Biochemical requirements for the maturation of mitochondrial c-type cytochromes

AbstractCytochromes c are metalloproteins that function in electron transfer reactions and contain a heme moiety covalently attached via thioether linkages between the co-factor and a CXXCH motif in the protein. Covalent attachment of the heme group occurs on the positive side of all energy-transducing membranes (bacterial periplasm, mitochondrial intermembrane space and thylakoid lumen) and requires minimally: 1) synthesis and translocation of the apocytochromes c and heme across at least one biological membrane, 2) reduction of apocytochromes c and heme and maintenance under a reduced form prior to 3) catalysis of the heme attachment reaction. Surprisingly, the conversion of apoforms of cytochromes c to their respective holoforms occurs through at least three different pathways (systems I, II and III). In this review, we detail the assembly process of soluble cytochrome c and membrane-bound cytochrome c1, the only two mitochondrial c-type cytochromes that function in respiration. Mitochondrial c-type cytochromes are matured in the intermembrane space via the system I or system III pathway, an intriguing finding considering that the biochemical requirements for cytochrome c maturation are believed to be common regardless of the energy-transducing membrane under study

Absorption foliaire des métaux présents dans des particules atmosphériques issues d'une usine de recyclage de batteries : biotest laitue

National audienceLes flux de polluants émis dans l'environnement ont été considérablement réduits en particulier par la mise en place par les industriels de système de filtres performants. Cependant les particules très fines et particulièrement réactives sont toujours émises dans l'environnement. De nombreuses études décrivent le transfert sol-plante des métaux mais très peu concernent la voie de transfert atmosphère plante. Pourtant, selon le rapport parlementaire de Miquel (2001), l'enrichissement actuel des sols en plomb provient pour 68% des retombées atmosphériques qui sont aussi interceptées par les plantes. Le transfert foliaire direct via des aérosols particulaires a été démontré pour des radionucléides (137Cs, 85Sr, 133Ba et 123mTe) par Madoz-Escande et al. (2004). Or les voies de transport des radionucléides et métaux sont aussi celles des "oligoélements" (Zn, Co, Mo, Cu) dans les plantes. C'est pourquoi il paraît pertinent de s'intéresser au transfert foliaire des métaux. De nombreuses questions scientifiques se posent en effet concernant le transfert foliaire des métaux. Est-il possible? Si oui sous quelle forme sont les métaux? Quels sont les mécanismes physico chimiques et biologiques impliqués? Quelle est l'importance de cette voie vis-à-vis du transfert sol plante ? Pour répondre à ces questions, le transfert du plomb et du cadmium vers les parties aériennes des plantes via le dépôt atmosphérique de particules industrielles riches en métaux a été expérimenté et modélisé

Influence of metal process micronic and submicronic particles on vegetables quality and ecosystems

International audienceImpact of atmospheric process particles enriched with metals (PM) on various vegetables was studied. Foliar metal interception was measured and calculated. Soil-plant transfer and phyto-toxicity were also studied. Influence of species and washing procedure on metal burning was observed. High correlation was obtained between measured and simulated lead plant uptake values. Ageing effect in polluted soils was highlighted with stabilisation or mobilization of metals in function of contact duration between soils and PM

Mortality from gastrointestinal congenital anomalies at 264 hospitals in 74 low-income, middle-income, and high-income countries: a multicentre, international, prospective cohort study

