Parasite Manipulation of the Invariant Chain and the Peptide Editor H2-DM Affects Major Histocompatibility Complex Class II Antigen Presentation during Toxoplasma gondii Infection

Toxoplasma gondii is an obligate intracellular protozoan parasite. This apicomplexan is the causative agent of toxoplasmosis, a leading cause of central nervous system disease in AIDS. It has long been known that T. gondii interferes with major histocompatibility complex class II (MHC-II) antigen presentation to attenuate CD4(+) T cell responses and establish persisting infections. Transcriptional downregulation of MHC-II genes by T. gondii was previously established, but the precise mechanisms inhibiting MHC-II function are currently unknown. Here, we show that, in addition to transcriptional regulation of MHC-II, the parasite modulates the expression of key components of the MHC-II antigen presentation pathway, namely, the MHC-II-associated invariant chain (Ii or CD74) and the peptide editor H2-DM, in professional antigen-presenting cells (pAPCs). Genetic deletion of CD74 restored the ability of infected dendritic cells to present a parasite antigen in the context of MHC-II in vitro. CD74 mRNA and protein levels were, surprisingly, elevated in infected cells, whereas MHC-II and H2-DM expression was inhibited. CD74 accumulated mainly in the endoplasmic reticulum (ER), and this phenotype required live parasites, but not active replication. Finally, we compared the impacts of genetic deletion of CD74 and H2-DM genes on parasite dissemination toward lymphoid organs in mice, as well as activation of CD4(+) T cells and interferon gamma (IFN-γ) levels during acute infection. Cyst burdens and survival during the chronic phase of infection were also evaluated in wild-type and knockout mice. These results highlight the fact that the infection is influenced by multiple levels of parasite manipulation of the MHC-II antigen presentation pathway

Fast and Flexible Software Polar List Decoders

International audienceFlexibility is one mandatory aspect of channel coding in modern wireless communication systems. Among other things, the channel decoder has to support several code lengths and code rates. This need for flexibility applies to polar codes that are considered for control channels in the future 5G standard. This paper presents a new generic and flexible implementation of a software Successive Cancellation List (SCL) decoder. A large set of parameters can be fine-tuned dynamically without re-compiling the software source code: the code length, the code rate, the frozen bits set, the puncturing patterns, the cyclic redundancy check, the list size, the type of decoding algorithm, the tree-pruning strategy and the data quantization. This generic and flexible SCL decoder enables to explore tradeoffs between throughput, latency and decoding performance. Several optimizations are proposed to achieve a competitive decoding speed despite the constraints induced by the genericity and the flexibility. The resulting polar list decoder is about 4 times faster than a generic software decoder and only 2 times slower than a non-flexible unrolled decoder. Thanks to the flexibility of the decoder, the fully adaptive SCL algorithm can be easily implemented and achieves higher throughput than any other similar decoder in the literature (up to 425 Mb/s on a single processor core for N = 2048 and K = 1723 at 4.5 dB)

The hydrophobic region of the Leishmania peroxin 14 : requirements for association with a glycosome mimetic membrane

This work was funded by operating grants from the Canadian Institutes of Health Research (CIHR) and a Natural Sciences Engineering Research Council of Canada (NSERC) Discovery grant [Fonds de recherche du Québec — Nature et technologies (FRQNT) Regroupement Stratégique grant to the Centre for Host-Parasite Interactions (A.J.)]. N.C. was supported by a doctoral research scholarship from FRQNT. E.B. was supported by a Banting postdoctoral fellowship from CIHR. This work was also supported in part by Wellcome Trust grants [086658 and 093228] to T.K.S. C.S. recognizes the financial support from the Natural Sciences and Engineering Research Council of Canada and a Canada Foundation for Innovation grant [16299].Protein import into the Leishmania glycosome requires docking of the cargo-loaded peroxin 5 (PEX5) receptor to the peroxin 14 (PEX14) bound to the glycosome surface. To examine the LdPEX14-membrane interaction, we purified L. donovani promastigote glycosomes and determined the phospholipid and fatty acid composition. These membranes contained predominately phosphatidylethanolamine, phosphatidylcholine, and phosphatidylglycerol (PG) modified primarily with C18 and C22 unsaturated fatty acid. Using large unilamellar vesicles (LUVs) with a lipid composition mimicking the glycosomal membrane in combination with sucrose density centrifugation and fluorescence-activated cell sorting technique, we established that the LdPEX14 membrane-binding activity was dependent on a predicted transmembrane helix found within residues 149-179. Monolayer experiments showed that the incorporation of PG and phospholipids with unsaturated fatty acids, which increase membrane fluidity and favor a liquid expanded phase, facilitated the penetration of LdPEX14 into biological membranes. Moreover, we demonstrated that the binding of LdPEX5 receptor or LdPEX5-PTS1 receptor-cargo complex was contingent on the presence of LdPEX14 at the surface of LUVs.PostprintPeer reviewe

