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Neurodegenerative disorders are frequently associated with β-sheet-rich amyloid deposits. Amyloid-forming proteins can aggregate under different structural conformations known as strains, which can exhibit a prion-like behaviour and distinct patho-phenotypes. Precise molecular determinants defining strain specificity and cross-strain interactions (cross-seeding) are currently unknown. The HET-s prion protein from the fungus Podospora anserina represents a model system to study the fundamental properties of prion amyloids. Here, we report the amyloid prion structure of HELLF, a distant homolog of the model prion HET-s. We find that these two amyloids, sharing only 17% sequence identity, have nearly identical β-solenoid folds but lack cross-seeding ability in vivo, indicating that prion specificity can differ in extremely similar amyloid folds. We engineer the HELLF sequence to explore the limits of the sequence-to-fold conservation and to pinpoint determinants of cross-seeding and prion specificity. We find that amyloid fold conservation occurs even at an exceedingly low level of identity to HET-s (5%). Next, we derive a HELLF-based sequence, termed HEC, able to breach the cross-seeding barrier in vivo between HELLF and HET-s, unveiling determinants controlling cross-seeding at residue level. These findings show that virtually identical amyloid backbone structures might not be sufficient for cross-seeding and that critical side-chain positions could determine the seeding specificity of an amyloid fold. Our work redefines the conceptual boundaries of prion strain and shed new light on key molecular features concerning an important class of pathogenic agents.

International audience; Due to the wealth of actors involved in the development of atherosclerosis, molecular imaging based on the targeting of specific markers would substantiate the diagnosis of life-threatening atheroma plaques. To this end, TEG4 antibody is a promising candidate targeting the activated platelets (integrin αIIbβ3) highly represented within the plaque. In this study, scFv antibody fragments were used to functionalize multimodal imaging nanoparticles. This grafting was performed in a regio-selective way to preserve TEG4 activity and the avidity of the nanoparticles was studied with respect to the number of grafted antibodies. Subsequently, taking advantage of the nanoparticle bimodality, both near infrared fluorescence and magnetic resonance imaging of the atheroma plaque were performed in the ApoE−/− mouse model. Here we describe the design of the targeted nanoparticles, and a quantification method for their detection in mice, both ex vivo and in vivo, highlighting their value as a potential diagnosis agent.

We present a new solid-state NMR proton-detected three-dimensional experiment dedicated to the observation of protein proton side chain resonances in nano-liter volumes. The experiment takes advantage of very fast magic angle spinning and double quantum 13C-13C transfer to establish efficient (H)CCH correlations detected on side chain protons. Our approach is demonstrated on the HET-s prion domain in its functional amyloid fibrillar form, fully protonated, with a sample amount of less than 500 g using a MAS frequency of 70kHz. The majority of aliphatic and aromatic side chain protons (70%) are observable, in addition to Halpha resonances, in a single experiment providing a complementary approach to the established proton-detected amide-based multidimensional solid-state NMR experiments for the study and resonance assignment of biosolid samples, in particular for aromatic side chain resonances.

The Samlat Formation is well exposed in coastal sections bordering the Atlantic Ocean south of Ad-Dakhla in southwestern Morocco. Here some 22 m of rhythmically-bedded, chert-rich, marine siltstones and marls are overlain by 1–1.5 m of vertebrate-bearing microconglomeratic sandstone, another 4–8 m of rhythmically-bedded siltstone and marl, and finally a second 3–6 m unit of vertebrate-bearing muddy sandstone. The microconglomeratic and muddy sandstones represent low sea stands in what is otherwise a deeper water sequence. Cetacean skeletons are rare but cetacean vertebrae are common in the lower sandstone (bed B1), where many show the effects of reworking. The cetaceans in bed B1represent a minimum of five species, from smallest to largest: cf. Saghacetus sp., cf. Stromerius sp., Dorudon atrox, cf. Dorudon sp., and Basilosaurus isis. Bed B1 yields rib fragments that may represent sirenians, but sirenians, if present, are rare. The only identifiable cetacean found in the upper sandstone (bed B2) is Basilosaurus sp. Dugongid sirenians identified as cf. Eosiren sp. are the most common mammal in bed B2. We interpret co-occurrence of the typically Early Priabonian species Dorudon atrox and Basilosaurus isis with smaller species more like Middle Priabonian genera Saghacetus osiris and Stromerius nidensis to indicate that bed B1 was deposited during low sea stand Pr-2 between the Early and Middle Priabonian (between the early and middle Late Eocene). Bed B2 is separated from B1 by an interval of deeper water sediment accumulation. Bed B2 could represent a later phase of Pr-2 or a subsequent Priabonian low sea stand (possibly Pr-3).