Summary Background Congenital anomalies are the fifth leading cause of mortality in children younger than 5 years globally. Many gastrointestinal congenital anomalies are fatal without timely access to neonatal surgical care, but few studies have been done on these conditions in low-income and middle-income countries (LMICs). We compared outcomes of the seven most common gastrointestinal congenital anomalies in low-income, middle-income, and high-income countries globally, and identified factors associated with mortality. Methods We did a multicentre, international prospective cohort study of patients younger than 16 years, presenting to hospital for the first time with oesophageal atresia, congenital diaphragmatic hernia, intestinal atresia, gastroschisis, exomphalos, anorectal malformation, and Hirschsprung’s disease. Recruitment was of consecutive patients for a minimum of 1 month between October, 2018, and April, 2019. We collected data on patient demographics, clinical status, interventions, and outcomes using the REDCap platform. Patients were followed up for 30 days after primary intervention, or 30 days after admission if they did not receive an intervention. The primary outcome was all-cause, in-hospital mortality for all conditions combined and each condition individually, stratified by country income status. We did a complete case analysis. Findings We included 3849 patients with 3975 study conditions (560 with oesophageal atresia, 448 with congenital diaphragmatic hernia, 681 with intestinal atresia, 453 with gastroschisis, 325 with exomphalos, 991 with anorectal malformation, and 517 with Hirschsprung’s disease) from 264 hospitals (89 in high-income countries, 166 in middleincome countries, and nine in low-income countries) in 74 countries. Of the 3849 patients, 2231 (58·0%) were male. Median gestational age at birth was 38 weeks (IQR 36–39) and median bodyweight at presentation was 2·8 kg (2·3–3·3). Mortality among all patients was 37 (39·8%) of 93 in low-income countries, 583 (20·4%) of 2860 in middle-income countries, and 50 (5·6%) of 896 in high-income countries (p<0·0001 between all country income groups). Gastroschisis had the greatest difference in mortality between country income strata (nine [90·0%] of ten in lowincome countries, 97 [31·9%] of 304 in middle-income countries, and two [1·4%] of 139 in high-income countries; p≤0·0001 between all country income groups). Factors significantly associated with higher mortality for all patients combined included country income status (low-income vs high-income countries, risk ratio 2·78 [95% CI 1·88–4·11], p<0·0001; middle-income vs high-income countries, 2·11 [1·59–2·79], p<0·0001), sepsis at presentation (1·20 [1·04–1·40], p=0·016), higher American Society of Anesthesiologists (ASA) score at primary intervention (ASA 4–5 vs ASA 1–2, 1·82 [1·40–2·35], p<0·0001; ASA 3 vs ASA 1–2, 1·58, [1·30–1·92], p<0·0001]), surgical safety checklist not used (1·39 [1·02–1·90], p=0·035), and ventilation or parenteral nutrition unavailable when needed (ventilation 1·96, [1·41–2·71], p=0·0001; parenteral nutrition 1·35, [1·05–1·74], p=0·018). Administration of parenteral nutrition (0·61, [0·47–0·79], p=0·0002) and use of a peripherally inserted central catheter (0·65 [0·50–0·86], p=0·0024) or percutaneous central line (0·69 [0·48–1·00], p=0·049) were associated with lower mortality. Interpretation Unacceptable differences in mortality exist for gastrointestinal congenital anomalies between lowincome, middle-income, and high-income countries. Improving access to quality neonatal surgical care in LMICs will be vital to achieve Sustainable Development Goal 3.2 of ending preventable deaths in neonates and children younger than 5 years by 2030

Etude d'AtCCME, une protéine mitochondriale liant l'hème au cours de la maturation des cytochromes de type c

STRASBOURG-Sc. et Techniques (674822102) / SudocSudocFranceF

La voie de réduction dans la maturation des cytochromes de type c mitochondriaux (Etude de la protéine CCMH d'Arabidopsis thaliana)

