 277: 7657-7663Abstract Full Text Full Text PDF PubMed Scopus (88) Google Scholar, 23Schulz H. Pellicioli E.C. Thöny-Meyer L. Mol. Microbiol. 2000; 37: 1379-1388Crossref PubMed Scopus (58) Google Scholar). In both bacteria and plant mitochondria, CcmF is predicted to be the final player of cytochrome c maturation. It is believed to be the site where heme and c-type apocytochromes are bound. However, no interaction between CcmF and c-type cytochromes had been established. Moreover, no interaction had been detected between cytochrome c1 and any Ccm protein. Here, we describe the submitochondrial localization of the 3 Arabidopsis CcmF proteins. We find them in a 500-kDa complex and show that they can be in direct interaction. Furthermore, CcmFN1 and CcmFN2 can bind CCMH and CcmFN2 is able to interact with both apocytochrome c and apocytochrome c1. These results together with previous work enable to propose a mechanistic model for the maturation of c-type cytochromes. Phylogenic Analysis—CcmF protein sequences were aligned with the Muscle version 3.52 program (24Edgar R.C. BMC Bioinformatics. 2004; 5: 113Crossref PubMed Scopus (5985) Google Scholar). The poorly aligned and too divergent positions were removed from alignment using Gblocks 0.91b program (25Talavera G. Castresana J. Syst. Biol. 2007; 56: 564-577Crossref PubMed Scopus (3515) Google Scholar). A phylogenetic analysis was performed using PhyML (26Guindon S. Gascuel O. Syst. Biol. 2003; 52: 696-704Crossref PubMed Scopus (14248) Google Scholar) and 100 boostraps support. The unrooted tree (Fig. 1A) was drawn with Treedyn (27Chevenet F. Brun C. Banuls A.L. Jacq B. Christen R. BMC Bioinformatics. 2006; 7: 439Crossref PubMed Scopus (819) Google Scholar). Cell Fractionation and Purification of Mitochondria—Arabidopsis thaliana var. Landsberg erecta suspension cultures were maintained in a Gamborg G0210 basal medium containing 1 mg/liter 2.4-Dichlorophenoxy acetic acid and 2% (w/v) sucrose (pH 5.8). Cell cultures of 100 ml were maintained in 250-ml conical flasks in the dark at 22 °C on a rotary shaker at 150 rpm. Every 7 days, 10 ml of the culture was subcultured into 100 ml of fresh media. Five-day-old cultures were used for the preparation of sub-cellular fractions as described previously (18Spielewoy N. Schulz H. Grienenberger J.M. Thony-Meyer L. Bonnard G. J. Biol. Chem. 2001; 276: 5491-5497Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar). For mitochondrial extraction, cells were harvested by filtration through 100μm nylon mesh and mitochondria were extracted and purified by Percoll gradient centrifugation as described previously (28Giegé P. Sweetlove L. Leaver C. Plant Mol. Biol. Reporter. 2003; 21: 133-144Crossref Scopus (27) Google Scholar). Mitochondria were fractionated into mitoplast, membrane, and soluble fractions as described previously (18Spielewoy N. Schulz H. Grienenberger J.M. Thony-Meyer L. Bonnard G. J. Biol. Chem. 2001; 276: 5491-5497Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar). The membrane fraction was subjected to alkaline treatment (0.1 m Na2CO3, pH 11.5, for 30 min at 4 °C) to extract peripheral proteins. Blue Native PAGE—Mitochondrial membrane complexes were resolved by Blue Native PAGE in the first dimension followed by SDS-PAGE in the second dimension as described previously (29Giegé P. Sweetlove L.J. Cognat V. Leaver C.J. Plant Cell. 2005; 17: 1497-1512Crossref PubMed Scopus (121) Google Scholar). 