The Involvement of SMILE/TMTC3 in Endoplasmic Reticulum Stress Response

The state of operational tolerance has been detected sporadically in some renal transplanted patients that stopped immunosuppressive drugs, demonstrating that allograft tolerance might exist in humans. Several years ago, a study by Brouard et al. identified a molecular signature of several genes that were significantly differentially expressed in the blood of such patients compared with patients with other clinical situations. The aim of the present study is to analyze the role of one of these molecules over-expressed in the blood of operationally tolerant patients, SMILE or TMTC3, a protein whose function is still unknown.We first confirmed that SMILE mRNA is differentially expressed in the blood of operationally tolerant patients with drug-free long term graft function compared to stable and rejecting patients. Using a yeast two-hybrid approach and a colocalization study by confocal microscopy we furthermore report an interaction of SMILE with PDIA3, a molecule resident in the endoplasmic reticulum (ER). In accordance with this observation, SMILE silencing in HeLa cells correlated with the modulation of several transcripts involved in proteolysis and a decrease in proteasome activity. Finally, SMILE silencing increased HeLa cell sensitivity to the proteasome inhibitor Bortezomib, a drug that induces ER stress via protein overload, and increased transcript expression of a stress response protein, XBP-1, in HeLa cells and keratinocytes.In this study we showed that SMILE is involved in the endoplasmic reticulum stress response, by modulating proteasome activity and XBP-1 transcript expression. This function of SMILE may influence immune cell behavior in the context of transplantation, and the analysis of endoplasmic reticulum stress in transplantation may reveal new pathways of regulation in long-term graft acceptance thereby increasing our understanding of tolerance

Étude du phénotype odontoblastique (caractérisation de deux nouvelles protéines)

LYON1-BU.Sciences (692662101) / SudocSudocFranceF

Les amelogenèses imparfaites (de la compréhension des mécanismes moléculaires au traitement d'un cas rencontré au CSTD de Lyon)

LYON1-BU Santé Odontologie (693882213) / SudocPARIS-BIUM (751062103) / SudocSudocFranceF

Expression et localisation de la rééline et de la sémaphorine 7A au cours de l'innervation du complexe dentino-pulpaire

Au cours de l'odontogenèse, la croissance et la mise en place des fibres nerveuses pulpaires sont contrôlées par des signaux fournis par des molécules de signalisation et d'adhésion. L'objectif de ce travail est de comprendre le rôle de la rééline et de la SEMA 7A et leur implication dans la relation nerfs/odontoblastes. L'expression de la rééline, de la SEMA 7A et de leurs récepteurs a été étudiée dans la pulpe dentaire humaine, dans les odontoblastes en culture et dans le ganglion trigéminal de rat. Nos résultats montrent une expression spécifique de ces molécules dans les odontoblastes humains in vivo et in vitro et une expression de leurs récepteurs dans le ganglion trigéminal et les fibres nerveuses pulpaires. Les co-cultures ganglions trigéminaux/odontoblastes ont révélé que la rééline pourrait être impliquée dans l'adhésion des fibres nerveuses au contact des odontoblastes, tandis que la SEMA 7A pourrait favoriser la croissance des fibres nerveuses au contact de ces derniersLYON1-BU Santé Odontologie (693882213) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

A micro-evolutionary change in target binding sites as a key determinant of Ultrabithorax function in Drosophila

Abstract Hox genes encode Homeodomain-containing transcription factors, which specify segmental identities along the anterior-posterior axis. Functional changes in Hox genes have been directly implicated in the evolution of body plans across the metazoan lineage. The Hox protein Ultrabithorax (Ubx) is expressed and required in developing third thoracic (T3) segments in holometabolous insects studied so far, particularly, of the order Coleoptera, Lepidoptera and Diptera. Ubx function is key to specify differential development of the second (T2) and T3 thoracic segments in these insects. While Ubx is expressed in the third thoracic segment in developing larvae of Hymenopteran Apis mellifera , the morphological differences between T2 and T3 are subtle. To identify evolutionary changes that are behind the differential function of Ubx in these two insects, which are diverged for more than 350 million years, we performed comparative analyses of genome wide Ubx-binding sites between Drosophila and Apis . Our studies reveal that a motif with a TAAAT core is a preferred binding site for Ubx in Drosophila , but not in Apis . Biochemical and transgenic assays suggest that in Drosophila , TAAAT core sequence in the Ubx binding sites is required for Ubx-mediated regulation of two of its target genes studied here. CG13222 , a gene that is normally upregulated by Ubx and vestigial ( vg ), whose expression is repressed by Ubx in T3. Interestingly, changing the TAAT site to a TAAAT site was sufficient to bring an otherwise unresponsive enhancer of the vg gene from Apis under the control of Ubx in a Drosophila transgenic assay. Taken together, our results suggest an evolutionary mechanism by which critical wing patterning genes might have come under the regulation of Ubx in the Dipteran lineage