La principale fonction des mitochondries est la synthèse d'énergie pour la cellule eucaryote sous forme d'ATP. Cette synthèse se fait au niveau de la membrane interne mitochondriale grâce à l'ATP synthase qui utilise le gradient de protons créé par la chaîne respiratoire, un ensemble de complexes multiprotéiques. Cette chaîne respiratoire mitochondriale comporte de nombreux transporteurs d'électrons contenant différents types de cofacteurs. Parmi ces transporteurs, les cytochromes de type c possèdent un cofacteur hémique qui est attaché de façon covalente sur les deux cystéines du motif CXXCH de l'apocytochrome. Dans les mitochondries, deux cytochromes de type c sont présents, le cytochrome c1 qui fait partie du complexe III et le cytochrome c qui transfère les électrons du complexe III au complexe IV. Ces deux protéines sont respectivement orientée ou localisée dans la membrane interne vers l'espace inter-membranaire.Les cytochromes de type c sont présents dans tous les organismes des procaryotes aux eucaryotes. Différentes étapes sont nécessaires à la biogenèse de ces hémoprotéines :le transport des apocytochromes et de l'hème vers le périplasme, l'espace intermembranaire ou le lumen selon les cas,le maintien sous forme réduite des groupements sulfhydrils des apocytochromes et du fer de l'hème,l'établissement d'une liaison covalente entre l'hème et l'apocytochrome.Trois voies de maturation des cytochromes de type c ont été mises en place au cours de l'évolution, l'une est présente chez certaines bactéries à Gram négatif (système I), l'autre chez les chloroplastes et les bactéries à Gram positif (système II), la troisième dans les mitochondries de levure et d'autres eucaryotes animaux (système III) (1). Dans les mitochondries de levure, l'assemblage de l'hème sur chaque apocytochrome est réalisé par une seule protéine, la cytochrome c hème lyase' pour le cytochrome c et la cytochrome c1 hème lyase' pour le cytochrome c1. Aucun équivalent de ces hème lyases n'a été identifié dans les mitochondries de plantes. Par contre, chez les plantes terrestres, le génome mitochondrial code pour quatre ou cinq protéines présentant de fortes similarités avec certaines protéines Ccm (pour "cytochrome c maturation") bactériennes (3-7). Chez Escherichia coli, organisme modèle pour le système I, 8 gènes ccm, organisés en opéron, sont impliqués dans les différentes étapes de la maturation des cytochromes de type c. Les gènes ccm codent pour un ABC-transporteur, une voie de livraison de l'hème contenant une protéine chaperon d'hème, une hème lyase potentielle et une voie de réduction composée d'une thioredoxine (CcmG) et d'une thiol-disulfide oxydoréductase (CcmH) qui transfèrent les électrons à l'apocytochrome (2).Le génome mitochondrial d'Arabidopsis thaliana ne codant que pour 3 des protéines Ccm orthologues d'E. coli, d'autres gènes ont été recherchés dans le génome nucléaire d'A. thaliana. Ainsi, au laboratoire, deux nouvelles protéines CCM d'A. thaliana, codées par le génome nucléaire puis importées dans les mitochondries, ont été identifiées et étudiées. Il s'agit d'AtCCME, un chaperon de l'hème impliqué dans la voie de livraison de l'hème (8) et d'AtCCMA, le domaine de fixation de l'ATP de l'ABC-transporteur (9).Le travail réalisé au cours de cette thèse a eu pour but d'isoler et de caractériser la protéine homologue à CcmH dans la voie de maturation des cytochromes c qui est spécifique aux mitochondries végétales. La protéine bactérienne CcmH est une protéine périplasmique indispensable à la formation d'holocytochrome c. Cette protéine serait directement impliquée dans la réduction des groupements thiols des cystéines de l'apocytochromes (10). CcmH est une protéine bipartite chez E. coli, la partie N-terminale d'EcCcmH est homologue à la protéine CycL de Bradyrhizobium japonicum et possède le site actif contenant le motif RCXXC. La partie C-terminale d'EcCcmH n'est pas indispensable pour la formation d'holocytochromes. [...]The main function of mitochondria is the synthesis of energy for the whole cell. This is done by the ATP synthase that utilises the proton gradient resulting from the activity of the respiratory chain. This respiratory chain is composed of multiprotein complexes that contain several electron transporters. c-type cytochromes are electron carriers that possess a heme as cofactor. They are characterised by the covalent attachment of this heme to two cysteinyl residues of the apocytochrome via two thioether bonds. In mitochondria, there are two c-type cytochromes : cytochrome c1 a component of complex III and cytochrome c that transfers electrons from complex III to complex IV. In these two proteins, the cofactors are respectively facing the inter-membrane space or present in the inter-membrane space.c-type cytochromes are present in all organisms from prokaryotes to eukaryotes. The biogenesis of these hemoproteins requires different steps :the transport of both apycytochrome and heme to the periplasm, inter-membrane space or lumen,the reduction of heme and cysteines of apocytochromes,the covalent ligation of heme to apocytochromes.Three distinct systems have evolved to achieve the conversion of apocytochromes to their holoforms (1). One is found in some Gram negative bacteria (system I), the second in chloroplast and Gram positive bacteria (System II) and the last in yeast and animals mitochondria (System III). In yeast mitochondria, a single protein, cytochrome c heme lyase is required for the assembly. No equivalent of heme lyase was found in plant mitochondria. Plant mitochondrial genomes encode four or five proteins similar to bacterial Ccm (for "cytochrome c maturation") proteins (3-7). In Escherichia coli, 8 ccm genes, organised in one operon, encode the proteins needed for the maturation of c-type cytochromes. This system involves an ABC-transporter, a heme delivery pathway, a putative heme lyase and a reducing pathway composed by CcmG, a thioredoxin and CcmH a thiol-disulfide oxidoreductase that transfers electrons to the apocytochrome (2).The mitochondrial genome of Arabidopsis thaliana encodes 3 orthologues of E. coli Ccm proteins. Other Ccm genes were searched on the nuclear genome. Two were identified in the laboratory and found to be imported into mitochondria. These are AtCCME, a heme chaperone involved in the heme delivery pathway (8) and AtCCMA, the ATP binding subunit of the ABC transporter (9).The aim of this thesis was to identify and study the plant counterpart of ccmH, a component of the reducing pathway. In bacteria, CcmH is a periplasmic thiol:disulfide oxidoreductase involved in the reduction of two cysteines of the heme-binding motif of apocytochromes (10). CcmH is a bipartite protein in E. coli, the N-terminal part of EcCcmH is homologuous to the Bradyrhizobium japonicum CycL protein and possesses as active site for redox activity the RCXXC motif. The C-terminal part of EcCcmH is not necessary for the formation of holocytochromes.[...]STRASBOURG-Sc. et Techniques (674822102) / SudocSudocFranceF