400 μg of mitochondrial membrane proteins were resuspended in ACA750 buffer containing 750 mm amino dicaproic acid, 50 mm bis-Tris, 4The abbreviations used are: bis-Tris2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diolTricineN-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycineADactivation domain. and 0.5 mm Na2EDTA (pH 7.0). Protein complexes were solubilized with digitonin, 5/1 detergent/protein (w/w) for 30 min on ice, centrifuged at 100,000 × g for 15 min at 4 °C, and 5% (v/v) Serva blue solution (750 mm ACA750 solution, 5% (w/v) Serva Blue G250) was added to the supernatant. For the first dimension, mitochondrial complexes were separated on 5–13% acrylamide (in 0.5 m amino dicaproic acid, 50 mm bis-Tris, pH 7.0, buffer) gradient gels, with 50 mm bis-Tris (pH 7.0) anode buffer and 50 mm Tricine, 15 mm bis-Tris, 0.02% (v/v) Serva Blue G-250 (pH 7.0) cathode buffer. Electrophoresis was carried out overnight at 5 mA. Gel lanes were cut out and denatured for 1 h at room temperature in 50 mm Tris-HCl (pH 6.8), 1% (w/v) SDS, and 1% (v/v) β-mercaptoethanol. For the second dimension, subunits of the various complexes were separated by SDS-PAGE according to the method of Schagger and von Jagow (30Schägger H. von Jagow G. Anal. Biochem. 1991; 199: 223-231Crossref PubMed Scopus (1910) Google Scholar). 2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diol N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine activation domain. Antibodies and Immunodetection—Polyclonal antibodies were raised in rabbits against LDAWRFRGSREGKRTH for CcmFN1 (amino acids 189–204) and MKQQASVRRTYKKEM for CcmFN2 (amino acids 163–177) peptides coupled to ovalbumin, and PSLLRQLQKDKLRWN (amino acids 428–442) for CcmFC coupled to KLH. SDS-PAGE and Blue Native gels were transferred to PVDF Immobilon-P membranes (Millipore). After blocking, membranes were incubated overnight with serum at dilutions of 1/20,000 for CcmFN1 and CcmFN2, 1/500 for purified CcmFC antibodies, 1/100,000 for wheat Nad9 (31Lamattina L. Gonzalez D. Gualberto J. Grienenberger J.M. Eur. J. Biochem. 1993; 217: 831-838Crossref PubMed Scopus (136) Google Scholar), 1/10,000 for tobacco MnSOD (32Bowler C. Alliotte T. Van den Bulcke M. Bauw G. Vandekerckhove J. Van Montagu M. Inze D. Proc. Natl. Acad. Sci. U. S. A. 1989; 86: 3237-3241Crossref PubMed Scopus (55) Google Scholar), 1/40,000 for Arabidopsis TRXh3 (33Laloi C. Rayapuram N. Chartier Y. Grienenberger J.M. Bonnard G. Meyer Y. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 14144-14149Crossref PubMed Scopus (223) Google Scholar), 1/20,000 for Chlamydomonas LHC II (34Vallon O. Bulte L. Dainese P. Olive J. Bassi R. Wollman F.A. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 8262-8266Crossref PubMed Scopus (145) Google Scholar), 1/50,000 for yeast Cytc1 (obtained from G. Schatz, Basel), and 1/5,000 for Arabidopsis CCME (18Spielewoy N. Schulz H. Grienenberger J.M. Thony-Meyer L. Bonnard G. J. Biol. Chem. 2001; 276: 5491-5497Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar). Goat anti-rabbit antibodies conjugated with horseradish peroxidase (Amersham Biosciences) were used as secondary antibodies and visualized with enhanced chemiluminescent reagents (Amersham Biosciences). Protein Interactions by the Split Ubiquitin System—cDNA fragments representing the full-length Arabidopsis CcmFN1, CcmFN2, CcmFC, apocytochrome c (At1g22840), apocytochrome c1 (At5g40810), and CCMH (9Meyer E.H. Giegé P. Gelhaye E. Rayapuram N. Ahuja U. Thony-Meyer L. Grienenberger J.M. Bonnard G. