Crystal structures of the Gon7/Pcc1 and Bud32/Cgi121 complexes provide a model for the complete yeast KEOPS complex.

International audienceThe yeast KEOPS protein complex comprising Kae1, Bud32, Cgi121, Pcc1 and Gon7 is responsible for the essential tRNA threonylcarbamoyladenosine (t(6)A) modification. Deletion of genes coding for the KEOPS subunits also affects telomere elongation and transcriptional regulation. In the present work, the crystal structure of Bud32/Cgi121 in complex with ADP revealed that ADP is bound in the catalytic site of Bud32 in a canonical manner characteristic of Protein Kinase A (PKA) family proteins. We found that Gon7 forms a stable heterodimer with Pcc1 and report the crystal structure of the Pcc1-Gon7 heterodimer. Gon7 interacts with the same Pcc1 region engaged in the archaeal Pcc1 homodimer. We further show that yeast KEOPS, unlike its archaeal counterpart, exists as a heteropentamer in which Gon7, Pcc1, Kae1, Bud32 and Cgi121 also adopt a linear arrangement. We constructed a model of yeast KEOPS that provides structural insight into the role of Gon7. The model also revealed the presence of a highly positively charged crater surrounding the entrance of Kae1 that likely binds tRNA

Entre l’antique et l’exotique, le projet comparatiste oublié du « Muséum des Antiques » en l’an III

L’éphémère « Muséum des Antiques » (an III-1795) a rassemblé, sous la Révolution, des objets à caractère ethnographique à partir des saisies révolutionnaires tant en France qu’en Europe, et en absorbant les collections du Muséum d’histoire naturelle. En exposant les Antiques du Cabinet des médailles aux côtés de l’exotique, la démarche se voulait délibérément comparative. L’article se propose de dresser la chronologie de ce musée méconnu de la Convention et de dégager le projet comparatiste entre l’antique et l’exotique.The short-lived « Muséum des Antiques » (an III - 1795) under the Revolution collected objects of an ethnographic nature from revolutionary confiscations in France and in Europe, assimilating them to the collections of the Museum of Natural History. By displaying ancient objects from the Cabinet des Medailles next to such exotica, the goal was deliberately comparative. This article aims to establish a chronology of this unknown museum of the National Convention and to identity the comparative program between the Ancient and the Exotic

Stratospheric aerosols from the Sarychev volcano eruption in the 2009 Arctic summer

Aerosols from the Sarychev volcano eruption (Kuril Islands, northeast of Japan) were observed in the Arctic lower stratosphere a few days after the strongest SO2 injection which occurred on 15 and 16 June 2009. From the observations provided by the Infrared Atmospheric Sounding Interferometer (IASI) an estimated 0.9 Tg of sulphur dioxide was injected into the upper troposphere and lower stratosphere (UTLS). The resultant stratospheric sulphate aerosols were detected from satellites by the Optical Spectrograph and Infrared Imaging System (OSIRIS) limb sounder and by the Cloud-Aerosol Lidar with Orthogonal Polarization (CALIOP) and from the surface by the Network for the Detection of Atmospheric Composition Changes (NDACC) lidar deployed at OHP (Observatoire de Haute-Provence, France). By the first week of July the aerosol plume had spread out over the entire Arctic region. The Sarychev-induced stratospheric aerosol over the Kiruna region (north of Sweden) was measured by the Stratospheric and Tropospheric Aerosol Counter (STAC) during eight balloon flights planned in August and September 2009. During this balloon campaign the Micro Radiomètre Ballon (MicroRADIBAL) and the Spectroscopie d'Absorption Lunaire pour l'Observation des Minoritaires Ozone et NOx (SALOMON) remote-sensing instruments also observed these aerosols. Aerosol concentrations returned to near-background levels by spring 2010. The effective radius, the surface area density (SAD), the aerosol extinction, and the total sulphur mass from STAC in situ measurements are enhanced with mean values in the range 0.15-0.21 μm, 5.5-14.7 μm2 cm-3, 5.5-29.5 × 10-4 km-1, and 4.9-12.6 × 10-10 kg[S] kg-1[air], respectively, between 14 km and 18 km. The observed and modelled e-folding time of sulphate aerosols from the Sarychev eruption is around 70-80 days, a value much shorter than the 12-14 months calculated for aerosols from the 1991 eruption of Mt Pinatubo. The OSIRIS stratospheric aerosol optical depth (AOD) at 750 nm is enhanced by a factor of 6, with a value of 0.02 in late July compared to 0.0035 before the eruption. The HadGEM2 and MIMOSA model outputs indicate that aerosol layers in polar region up to 14-15 km are largely modulated by stratosphere-troposphere exchange processes. The spatial extent of the Sarychev plume is well represented in the HadGEM2 model with lower altitudes of the plume being controlled by upper tropospheric troughs which displace the plume downward and upper altitudes around 18-20 km, in agreement with lidar observations. Good consistency is found between the HadGEM2 sulphur mass density and the value inferred from the STAC observations, with a maximum located about 1 km above the tropopause ranging from 1 to 2 × 10 -9 kg[S] kg-1[air], which is one order of magnitude higher than the background level. © Author(s) 2013.The authors thank the CNES balloon launching team for successful operations and the Swedish Space Corporation at Esrange. The ETHER database (CNES-INSUCNRS) and the CNES “sous-direction Ballon” are partners of the project. The StraPolEt ´ e project has been funded by the French ´ “Agence Nationale de la Recherche” (ANR-BLAN08-1-31627), the “Centre National d’Etudes Spatiales” (CNES), and the “Institut ´ Polaire Paul-Emile Victor” (IPEV). The AEROWAVE (Aerosols, Water Vapor and Electricity) and the HALOHA (HALOgen in High Altitudes) projects have been funded by the recently created French CNES-INSU Balloon Committee (so-called CSTB). We are grateful to Slimane Bekki and David Cugniet for their constructive comments about the AER-UPMC 2-D model, to Marc-Antoine Drouin for his help about the MIMOSA model, and to the LPC2E technical team for this successful campaign. Jim Haywood and Andy Jones were supported by the Joint DECC/Defra Met Office Hadley Centre Climate Programme (GA01101). IASI was developed and built under the responsibility of the Centre National d’Etudes Spatiales (CNES, France). It is flown on board the Metop ´ satellites as part of the EUMETSAT Polar System. The IASI L1 data are received through the EUMETCast near-real-time data distribution service. L. Clarisse is a postdoctoral researcher with FRS-FNRS. We acknowledge the CALIOP team for acquiring and processing data as well as the ICARE team for providing and maintaining the computational facilities to store them. Odin is a Swedish-led satellite project funded jointly by Sweden (SNSB), Canada (CSA), France (CNES), and Finland (Tekes). This study was supported by the French VOLTAIRE Labex (Laboratoire d’Excellence ANR-10-LABX-100-01) managed by the University of Orleans