Transfer RNA maturation in Chlamydomonas mitochondria, chloroplast and the nucleus by a single RNase P protein.

The maturation of tRNA precursors involves the 5' cleavage of leader sequences by an essential endonuclease called RNase P. Beyond the ancestral ribonucleoprotein (RNP) RNase P, a second type of RNase P called PRORP (protein-only RNase P) evolved in eukaryotes. The current view on the distribution of RNase P in cells is that multiple RNPs, multiple PRORPs or a combination of both, perform specialised RNase P activities in the different compartments where gene expression occurs. Here, we identify a single gene encoding PRORP in the green alga Chlamydomonas reinhardtii while no RNP is found. We show that its product, CrPRORP, is triple-localised to mitochondria, the chloroplast and the nucleus. Its downregulation results in impaired tRNA biogenesis in both organelles and the nucleus. CrPRORP, as a single-subunit RNase P for an entire organism, makes up the most compact and versatile RNase P machinery described in either prokaryotes or eukaryotes

Resemblance and Dissemblance of Arabidopsis Type II Peroxiredoxins: Similar Sequences for Divergent Gene Expression, Protein Localization, and Activity

The Arabidopsis type II peroxiredoxin (PRXII) family is composed of six different genes, five of which are expressed. On the basis of the nucleotide and protein sequences, we were able to define three subgroups among the PRXII family. The first subgroup is composed of AtPRXII-B, -C, and -D, which are highly similar and localized in the cytosol. AtPRXII-B is ubiquitously expressed. More striking is the specific expression of AtPRXII-C and AtPRXII-D localized in pollen. The second subgroup comprises the mitochondrial AtPRXII-F, the corresponding gene of which is expressed constitutively. We show that AtPRXII-E, belonging to the last subgroup, is expressed mostly in reproductive tissues and that its product is addressed to the plastid. By in vitro enzymatic experiments, we demonstrate that glutaredoxin is the electron donor of recombinant AtPRXII-B for peroxidase reaction, but the donors of AtPRXII-E and AtPRXII-F have still to be identified

Inactivation of Thioredoxin Reductases Reveals a Complex Interplay between Thioredoxin and Glutathione Pathways in Arabidopsis Development[W]

NADPH-dependent thioredoxin reductases (NTRs) are key regulatory enzymes determining the redox state of the thioredoxin system. The Arabidopsis thaliana genome has two genes coding for NTRs (NTRA and NTRB), both of which encode mitochondrial and cytosolic isoforms. Surprisingly, plants of the ntra ntrb knockout mutant are viable and fertile, although with a wrinkled seed phenotype, slower plant growth, and pollen with reduced fitness. Thus, in contrast with mammals, our data demonstrate that neither cytosolic nor mitochondrial NTRs are essential in plants. Nevertheless, in the double mutant, the cytosolic thioredoxin h3 is only partially oxidized, suggesting an alternative mechanism for thioredoxin reduction. Plant growth in ntra ntrb plants is hypersensitive to buthionine sulfoximine (BSO), a specific inhibitor of glutathione biosynthesis, and thioredoxin h3 is totally oxidized under this treatment. Interestingly, this BSO-mediated growth arrest is fully reversible, suggesting that BSO induces a growth arrest signal but not a toxic accumulation of activated oxygen species. Moreover, crossing ntra ntrb with rootmeristemless1, a mutant blocked in root growth due to strongly reduced glutathione synthesis, led to complete inhibition of both shoot and root growth, indicating that either the NTR or the glutathione pathway is required for postembryonic activity in the apical meristem