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 16113-16118Crossref PubMed Scopus (70) Google Scholar) were amplified by PCR with oligonucleotides containing B1 and B2 recombination sites necessary for the entry of the cDNA fragments into vectors by in vivo recombination in yeast (sequences available upon request to authors). PCR products were recombined with linearized pMet-X-Cub-PLV, pX-NubG, and pNubG-X vectors also containing B1 and B2 sites (35Obrdlik P. El-Bakkoury M. Hamacher T. Cappellaro C. Vilarino C. Fleischer C. Ellerbrok H. Kamuzinzi R. Ledent V. Blaudez D. Sanders D. Revuelta J.L. Boles E. Andre B. Frommer W.B. Proc. Natl. Acad. Sci. U. S. A. 2004; 101: 12242-12247Crossref PubMed Scopus (243) Google Scholar) to obtain “XCub, XNub, and NubX” constructs. XCub constructs express a fusion between the protein of interest, the C-terminal domain of ubiquitin, and the chimeric transcription factor PLV consisting of protein A, LexA, and VP16 under control of the methionine repressible pMET25 promoter. XNub and NubX constructs express N-terminal and C-terminal fusions of the protein of interest with the N-terminal domain of ubiquitin (containing a point mutation that abolishes its ability to associate spontaneously with Cub) and the 3HA epitope, under the control of the pADH promoter. XCub constructs were transformed by heat shock in the haploid yeast strain AP4 (MATa), and XNub and NubX constructs in the haploid yeast strain AP5 (MATα) according to standard methods (36Sambrook J. Fritsch E.F. Maniatis T. Molecular Cloning: A Laboratory Manual. 2nd Ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY1989Google Scholar). Transformation in yeast was controlled by the growth on –Leu and –Trp media (-LW). Protein interaction was monitored by the expression of the reporter genes ADE2, HIS3, and lacZ. The expression of ADE2 and HIS3 was visualized by the growth on –Ade–His media (-AH). The expression of lacZ was followed by measuring at OD420 the accumulation of the product metabolized by β-galactosidase with 2.2 mm 2-nitrophenyl β-d-galactopyranoside (Sigma) as substrate. Yeast Two-hybrid Assays—cDNA fragments representing domain 3 (residues 50–80) and domain 5 (residues 240–382) of CcmFN1, domain 2 of CcmFN2 (residues 40–165), and domain 6 of CcmFC (residues 262–409) were cloned in pGBKT7 and pGADT7 vectors (Clontech) to express proteins fused to the DNA-binding domain and the activation domain (AD) of GAL4. Interaction assays were performed as previously described (9Meyer E.H. Giegé P. Gelhaye E. Rayapuram N. Ahuja U. Thony-Meyer L. Grienenberger J.M. Bonnard G. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 16113-16118Crossref PubMed Scopus (70) Google Scholar). ccmF in Mitochondrial Genomes—In plants, ccm genes were inherited from the α proteobacterial ancestor of mitochondria. Presently, some lineages have retained Ccm proteins, their genes are encoded in both mitochondria and the nucleus. Still, ccmF genes are only found in mitochondria. A high number of complete mitochondrial genomes covering the entire diversity of eukaryotic lineages have become available (37Gray M.W. Lang B.F. Burger G. Annu. Rev. Genet. 2004; 38: 477-524Crossref PubMed Scopus (252) Google Scholar). We have searched mitochondrial genomes deposited in GOBASE (rel. 21, 2008) (gobase.bcm.umontreal.ca/) to try to understand the evolutionary history of ccmF genes among eukaryotes. ccmF, similar to all ccm genes has been lost from all holozoa (including animals) and from fungi. In viridiplantae, ccmF genes are absent from all the 10 mitochondrial genomes of chlorophytes available (including Mesostigma). In land plants including the bryophyte Marchantia polymorpha and the moss Physcomitrella patens fully sequenced to date (38Oda K. Kohchi T. Ohyama K. Biosci. Biotech. Biochem. 1992; 56: 132-135Crossref PubMed Scopus (33) Google Scholar, 39Terasawa K. Odahara M. Kabeya Y. Kikugawa T. Sekine Y. Fujiwara M. Sato N. Mol. Biol. Evol. 2007; 24: 699-709Crossref PubMed Scopus (110) Google Scholar), ccmF genes are always found. ccmF is also found in a wide array of other lineages: in charophytes, Rhodophytes, alveolates, Discicristata, and jako-bids (17Giegé P. Grienenberger J.M. Bonnard G. Mitochondrion. 