A micro-evolutionary change in target binding sites as a key determinant of Ultrabithorax function in Drosophila

International audienceAbstract Hox genes encode Homeodomain-containing transcription factors, which specify segmental identities along the anterior-posterior axis. Functional changes in Hox genes have been directly implicated in the evolution of body plans across the metazoan lineage. The Hox protein Ultrabithorax (Ubx) is expressed and required in developing third thoracic (T3) segments in holometabolous insects studied so far, particularly, of the order Coleoptera, Lepidoptera and Diptera. Ubx function is key to specify differential development of the second (T2) and T3 thoracic segments in these insects. While Ubx is expressed in the third thoracic segment in developing larvae of Hymenopteran Apis mellifera , the morphological differences between T2 and T3 are subtle. To identify evolutionary changes that are behind the differential function of Ubx in these two insects, which are diverged for more than 350 million years, we performed comparative analyses of genome wide Ubx-binding sites between Drosophila and Apis . Our studies reveal that a motif with a TAAAT core is a preferred binding site for Ubx in Drosophila , but not in Apis . Biochemical and transgenic assays suggest that in Drosophila , TAAAT core sequence in the Ubx binding sites is required for Ubx-mediated regulation of two of its target genes studied here. CG13222 , a gene that is normally upregulated by Ubx and vestigial ( vg ), whose expression is repressed by Ubx in T3. Interestingly, changing the TAAT site to a TAAAT site was sufficient to bring an otherwise unresponsive enhancer of the vg gene from Apis under the control of Ubx in a Drosophila transgenic assay. Taken together, our results suggest an evolutionary mechanism by which critical wing patterning genes might have come under the regulation of Ubx in the Dipteran lineage

Cytokine production by human odontoblast-like cells upon Toll-like receptor-2 engagement

International audienceRecent studies have suggested that odontoblasts are involved in the dental pulp immune response to oral pathogens that invade human dentin during the caries process. How odontoblasts regulate the early inflammatory and immune pulp response to Gram-positive bacteria, which predominate in shallow and moderate dentin caries, is still poorly understood. In this study, we investigated the production of pro-and anti-inflammatory cytokines by odontoblast-like cells upon engagement of Toll-like receptor (TLR) 2, a pattern recognition molecule activated by Gram-positive bacteria components. We used a highly sensitive Milliplex (R) kit for detecting cytokines released by cells stimulated with lipoteichoic acid (LTA), a cell wall component of Gram-positive bacteria, or with the potent TLR2 synthetic agonist Pam2CSK4. We found that odontoblasts produce the pro-inflammatory cytokines interleukin (IL)-6 and CXCL8, as well as the immunosuppressive cytokine IL-10 in response to TLR2 agonists. GM-CSF, IFN gamma, IL-1 beta, IL-2, IL-4, IL-5, IL-7, IL-12(p70), IL-13 and TNF-alpha were not detected. These data indicate that TLR2 activation in human odontoblasts selectively induces production of mediators known to influence positively or negatively inflammatory and immune responses in pathogen-challenged tissues. We suggest that these molecules might be important in regulating the fine tuning of the pulp response to Gram-positive bacteria which enter dentin during the caries process. (C) 2010 Elsevier GmbH. All rights reserved

Human HOX Proteins Use Diverse and Context-Dependent Motifs to Interact with TALE Class Cofactors

Summary: HOX proteins achieve numerous functions by interacting with the TALE class PBX and MEIS cofactors. In contrast to this established partnership in development and disease, how HOX proteins could interact with PBX and MEIS remains unclear. Here, we present a systematic analysis of HOX/PBX/MEIS interaction properties, scanning all paralog groups with human and mouse HOX proteins in vitro and in live cells. We demonstrate that a previously characterized HOX protein motif known to be critical for HOX-PBX interactions becomes dispensable in the presence of MEIS in all except the two most anterior paralog groups. We further identify paralog-specific TALE-binding sites that are used in a highly context-dependent manner. One of these binding sites is involved in the proliferative activity of HOXA7 in breast cancer cells. Together these findings reveal an extraordinary level of interaction flexibility between HOX proteins and their major class of developmental cofactors. : Dard et al. examine interaction flexibility between HOX proteins and TALE cofactors. The authors identify specific TALE-binding sites in different human HOX proteins. Most of these TALE-binding sites correspond to short motifs that are differently used depending on the DNA-binding site or cell context. Keywords: HOX, PBX, MEIS, BiFC, SLiM, TALE, homeodomain, transcription, protein interactio

Human odontoblast-like cells produce nitric oxide with antibacterial activity upon TLR2 activation

International audienc