The Protozoan Parasite Toxoplasma gondii Selectively Reprograms the Host Cell Translatome

The intracellular parasite Toxoplasma gondii promotes infection by targeting multiple host cell processes; however, whether it modulates mRNA translation is currently unknown. Here, we show that infection of primary murine macrophages with type I or II T. gondii strains causes a profound perturbation of the host cell translatome. Notably, translation of transcripts encoding proteins involved in metabolic activity and components of the translation machinery was activated upon infection. In contrast, the translational efficiency of mRNAs related to immune cell activation and cytoskeleton/cytoplasm organization was largely suppressed. Mechanistically, T. gondii bolstered mechanistic target of rapamycin (mTOR) signaling to selectively activate the translation of mTOR-sensitive mRNAs, including those with a 5'-terminal oligopyrimidine (5' TOP) motif and those encoding mitochondrion-related proteins. Consistent with parasite modulation of host mTOR-sensitive translation to promote infection, inhibition of mTOR activity suppressed T. gondii replication. Thus, selective reprogramming of host mRNA translation represents an important subversion strategy during T. gondii infection

Damaged DNA Binding Protein 2 Plays a Role in Breast Cancer Cell Growth

The Damaged DNA binding protein 2 (DDB2), is involved in nucleotide excision repair as well as in other biological processes in normal cells, including transcription and cell cycle regulation. Loss of DDB2 function may be related to tumor susceptibility. However, hypothesis of this study was that DDB2 could play a role in breast cancer cell growth, resulting in its well known interaction with the proliferative marker E2F1 in breast neoplasia. DDB2 gene was overexpressed in estrogen receptor (ER)-positive (MCF-7 and T47D), but not in ER-negative breast cancer (MDA-MB231 and SKBR3) or normal mammary epithelial cell lines. In addition, DDB2 expression was significantly (3.0-fold) higher in ER-positive than in ER-negative tumor samples (P = 0.0208) from 16 patients with breast carcinoma. Knockdown of DDB2 by small interfering RNA in MCF-7 cells caused a decrease in cancer cell growth and colony formation. Inversely, introduction of the DDB2 gene into MDA-MB231 cells stimulated growth and colony formation. Cell cycle distribution and 5 Bromodeoxyuridine incorporation by flow cytometry analysis showed that the growth-inhibiting effect of DDB2 knockdown was the consequence of a delayed G1/S transition and a slowed progression through the S phase of MCF-7 cells. These results were supported by a strong decrease in the expression of S phase markers (Proliferating Cell Nuclear Antigen, cyclin E and dihydrofolate reductase). These findings demonstrate for the first time that DDB2 can play a role as oncogene and may become a promising candidate as a predictive marker in breast cancer

Subversion of MHC-II antigen presentation by «Toxoplasma gondii» involves parasite secretory organelles and the modulation of host immune effectors in the endocytic pathway