2008; 8: 61-73Crossref PubMed Scopus (82) Google Scholar) (Fig. 1A). For all these lineages, ccmF is found in some species but not in all of them. Altogether, the presence of ccmF genes in evolutionary distant lineages, in some mitochondrial genomes, but not in all of them in the respective lineages, suggest that ccmF genes were lost from mitochondrial genomes at several time points during the evolution of eukaryotes. In particular in viridiplantae, the sequence data available suggests that they were lost at least twice: at the separation between streptophytes and chlorophytes and during the evolution of charophytes. The Orthologue of Bacterial ccmF Is Encoded by Multiple Genes in Land Plants—In land plants, beyond the 16 fully sequenced mitochondrial genomes, ccmF genes are also found in all the incomplete genomic sequences available, in mitochondrial genomes of evolutionary distant organisms, such as e.g. Ginkgo or Amborella. Thus it appears to be strictly conserved in land plants. However, in this lineage, contrary to bacteria, ccmF is not encoded by single genes. ccmF has been split in multiple genes, each orthologue to different domains of the bacterial ccmF (Fig. 1B). This separation is differential according to species. In most plants, ccmF is split into 2 genes, e.g. in wheat (4Gonzalez D.H. Bonnard G. Grienenberger J.M. Curr. Genet. 1993; 21: 248-255Crossref Scopus (61) Google Scholar, 12Giegé P. Rayapuram N. Meyer E.H. Grienenberger J.M. Bonnard G. FEBS Lett. 2004; 563: 165-169Crossref PubMed Scopus (23) Google Scholar). In M. polymorpha, the part encoding the C-terminal end of ccmF is further split in 2 genes (40Oda K. Yamato K. Ohta E. Nakamura Y. Takemura M. Nozato N. Akashi K. Ohyama K. Nucleic Acids Res. 1992; 20: 3773-3777Crossref PubMed Scopus (35) Google Scholar). In Brasicacea such as Arabidopsis the part encoding the N-terminal end of ccmF is encoded by 2 genes (41Handa H. Bonnard G. Grienenberger J.M. Mol. Gen. Genet. 1996; 252: 292-302Crossref PubMed Scopus (26) Google Scholar, 42Menassa R. El-Rouby N. Brown G.G. Curr. Genet. 1997; 31: 70-79Crossref PubMed Scopus (16) Google Scholar). Thus, the 3 Arabidopsis ccmF genes are named ccmFN1, ccmFN2, and ccmFC. ccmFN2 that encodes the highly conserved WWD domain does not start by an ATG initiation codon (2Unseld M. Marienfeld J.R. Brandt P. Brennicke A. Nat. Genet. 1997; 15: 57-61Crossref PubMed Scopus (721) Google Scholar). The 3 Arabidopsis ccmF sequences have an estimated 1027 codons, as compared with the 647 of ccmF in E. coli. This difference is explained by the presence of large insertions in plant between the regions of high sequence conservation between prokaryotes and eukaryotes (Fig. 1B). The Three Arabidopsis ccmF Genes Are Translated—Gene expression has been documented for ccmF genes for various plant mitochondrial genomes. In wheat, the translation of CcmFN and CcmFC has also been reported (4Gonzalez D.H. Bonnard G. Grienenberger J.M. Curr. Genet. 1993; 21: 248-255Crossref Scopus (61) Google Scholar, 12Giegé P. Rayapuram N. Meyer E.H. Grienenberger J.M. Bonnard G. FEBS Lett. 2004; 563: 165-169Crossref PubMed Scopus (23) Google Scholar). In Arabidopsis, the absence of the ATG initiation codon in ccmFN2 suggested that ccmFN2 could be a pseudo gene. However, transcription and RNA editing had been observed for the 3 Arabidopsis ccmF genes (10Giegé P. Brennicke A. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 15324-15329Crossref PubMed Scopus (319) Google Scholar). Still, in plant mitochondria, many sequences and reading frames are transcribed but not translated (44Giegé P. Hoffmann M. Binder S. Brennicke A. EMBO Rep. 2000; 1: 164-170Crossref PubMed Scopus (74) Google Scholar, 45Holec S. Lange H. Kuhn K. Alioua M. Borner T. Gagliardi D. Mol. Cell. Biol. 2006; 26: 2869-2876Crossref PubMed Scopus (90) Google Scholar). Similarly, RNA editing has been observed for pseudogene transcripts (46Giegé P. Knoop V. Brennicke A. Curr. Genet. 