The highly conserved eukaryotic DRG factors are required for efficient translation in a manner redundant with the putative RNA helicase Slh1

Eukaryotic and archaeal DRG factors are highly conserved proteins with characteristic GTPase motifs. This suggests their implication in a central biological process, which has so far escaped detection. We show here that the two Saccharomyces cerevisiae DRGs form distinct complexes, RBG1 and RBG2, and that the former co-fractionate with translating ribosomes. A genetic screen for triple synthetic interaction demonstrates that yeast DRGs have redundant function with Slh1, a putative RNA helicase also associating with translating ribosomes. Translation and cell growth are severely impaired in a triple mutant lacking both yeast DRGs and Slh1, but not in double mutants. This new genetic assay allowed us to characterize the roles of conserved motifs present in these proteins for efficient translation and/or association with ribosomes. Altogether, our results demonstrate for the first time a direct role of the highly conserved DRG factors in translation and indicate that this function is redundantly shared by three factors. Furthermore, our data suggest that important cellular processes are highly buffered against external perturbation and, consequently, that redundantly acting factors may escape detection in current high-throughput binary genetic interaction screens

The enzyme activities of Caf1 and Ccr4 are both required for deadenylation by the human Ccr4-Not nuclease module

In eukaryotic cells, the shortening and removal of the poly(A) tail (deadenylation) of cytoplasmic mRNA is a key event in regulated mRNA degradation. A major enzyme involved in deadenylation is the Ccr4-Not deadenylase complex, which can be recruited to its target mRNA by RNA-binding proteins or the miRNA repression complex. In addition to six non-catalytic components, the complex contains two enzymatic subunits with ribonuclease activity: Ccr4 and Caf1 (Pop2). In vertebrates, each deadenylase subunit is encoded by two paralogues: Caf1, which can interact with the anti-proliferative protein BTG2, is encoded by CNOT7 and CNOT8, while Ccr4 is encoded by the highly similar genes CNOT6 and CNOT6L. Currently, it is unclear whether the catalytic subunits work cooperatively, or whether the nuclease components have unique roles in deadenylation. We therefore developed a method to express and purify a minimal human BTG2-Caf1-Ccr4 nuclease sub-complex from bacterial cells. By using chemical inhibition and well-characterised inactivating amino acid substitutions, we demonstrate that the enzyme activities of Caf1 and Ccr4 are both required for deadenylation in vitro. These results indicate that Caf1 and Ccr4 cooperate in mRNA deadenylation and suggest that the enzyme activities of Caf1 and Ccr4 are regulated via allosteric interactions within the nuclease module

Diversité, structure et endémicité des communautés de vers de terre et de collemboles dans une hêtraie peu aménagée des Pyrénées (France)