The obligate intracellular protozoan parasite Toxoplasma gondii, the causative agent of toxoplasmosis, is a highly ubiquitous pathogen that infects virtually any warm-blooded animal. Although the infection generally remains asymptomatic in healthy individuals, the parasite invariably encysts. It has been shown that T. gondii is able to achieve this goal at least by interfering with MHC-II antigen presentation to dampen the development of the CD4+ T helper cell response and gain a head start on the host adaptive immune system. Previous reports have shown that T. gondii inhibits transcription of MHC-II and several other related genes, but the causative inhibitory molecules have yet to be identified.In an attempt to identify these molecules, a forward genetic screening strategy was initially elaborated, with a genome-wide insertional mutagenesis carried out in order to disrupt the genes encoding for the inhibitory molecules followed. By specifically isolating mutants unable to inhibit MHC-II expression and antigen presentation by flow cytometry, isolation of the disrupted loci and database mining could have enabled the identification by of the encoded inhibitory molecules. However, the screen was not carried out due to experimental limitations.Biochemical analyses were employed to further characterize the MHC-II inhibitory activity, revealing that this activity segregated with the high-speed supernatant (HSS) prepared from sonicated parasites and was enriched with increasing centrifugal speeds. The inhibitory activity was protein dose-dependent and was completely abrogated when the HSS was treated with a broad-spectrum protease. Subcellular fractionation revealed that the inhibitory activity was found in fractions enriched with secretory organelles, specifically rhoptries (ROP) and/or dense granules (GRA). Furthermore, excreted-secreted antigens (ESA) from freshly egressed tachyzoites displayed inhibitory activity. Proteins from ESA preparations were separated by a two-step fractionation using ion exchange chromatography followed size-exclusion chromatography, and analyzed by tandem mass spectrometry (MS/MS). Database mining of the MS/MS results generated a list of possible candidates, the majority of which originated from secretory organelles.Although MHC-II expression was inhibited, low levels of MHC-II molecules were still detected in infected or lysate-treated cells. Experimental results argued that the first layer of transcriptional regulation has to be complemented by a post-translational layer of interference in the host cell endocytic pathway, involving the MHC-II associated invariant chain (Ii), and peptide editor H2-DM. Ii mRNA and protein levels were induced in T. gondii-infected cells, while MHC-II and H2-DM IFN-induced expression was down-regulated. Ii accumulated in infected cells from 20 hour post-infection until host cell lysis, mainly in the ER. In Ii KO cells, the absence of Ii restored the ability of infected bone marrow-derived dendritic cells to present a parasite antigen in the context of MHC-II, arguing that Ii acts as a dominant negative on MHC-II-restricted antigen presentation of endogenously acquired parasite antigens. Keys host proteases, namely legumain, and cathepsins L and S, and the acidification of endosomal compartments were modulated by the parasite, pointing toward a wider manipulation of the host endocytic pathway by T. gondii. Opposing expression patterns of Ii and H2-DM had a drastic effect in vivo not only on parasite dissemination towards lymphoid organs, CD4+ T cell activation, and IFNγ production during acute infection, but also on cyst numbers and survival at the chronic phase of the infection. Altogether, these findings reveal a broader manipulation of host cell processes by T. gondii, and shed new light on the intricate interactions between intracellular pathogens and host cells, and their ability to subvert immune functions to establish successful infections.Le parasite protozoaire intracellulaire obligatoire Toxoplasma gondii, l'agent causant la toxoplasmose, est un pathogène ubiquitaire capable d'infecter tout animal à sang chaud. En dépit du fait que l'infection reste généralement asymptomatique chez les individus en bonne santé, le parasite s'enkyste inévitablement. Il a été démontré que T. gondii est capable d'atteindre ce but en partie en interférant avec la présentation d'antigène par le complexe majeur d'histocompatibilité (CMH)-II pour diminuer le développement de la réponse des cellules T CD4+ et pour ainsi devancer la réponse adaptative du système immunitaire. Il a été démontré que T. gondii inhibe la transcription de CMH-II et autres gènes liés, mais les molécules inhibitrices causatives restent inconnues.Pour identifier ces molécules, une stratégie pour un criblage génétique avait été élaborée. Une mutagénèse insertionelle à travers le génome devait être conduite pour perturber les gènes codant pour ces molécules inhibitrices, suivie d'un tri par cytométrie en flux de cellules infectées avec des mutants. En isolant les mutants incapables d'inhiber l'expression de CMH-II, l'isolement des locus génétiques et l'exploration des bases de données auraient pu permettre l'identification des molécules codées. Cependant, ce criblage ne fut complété dû à des limitations expérimentales. Des analyses biochimiques ont démontré que l'activité inhibitrice se retrouvait dans le surnageant d'haute-vitesse (HSS) préparé à partir de parasites soniqués, et était enrichie avec des vitesses de centrifugation croissantes. L'activité inhibitrice était dose-dépendante de protéines et était complètement abrogée lorsque le SHV était préalablement traité avec une protéase. Un fractionnement subcellulaire révéla que l'activité se retrouvait dans les fractions enrichies d'organelles sécrétoires (rhoptries et granules denses). Aussi, des antigènes excrétés-sécrétés (ESA) obtenus de tachyzoïtes fraîchement lysés possédaient une activité inhibitrice. Les protéines d'ESA furent séparées par fractionnement en deux étapes, commençant par une chromatographie par échange d'ions, suivi par une chromatographie d'exclusion par taille et analysées par spectrométrie de masse en tandem (MS/MS). Les résultats obtenus furent comparés aux bases de données, et une liste fut dressée avec de candidats potentiels, la majorité d'entre eux provenant d'organelles de sécrétion.Malgré l'expression réduite, quelques molécules de CMH-II étaient détectés dans les cellules infectées ou traitées. Nos résultats démontrent que la régulation transcriptionelle doit être complémentée par une interférence au niveau post-traductionel dans la voie endocytique de la cellule hôte qui implique la chaîne invariante associée au CMH-II (Ii) et l'éditeur de peptide H2-DM. Les niveaux d'ARNm et de protéines d'Ii étaient induits dans les cellules infectées, alors que ceux de CMH-II et d'H2-DM étaient inhibés. Ii s'accumulait dans les cellules infectées dès 20 heures post-infection, principalement dans le RE. Dans les cellules Ii KO, l'absence d'Ii rétablit la capacité de cellules dendritiques à présenter un antigène du parasite dans le contexte de CMH-II, proposant ainsi qu'Ii agit comme un dominant négatif sur la présentation d'antigènes endogènes provenant du parasite. Des protéases de l'hôte (légumaine, cathepsines L et S) et l'acidification des compartiments endosomaux étaient modulées par le parasite, révélant une manipulation plus étendue de la voie endocytique. Les modes d'expression opposés d'Ii et H2-DM avaient un effet in vivo sur la dissémination des parasites vers des organes lymphoïdes, l'activation des cellules T CD4+, la production d'IFN, le nombre de kystes et la survie. Collectivement, nos résultats montrent l'étendue des processus de manipulation par T. gondii, révélant de complexes interactions avec l'hôte et sa capacité de subvertir les fonctions immunitaires pour établir une infection chronique

