Use of resonance Raman spectroscopy to study the phase diagram of PbZr0.52Ti0.48O3

Evidence is presented for the first time that the sharp and continuous spectral changes observed in PbZr0.52Ti0.48O3 (PZT) between 350 and 10 K with the 647.1 nm wavelength are due to a resonance Raman effect. Such a phenomenon can be explained by means of a self-trapped exciton emission oxygen deficient complex (TiTi' - VO-) of PZT powder whose energy is close to the radiation line of the laser. This kind of approach should also be very useful to distinguish the phase transition sequence for other related ferro/ piezoelectric systems

From Safety Analysis to Experimental Validation by Fault Injection—Case of Automotive Embedded Systems

En raison de la complexité croissante des systèmes automobiles embarqués, la sûreté de fonctionnement est devenue un enjeu majeur de l’industrie automobile. Cet intérêt croissant s’est traduit par la sortie en 2011 de la norme ISO 26262 sur la sécurité fonctionnelle. Les défis auxquelles sont confrontés les acteurs du domaine sont donc les suivants : d’une part, la conception de systèmes sûrs, et d’autre part, la conformité aux exigences de la norme ISO 26262. Notre approche se base sur l’application systématique de l’injection de fautes pour la vérification et la validation des exigences de sécurité, tout au long du cycle de développement, des phases de conception jusqu’à l’implémentation. L’injection de fautes nous permet en particulier de vérifier que les mécanismes de tolérance aux fautes sont efficaces et que les exigences non-fonctionnelles sont respectées. L’injection de faute est une technique de vérification très ancienne. Cependant, son rôle lors de la phase de conception et ses complémentarités avec la validation expérimentale, méritent d’être étudiés. Notre approche s’appuie sur l’application du modèle FARM (Fautes, Activations, Relevés et Mesures) tout au long du processus de développement. Les analyses de sûreté sont le point de départ de notre approche, avec l'identification des mécanismes de tolérance aux fautes et des exigences non-fonctionnelles, et se terminent par la validation de ces mécanismes par les expériences classiques d'injection de fautes. Enfin, nous montrons que notre approche peut être intégrée dans le processus de développement des systèmes embarqués automobiles décrits dans la norme ISO 26262. Les contributions de la thèse sont illustrées sur l’étude de cas d’un système d’éclairage avant d’une automobile. ABSTRACT : Due to the rising complexity of automotive Electric/Electronic embedded systems, Functional Safety becomes a main issue in the automotive industry. This issue has been formalized by the introduction of the ISO 26262 standard for functional safety in 2011. The challenges are, on the one hand to design safe systems based on a systematic verification and validation approach, and on the other hand, the fulfilment of the requirements of the ISO 26262 standard. Following ISO 26262 recommendations, our approach, based on fault injection, aims at verifying fault tolerance mechanisms and non-functional requirements at all steps of the development cycle, from early design phases down to implementation. Fault injection is a verification technique that has been investigated for a long time. However, the role of fault injection during design phase and its complementarities with the experimental validation of the target have not been explored. In this work, we investigate a fault injection continuum, from system design validation to experiments on implemented targets. The proposed approach considers the safety analyses as a starting point, with the identification of safety mechanisms and safety requirements, and goes down to the validation of the implementation of safety mechanisms through fault injection experiments. The whole approach is based on a key fault injection framework, called FARM (Fault, Activation, Readouts and Measures). We show that this approach can be integrated in the development process of the automotive embedded systems described in the ISO 26262 standard. Our approach is illustrated on an automotive case study: a Front-Light system

The Conserved nhaAR Operon Is Drastically Divergent between B2 and Non-B2 Escherichia coli and Is Involved in Extra-Intestinal Virulence

The Escherichia coli species is divided in phylogenetic groups that differ in their virulence and commensal distribution. Strains belonging to the B2 group are involved in extra-intestinal pathologies but also appear to be more prevalent as commensals among human occidental populations. To investigate the genetic specificities of B2 sub-group, we used 128 sequenced genomes and identified genes of the core genome that showed marked difference between B2 and non-B2 genomes. We focused on the gene and its surrounding region with the strongest divergence between B2 and non-B2, the antiporter gene nhaA. This gene is part of the nhaAR operon, which is in the core genome but flanked by mobile regions, and is involved in growth at high pH and high sodium concentrations. Consistently, we found that a panel of non-B2 strains grew faster than B2 at high pH and high sodium concentrations. However, we could not identify differences in expression of the nhaAR operon using fluorescence reporter plasmids. Furthermore, the operon deletion had no differential impact between B2 and non-B2 strains, and did not result in a fitness modification in a murine model of gut colonization. Nevertheless, sequence analysis and experiments in a murine model of septicemia revealed that recombination in nhaA among B2 strains was observed in strains with low virulence. Finally, nhaA and nhaAR operon deletions drastically decreased virulence in one B2 strain. This effect of nhaAR deletion appeared to be stronger than deletion of all pathogenicity islands. Thus, a population genetic approach allowed us to identify an operon in the core genome without strong effect in commensalism but with an important role in extra-intestinal virulence, a landmark of the B2 strains

Mechanisms of nuclear pore complex disassembly by the mitotic Polo-like kinase 1 (PLK-1) in C. elegans embryos