1998; 34: 313-317Crossref PubMed Scopus (21) Google Scholar). A recent study where Arabidopsis mitochondrial transcript ends were mapped by circular reverse transcriptase-PCR shows that no ATG codon has been brought in 5′ of ccmFN2 by trans-splicing (47Forner J. Weber B. Thuss S. Wildum S. Binder S. Nucleic Acids Res. 2007; 35: 3676-3692Crossref PubMed Scopus (108) Google Scholar), nor RNA editing (10Giegé P. Brennicke A. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 15324-15329Crossref PubMed Scopus (319) Google Scholar). Thus we had to test whether the 3 ccmF were indeed translated and that ccmFN2 was not a pseudo gene. Therefore, we raised antibodies against peptides representing the 3 CcmF. The antibodies were used to probe Arabidopsis cell fractions. Single bands were detected in the mitochondrial fractions only, at 42, 30, and 60 kDa for CcmFN1, CcmFN2, and CcmFC antibodies, respectively (supplementary Fig. S1A). These sizes correspond to the calculated sizes of the proteins, 382, 203, and 442 amino acids long, respectively. For the CcmFN2 reading frame, we calculated its predicted size starting from a GUG codon (see below). Thus, it confirms that the 3 CcmF proteins, including CcmFN2, are indeed translated. The quality of cell fractionation was assessed with antibodies directed against cytosolic, chloroplastic, and mitochondrial proteins. The exact position of CcmFN2 translation start, however, remains uncertain. A precise answer would be brought by the direct N-terminal sequencing of CcmFN2 purified from Arabidopsis mitochondria. The antibodies were also used with mitochondrial extracts from other plants. The CcmFN2 antibodies detect bands of 53 and 58 kDa for wheat mitochondria. The 58-kDa protein is only detected in a soluble protein fraction thus suggesting that it is not CcmFN, whereas the band of 53 kDa corresponds to the calculated size of wheat CcmFN and is mostly found in a membrane fraction. For CcmFC, as expected, a band of 60 kDa is detected in both wheat and Arabidopsis mitochondrial extracts. These results confirm at the protein level the differential organization of ccmF genes between wheat and Arabidopsis (supplementary Fig. S1B). Submitochondrial Localization and Topology of CcmF Proteins—The submitochondrial localization, the biochemical properties, and the topologies of the 3 Arabidopsis CcmF proteins were investigated to establish whether they correspond or not to the predicted function of CcmF. Mitochondria were fractionated into soluble and membrane fractions. The antibodies detected signals in the membrane fractions only for CcmFN1, CcmFN2, and CcmFC (Fig. 2A). Membrane proteins were further fractionated into peripheral and intrinsic membrane proteins by alkaline treatment. The 3 CcmF proteins were all found to be intrinsic membrane proteins. In bacteria, the topology of Rhodobacter CcmF was determined by fusing PhoA and LacZ to the predicted soluble domains of CcmF and by measuring the corresponding enzymatic activities (19Goldman B.S. Beck D.L. Monika E.M. Kranz R.G. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 5003-5008Crossref PubMed Scopus (93) Google Scholar, 48Rios-Velazquez C. Coller R. Donohue T.J. J. Bacteriol. 2003; 185: 422-431Crossref PubMed Scopus (12) Google Scholar). We used this data as well the ConPred II transmembrane and the topology prediction program (49Arai M. Mitsuke H. Ikeda M. Xia J.X. Kikuchi T. Satake M. Shimizu T. Nucleic Acids Res. 2004; 32: W390-W393Crossref PubMed Scopus (189) Google Scholar) to build topology models for the 3 Arabidopsis CcmF proteins (Fig. 2B). CcmFN1 could have 3 to 5" @default.