Spondylodiscitis Due to Clostridium ramosum Infection in an Immunocompetent Elderly Patient

The first ever case of spondylodiscitis caused by Clostridium ramosum in an elderly immunocompetent patient has been reported. C. ramosum is usually an intestinal bacterium but may occasionally be isolated in clinical specimens as an opportunistic pathogen. This report shows that this anaerobic organism can cause bone tropism without there having been any contamination due to spinal surgery. The infection cleared after empirical therapy using intravenous amoxicillin and oral metronidazole

Low-temperature crystallization of MgSiO3 glasses under electron irradiation: Possible implications for silicate dust evolution in circumstellar environments

Synthetic MgSiO3 glasses were irradiated at room temperature by 300 keV electrons in a transmission electron microscope. One of the samples had been previously irradiated by 50 keV He+ ions. Electron irradiation induces the nucleation and growth of randomly oriented nanometer-sized crystallites. The crystallites first consist of MgO and subsequently of forsterite (Mg2SiO4). Both are seen to form within an amorphous SiO2 matrix. The rate of crystallisation of the samples has been monitored by conventional TEM imaging and electron diffraction. The sample that had been pre-irradiated with He+ ions is found to transform faster than the as-quenched glass. The crystallization of metastable MgSiO3 glasses is explained by ionising radiation-induced elemental diffusion which allows the reorganization of matter into a more favourable thermodynamic state. These results show that ionizing radiation interactions could account for crystal formation as observed in IR spectroscopy in some young stellar environments.The Meteoritics & Planetary Science archives are made available by the Meteoritical Society and the University of Arizona Libraries. Contact [email protected] for further information.Migrated from OJS platform February 202