The nuclear envelope, which protects and organizes the genome, is dismantled during mitosis. In the Caenorhabditis elegans zygote, nuclear envelope breakdown (NEBD) of the parental pronuclei is spatially and temporally regulated during mitosis to promote the unification of the maternal and paternal genomes. Nuclear pore complex (NPC) disassembly is a decisive step of NEBD, essential for nuclear permeabilization. By combining live imaging, biochemistry, and phosphoproteomics, we show that NPC disassembly is a stepwise process that involves Polo-like kinase 1 (PLK-1)–dependent and –independent steps. PLK-1 targets multiple NPC subcomplexes, including the cytoplasmic filaments, central channel, and inner ring. PLK-1 is recruited to and phosphorylates intrinsically disordered regions (IDRs) of several multivalent linker nucleoporins. Notably, although the phosphosites are not conserved between human and C. elegans nucleoporins, they are located in IDRs in both species. Our results suggest that targeting IDRs of multivalent linker nucleoporins is an evolutionarily conserved driver of NPC disassembly during mitosis.This work was supported by a PhD fellowship from the French Ministry of Higher Education and Research (to S.N.N.), fourth year PhD fellowship from the Foundation ARC (to S.N.N.), CONACYT grant CVU 364106, and the CM/SECTEI/201/2022 fellowships (to G.V.-A.). Research in the Seydoux laboratory is supported by the NIH (grant no. R37 HD37047) and by the Howard Hughes Medical Institute. This work was also supported by the Agence Nationale pour la Recherche (ANR), France - ANR-17-CE13-0011 (to L.P.); Ligue Nationale Contre le Cancer Equipe Labéllisée, France (to L.P.); Spanish State Research Agency, the European Union, and the European Regional Development Fund (CEX2020-001088-M and PID2019-105069GB-I00; doi: 10.13039/501100011033) (to P.A.).With funding from the Spanish government through the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2020-001088-M).Peer reviewe

Structural basis for Cul3 protein assembly with the BTB-Kelch family of E3 ubiquitin ligases

Cullin-RING ligases are multisubunit E3 ubiquitin ligases that recruit substrate-specific adaptors to catalyze protein ubiquitylation. Cul3-based Cullin-RING ligases are uniquely associated with BTB adaptors that incorporate homodimerization, Cul3 assembly, and substrate recognition into a single multidomain protein, of which the best known are BTB-BACK-Kelch domain proteins, including KEAP1. Cul3 assembly requires a BTB protein "3-box" motif, analogous to the F-box and SOCS box motifs of other Cullin-based E3s. To define the molecular basis for this assembly and the overall architecture of the E3, we determined the crystal structures of the BTB-BACK domains of KLHL11 both alone and in complex with Cul3, along with the Kelch domain structures of KLHL2 (Mayven), KLHL7, KLHL12, and KBTBD5. We show that Cul3 interaction is dependent on a unique N-terminal extension sequence that packs against the 3-box in a hydrophobic groove centrally located between the BTB and BACK domains. Deletion of this N-terminal region results in a 30-fold loss in affinity. The presented data offer a model for the quaternary assembly of this E3 class that supports the bivalent capture of Nrf2 and reveals potential new sites for E3 inhibitor design

A Ubiquitin Ligase Complex Regulates Caspase Activation During Sperm Differentiation in Drosophila

In both insects and mammals, spermatids eliminate their bulk cytoplasm as they undergo terminal differentiation. In Drosophila, this process of dramatic cellular remodeling requires apoptotic proteins, including caspases. To gain further insight into the regulation of caspases, we screened a large collection of sterile male flies for mutants that block effector caspase activation at the onset of spermatid individualization. Here, we describe the identification and characterization of a testis-specific, Cullin-3–dependent ubiquitin ligase complex that is required for caspase activation in spermatids. Mutations in either a testis-specific isoform of Cullin-3 (Cul3Testis), the small RING protein Roc1b, or a Drosophila orthologue of the mammalian BTB-Kelch protein Klhl10 all reduce or eliminate effector caspase activation in spermatids. Importantly, all three genes encode proteins that can physically interact to form a ubiquitin ligase complex. Roc1b binds to the catalytic core of Cullin-3, and Klhl10 binds specifically to a unique testis-specific N-terminal Cullin-3 (TeNC) domain of Cul3Testis that is required for activation of effector caspase in spermatids. Finally, the BIR domain region of the giant inhibitor of apoptosis–like protein dBruce is sufficient to bind to Klhl10, which is consistent with the idea that dBruce is a substrate for the Cullin-3-based E3-ligase complex. These findings reveal a novel role of Cullin-based ubiquitin ligases in caspase regulation

Drosophila Kelch functions with Cullin-3 to organize the ring canal actin cytoskeleton

In addition to cross-linking F-actin, Drosophila Kelch is a component of a cullin-RING ubiquitin ligase complex required for morphogenesis of ring canals during oogenesis

Characterization of the mammalian family of DCN-type NEDD8 E3 ligases

Cullin-RING ligases (CRL) are ubiquitin E3s that bind substrates through variable substrate-receptor proteins. CRLs are activated by attachment of the ubiquitin-like protein NEDD8 to the Cullin subunit and DCNs are NEDD8 E3 ligases that promote neddylation. Mammalian cells express five DCN-like proteins and little is known about their specific functions or interaction partners. We found that DCNLs form stable stoichiometric complexes with CAND1 and Cullins that can only be neddylated in the presence of substrate adaptor. These DCNL-CUL-CAND1 complexes may represent “reserve” CRLs that can be rapidly activated when needed. We further found that all DCNLs interact with most Cullin subtypes, but that they are likely responsible for the neddylation of different subpopulations of any given Cullin. This is consistent with the fact that the subcellular localization of DCNLs in tissue culture cells differs and that they show unique tissue specific expression patterns in mice. Thus, the specificity between DCNL-type NEDD8 E3 enzymes and their Cullin substrates is only apparent in well-defined physiological contexts and related to their subcellular distribution and restricted expression