A vast majority of the cardioprotective properties exhibited by High-Density Lipoprotein (HDL) is mediated by its major protein component Apolipoprotein A-I (ApoA1). In order to develop a simplified bioprocess for producing recombinant human Apolipoprotein A-I (rhApoA1) in its near-native form, rhApoA1was expressed without the use of an affinity tag in view of its potential therapeutic applications. Expressed in Pichia pastoris at expression levels of 58.2 mg ApoA1 per litre of culture in a reproducible manner, the target protein was purified by mixed-mode chromatography using Capto™ MMC ligand with a purity and recovery of 84% and 68%, respectively. ApoA1 purification was scaled up to Mixed-mode Expanded Bed Adsorption chromatography to establish an 'on-line' process for the efficient capture of rhApoA1 directly from the P. pastoris expression broth. A polishing step using anion exchange chromatography enabled the recovery of ApoA1 up to 96% purity. Purified ApoA1 was identified and verified by RPLC-ESI-Q-TOF mass spectrometry. This two-step process would reduce processing times and therefore costs in comparison to the twelve-step procedure currently used for recovering rhApoA1 from P. pastoris.

Early Paleogene mammals are rare in Africa. They are mainly found in the northwestern part of the continent. Nearshore marine deposits from the Ouarzazate basin, in the southern rim of the Central High Atlas (Morocco), have yielded diverse Thanetian and Ypresian selachian and mammalian faunas [Cappetta et al., 1978, 1987; Gheerbrant et al., 1993]. These faunas from the Paleocene/Eocene transition document the oldest known placental mammals of Africa [Gheerbrant, 1992, 1994, 1995; Gheerbrant et al., 1998; Sigé et al., 1990; Sudre et al., 1993]. New field work in the central part of the basin, near El-Kelaâ M’Gouna, has led to the discovery of a new locality with continental and marine remains, including some non-determinable ostracods and charophytes plus planktonic and benthic foraminifera. The tests of the planktonic foraminifera were globally poorly preserved and highly re-crystallized. However, we found Globigerapsis index (dominant in the sample), Globigerapsis aff. kugleri, Subbotina frontosa, Subbotina inaequispira, Acarinina bullbrooki, Acarinina aff. pentacamerata, Morozovella aragonensis, Morozovella caucasica, Morozovella crater and Pseudohastigerina sp. Following Berggren et al. [1995], the lowest occurrence of the genus Globigerapsis denotes the P10/P11 zonal boundary and the highest occurrence of M. aragonensis denotes the P11/P12 zonal boundary. Both these taxa were represented by several relatively well-preserved specimens. Then, we consider that the sample is correlative to the planktonic foraminiferal Zone P11, for which a biochronal equivalent has been dated between 45.8 and 43.6 Ma. Consequently, the middle Lutetian Aznag locality represents the only known Eocene mammalian site to be adequately dated in Africa. The selachian fauna is relatively rich with nearly 30 species, many of them being probably new. The genera Squatiscyllium, Protoginglymostoma, Ouledia and Garabatis are confidently identified; these taxa were unknown in the Lutetian. We mention the first occurrence in the Eocene of Morocco of « septentrional » taxa Protoginglymostoma, Hemiscyllium and Rhinobatos bruxelliensis and we confirm the presence of genera Eomobula and Rhynchobatus. These data suggest significant North to South faunal exchanges, that contrast with the South to North tendency observed during the early Eocene [Noubhani and Cappetta, 1997]. Nevertheless, the selachian fauna from Aznag is clearly tropical with the occurrence of endemic taxa only known from Morocco (Garabatis, Orectolobiformes nov. gen.) and from the equatorial Tethys-Central Atlantic (Chiloscyllium aff. meraense, Ginglymostoma aff. angolense, Squatiscyllium, « Dasyatis » aff. sudrei, Ouledia, Arechia, Odontorhytis). Only 15 mammalian teeth have been recovered, most of them are very fragmentary and of small size; they document at least 7 species. A probable soricomorph “insectivore” documented by a complete lower molar is very atypical by its tiny size and the entoconid and hypoconulid poorly differentiated from the postcristid. Another “insectivore” is documented by a trigonid of p4, it is reminiscent to the zalambdodont groups. An incisor of rodent shows an enamel microstructure with uniserial Hunter-Schreger bands ; this structure is observed in several Eocene rodents from Eurasia and could be related to the pauciserial to uniserial transition observed in Zegdoumys sebtlai from Chambi (early Eocene, Tunisia) [Martin, 1999]. An upper molar of chiropteran cannot be assigned to one of the known microchiropteran superfamilies; the species from Aznag is characterized by derived traits (e.g. crestiform protocone, hypocone present) and primitive ones such as the presence of a paraconule which evokes the “eochiropteran” grade and the primitive vespertilionoids (paleochiropterygids). A small primate is documented by two fragmentary lower molars and characterized by a well-marked bunodonty and a large paraconid ; this form differs from contemporaneous anthropoids but is very similar to Altiatlasius from the Thanetian of the Adrar Mgorn 1. Two “condylarths” evoke European taxa; a M3 is tentatively assigned to the genus Paschatherium and a DP3or4 is reminiscent to that of Microhyus. This specimen shows also affinities with the ?DP4 of Chambius, a primitive macroscelidid from Chambi. The presence of these mammals strengthen the hypothesis of faunal exchanges between Africa and Europe during the early Eocene. During the middle Eocene, in the south of the Central High Atlas, palynologic data suggested a fairly dry, scarcely vegetated hinterland, and a tropical coastal vegetation (mangrove swamp and salt-marsh) [Fechner, 1988]. The mammals of Aznag, which are characteristic of a closed environment (chiropterans, primate, small ungulates), were certainly confined to a thin belt of coastal vegetation (forest gallery). The fossiliferous level is characterized by a cyclicity of the deposits which could suggest a seasonal variation. Among the selacians, the abundance of the batoids, the absence of both macrofauna and deep-water selacians, and the fragmentary elements indicate hydrodynamical transport in a shallow channel or delta.