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• W1973492092 title "The Three Mitochondrial Encoded CcmF Proteins Form a Complex That Interacts with CCMH and c-Type Apocytochromes in Arabidopsis" @default.
• W1973492092 cites W1519266993 @default.
• W1973492092 cites W1521650410 @default.
• W1973492092 cites W1574378255 @default.
• W1973492092 cites W1606103640 @default.
• W1973492092 cites W1966493111 @default.
• W1973492092 cites W1966662386 @default.
• W1973492092 cites W1966824234 @default.
• W1973492092 cites W1967581443 @default.
• W1973492092 cites W1969538062 @default.
• W1973492092 cites W1981126897 @default.
• W1973492092 cites W1985718139 @default.
• W1973492092 cites W1986248185 @default.
• W1973492092 cites W1987135257 @default.
• W1973492092 cites W1989835605 @default.
• W1973492092 cites W1990122515 @default.
• W1973492092 cites W1995891760 @default.
• W1973492092 cites W1997039036 @default.
• W1973492092 cites W2005678783 @default.
• W1973492092 cites W2011237790 @default.
• W1973492092 cites W2012351178 @default.
• W1973492092 cites W2019116905 @default.
• W1973492092 cites W2019484805 @default.
• W1973492092 cites W2019769257 @default.
• W1973492092 cites W2021795499 @default.
• W1973492092 cites W2029702158 @default.
• W1973492092 cites W2037212449 @default.
• W1973492092 cites W2045788063 @default.
• W1973492092 cites W2054370312 @default.
• W1973492092 cites W2055350245 @default.
• W1973492092 cites W2063842247 @default.
• W1973492092 cites W2066167419 @default.
• W1973492092 cites W2072655346 @default.
• W1973492092 cites W2074580579 @default.
• W1973492092 cites W2078880075 @default.
• W1973492092 cites W2081433217 @default.
• W1973492092 cites W2085101416 @default.
• W1973492092 cites W2086879031 @default.
• W1973492092 cites W2090273203 @default.
• W1973492092 cites W2091287125 @default.
• W1973492092 cites W2091896722 @default.
• W1973492092 cites W2092833610 @default.
• W1973492092 cites W2101670640 @default.
• W1973492092 cites W2103546861 @default.
• W1973492092 cites W2108338619 @default.
• W1973492092 cites W2108978779 @default.
• W1973492092 cites W2109291482 @default.
• W1973492092 cites W2112050041 @default.
• W1973492092 cites W2112905569 @default.
• W1973492092 cites W2121607333 @default.
• W1973492092 cites W2130522675 @default.
• W1973492092 cites W2133142105 @default.
• W1973492092 cites W2137541332 @default.
• W1973492092 cites W2140436264 @default.
• W1973492092 cites W2141158635 @default.
• W1973492092 cites W2153317207 @default.
• W1973492092 cites W2156371987 @default.
• W1973492092 cites W2159154881 @default.
• W1973492092 cites W2159957100 @default.
• W1973492092 cites W2163990363 @default.
• W1973492092 cites W2408999259 @default.
• W1973492092 cites W91108846 @default.
• W1973492092 cites W4244773408 @default.
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