We assessed and compared patterns of biodiversity and the composition of two soil invertebrate communities (Collembola and Lumbricidae) in three slightly managed plots in a beech forest of the French Pyrenees. The plots were managed in three different ways: an even-aged (REG), a closed unevenaged stand (NAT ), and an open uneven-aged full-grown stand (IRR ). At each sampling point, earthworms and Collembola from litter, soil, and pitfall traps were collected, and nine edaphic and environmental parameters were measured. The fauna collected was rich in species, and endemic and rare taxa. No clear-cut differences in species richness appeared between plots. Nevertheless, (1) the less disturbed plot, i.e. NAT , hosted a slightly larger number of endemic and rare species than IRR and REG; (2) the structure of both communities in NAT, REG and IRR differed significantly depending on the site elevation and organic nitrogen content (higher in NAT ), soil surface temperature and soil pH level (higher in IRR ), and humus index and soil water content (higher in REG ). Those factors, except elevation, may be explained by the canopy opening partly controlled by management practices; (3) a side aspect of the study was to show that sampling exhaustiveness required more sampling effort for soil than for litter, and more for Lumbricidae than for Collembola. In a broader context, the case documented in this study suggests that management practices with limited clearing of forest cover affect soil biodiversity only slightly, which is in sharp contrast with the collapse in endemic biodiversity induced by reafforestationNous avons évalué et comparé les patrons de biodiversité et la composition de deux communautés d'invertébrés du sol (collemboles et lombrics) dans trois parcelles peu aménagées d'une hêtraie-sapinière dans les Pyrénées françaises. Ces parcelles sont gérées selon trois modalités différentes: une futaie régulière (REG), une futaie irrégulière fermée (NAT) et une futaie irrégulière ouverte (IRR). À chaque point d'échantillonnage les vers de terre et les collemboles ont été collectés dans la litière, le sol et au moyen de pièges Barber, et neuf paramètres édaphiques et environnementaux ont été mesurés. La faune récoltée était riche en espèces ainsi qu'en taxons rares et endémiques. Aucune différence marquée de la richesse spécifique n'est apparue entre les parcelles. Néanmoins (1) le site le moins perturbé, i.e. NAT, hébergeait un nombre légèrement plus élevé d'espèces endémiques et rares que IRR et REG; (2) la structure des deux communautés différait significativement entre NAT, REG et IRR selon l'altitude du site et la teneur en azote organique (plus forte dans NAT), la température à la surface du sol et le pH du sol (plus élevés dans IRR), ainsi que l'indice d'humus et la teneur en eau du sol (plus élevés en REG). Ces facteurs, à l'exception de l'altitude, peuvent s'expliquer par l'ouverture de la canopée partiellement contrôlée par les pratiques de gestion; (3) un à-côté de cette étude est d'avoir montré que l'exhaustivité de l'échantillonnage requiert plus d'efforts pour le sol que pour la litière, et plus pour les lombrics que pour les collemboles. Dans un contexte plus large, le cas traité dans cette étude suggère que des pratiques de gestion avec un éclaircissement limité du couvert forestier n'affectent que légèrement la biodiversité du sol, ce qui contraste fortement avec l'effondrement de la biodiversité endémique qu'induit la reforestation

60S ribosomal subunit assembly dynamics defined by semi-quantitative mass spectrometry of purified complexes

During the highly conserved process of eukaryotic ribosome formation, RNA follows a maturation path with well-defined, successive intermediates that dynamically associate with many pre-ribosomal proteins. A comprehensive description of the assembly process is still lacking. To obtain data on the timing and order of association of the different pre-ribosomal factors, a strategy consists in the use of pre-ribsomal particles isolated from mutants that block ribosome formation at different steps. Immunoblots, inherently limited to only a few factors, have been applied to evaluate the accumulation or decrease of pre-ribosomal intermediates under mutant conditions. For a global protein-level description of different 60S ribosomal subunit maturation intermediates in yeast, we have adapted a method of in vivo isotopic labelling and mass spectrometry to study pre-60S complexes isolated from strains in which rRNA processing was affected by individual depletion of five factors: Ebp2, Nog1, Nsa2, Nog2 or Pop3. We obtained quantitative data for 45 distinct pre-60S proteins and detected coordinated changes for over 30 pre-60S factors in the analysed mutants. These results led to the characterisation of the composition of early, intermediate and late pre-ribosomal complexes, specific for crucial maturation steps during 60S assembly in eukaryotes

Gcn4 misregulation reveals a direct role for the evolutionary conserved EKC/KEOPS in the t6A modification of tRNAs

The EKC/KEOPS complex is universally conserved in Archaea and Eukarya and has been implicated in several cellular processes, including transcription, telomere homeostasis and genomic instability. However, the molecular function of the complex has remained elusive so far. We analyzed the transcriptome of EKC/KEOPS mutants and observed a specific profile that is highly enriched in targets of the Gcn4p transcriptional activator. GCN4 expression was found to be activated at the translational level in mutants via the defective recognition of the inhibitory upstream ORFs (uORFs) present in its leader. We show that EKC/KEOPS mutants are defective for the N6-threonylcarbamoyl adenosine modification at position 37 (t6A37) of tRNAs decoding ANN codons, which affects initiation at the inhibitory uORFs and provokes Gcn4 de-repression. Structural modeling reveals similarities between Kae1 and bacterial enzymes involved in carbamoylation reactions analogous to t6A37 formation, supporting a direct role for the EKC in tRNA modification. These findings are further supported by strong genetic interactions of EKC mutants with a translation initiation factor and with threonine biosynthesis genes. Overall, our data provide a novel twist to understanding the primary function of the EKC/KEOPS and its impact on several essential cellular functions like transcription and telomere homeostasis