International audience; Hepcidin was first identified as an antimicrobial peptide present in human serum and urine. It was later demonstrated that hepcidin is the long-sought hormone that regulates iron homeostasis in mammals. Recombinant human Hepcidin-25 (Hepc25) was expressed in Pichia pastoris using a modified version of the pPICZ alpha A vector. Hepc25 was then purified by a simple two-step chromatographic process to obtain 1.9 mg of soluble recombinant human Hepc25 per liter of culture at 96% purity. The sequence of Hepc25 and the presence of four disulfide bridges were confirmed by mass spectrometry analyses, and the recombinant Hepc25 exhibited antibacterial activity. This protocol of production and purification is the first step toward the production of human Hepc25 at a greater scale. (C) 2015 Elsevier B.V. All rights reserved.

In yeast, trehalose is synthesized by a multimeric enzymatic complex: TPS1 encodes trehalose 6-phosphate synthase, which belongs to a complex that is composed of at least three other subunits, including trehalose 6-phosphate phosphatase Tps2 and the redundant regulatory subunits Tps3 and Tsl1. The product of the TPS1 gene plays an essential role in the control of the glycolytic pathway by restricting the influx of glucose into glycolysis. In this paper, we investigated whether the trehalose synthesis pathway could be involved in the control of the other energy-generating pathway: oxidative phosphorylation. We show that the different mutants of the trehalose synthesis pathway (tps1Delta, tps2Delta, and tps1,2Delta) exhibit modulation in the amount of respiratory chains, in terms of cytochrome content and maximal respiratory activity. Furthermore, these variations in mitochondrial enzymatic content are positively linked to the intracellular concentration in cAMP that is modulated by Tps1p through hexokinase2. This is the first time that a pathway involved in sugar storage, i.e. trehalose, is shown to regulate the mitochondrial enzymatic content.

When the yeast protein Ypr140w was expressed in Escherichia coli, a lyso-PC [lysophosphatidylcholine (1-acylglycerophosphorylcholine)] acyltransferase activity was found associated with the membranes of the bacteria. To our knowledge, this is the first identification of a protein capable of catalysing the acylation of lyso-PC molecules to form PC. Fluorescence microscopy analysis of living yeasts revealed that the fusion protein Ypr140w–green fluorescent protein is targeted to the mitochondria. Moreover, in contrast with wild-type cells, in the absence of acyl-CoA, the yeast mutant deleted for the YPR140w gene has no lyso-PC acyltransferase activity associated with the mitochondrial fraction. When yeast cells were grown in the presence of lactate, the mutant synthesized 2-fold more triacylglycerols when compared with the wild-type. Moreover, its mitochondrial membranes contained a lesser amount of PC and cardiolipin, and the fatty acid composition of these latter was greatly changed. These modifications were accompanied by a 2-fold increase in the respiration rates (states 3 and 4) of the mitochondria. The relationship between the deletion of the YPR140w gene and the lipid composition of the ypr140wΔ cells is discussed.