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International audience; ER stress is mediated by three sensors and the most evolutionary conserved IRE1α signals through its cytosolic kinase and endoribonuclease (RNase) activities. IRE1α RNase activity can either catalyze the initial step of XBP1 mRNA unconventional splicing or degrade a number of RNAs through regulated IRE1-dependent decay. Until now, the biochemical and biological outputs of IRE1α RNase activity have been well documented; however, the precise mechanisms controlling whether IRE1α signaling is adaptive or pro-death (terminal) remain unclear. We investigated those mechanisms and hypothesized that XBP1 mRNA splicing and regulated IRE1-dependent decay activity could be co-regulated by the IRE1α RNase regulatory network. We identified that RtcB, the tRNA ligase responsible for XBP1 mRNA splicing, is tyrosine-phosphorylated by c-Abl and dephosphorylated by PTP1B. Moreover, we show that the phosphorylation of RtcB at Y306 perturbs RtcB interaction with IRE1α, thereby attenuating XBP1 mRNA splicing. Our results demonstrate that the IRE1α RNase regulatory network is dynamically fine-tuned by tyrosine kinases and phosphatases upon various stresses and that the extent of RtcB tyrosine phosphorylation determines cell adaptive or death outputs.

Endoplasmic Reticulum (ER) stress is a hallmark of various diseases. Cells cope with ER stress through the activation of an adaptive signaling pathway named the Unfolded Protein Response (UPR) which is mediated by three ER-resident sensors. The most evolutionary conserved of the UPR sensors is IRE1alpha that elicits diverse downstream effects through its kinase and endoribonuclease (RNase) activities. IRE1alpha RNase activity can either catalyze the initial step of XBP1 mRNA unconventional splicing or degrade a number of RNAs through Regulated IRE1-Dependent Decay (RIDD). The degree of exertion of these two activities plays an instrumental role in cells life and death decisions upon ER stress. Until now, the biochemical and biological outputs of IRE1alpha RNase activity have been well documented, however, the precise mechanisms controlling whether IRE1 signaling is adaptive or pro-death (terminal) remain unclear. This prompted us to further investigate those mechanisms and we hypothesized that XBP1 mRNA splicing and RIDD activity could be co-regulated within the context of the IRE1alpha RNase regulatory network. We showed that a key nexus in this pathway is the tRNA ligase RtcB which, together with IRE1alpha is responsible for XBP1 mRNA splicing. We demonstrated that RtcB is tyrosine phosphorylated by c-Abl and dephosphorylated by PTP1B. Moreover, we identified RtcB Y306 as a key residue which, when phosphorylated, perturbs RtcB interaction with IRE1alpha, thereby attenuating XBP1 mRNA splicing and favoring RIDD. Our results demonstrate that the IRE1alpha RNase regulatory network is dynamically fine-tuned by tyrosine kinases and phosphatases upon various stresses and depending on the nature of the stress determines cell adaptive or death outputs.

International audience; Cancer cells are exposed to intrinsic (oncogene) or extrinsic (microenvironmental) challenges, leading to activation of stress response pathways. The unfolded protein response (UPR) is the cellular response to endoplasmic reticulum (ER) stress and plays a pivotal role in tumor development. Depending on ER stress intensity and duration, the UPR is either pro-survival to preserve ER homeostasis or pro-death if the stress cannot be resolved. On one hand, the adaptive arm of the UPR is essential for cancer cells to survive the harsh conditions they are facing, and on the other hand, cancer cells have evolved mechanisms to bypass ER stress-induced cell death, thereby conferring them with a selective advantage for malignant transformation. Therefore, the mechanisms involved in the balance between survival and death outcomes of the UPR may be exploited as therapeutic tools to treat cancer

C’est a partir d’echanges avec Francoise Galinon, psychiatre psychanalyste, fondatrice de l’association « Bon Pied Bon Œil » que j’ai pu ecrire cet article qui retrace l’histoire de ce gem afin d’eclairer ce que je peux y proposer en tant que vacataire. J’interviens une fois par mois sur l’atelier « aide aux projets ». Cet atelier est ouvert a tout membre qui souhaite envisager un projet d’insertion (le projet doit etre entendu comme toute proposition dans le champ social).

Inositol-requiring enzyme 1 (IRE1) is the most conserved transducer of the unfolded protein response (UPR), a homeostatic response that preserves proteostasis. Intriguingly, via its endoribonuclease activity, IRE1 produces either adaptive or death signals. This occurs through both unconventional splicing of XBP1 mRNA and regulated IRE1-dependent decay of mRNA (RIDD). Whereas XBP1 mRNA splicing is cytoprotective in response to endoplasmic reticulum (ER) stress, RIDD has revealed many unexpected features. For instance, RIDD cleaves RNA at an XBP1-like consensus site but with an activity divergent from XBP1 mRNA splicing and can either preserve ER homeostasis or induce cell death. Here we review recent findings on RIDD and propose a model of how IRE1 RNase activity might control cell fate decisions.

The endoplasmic reticulum (ER)-induced unfolded protein response (ERUPR) is an adaptive mechanism that is activated upon accumulation of misfolded proteins in the ER and aims at restoring ER homeostasis. The ERUPR is transduced by three major ER-resident stress sensors, namely PKR-like endoplasmic reticulum kinase (PERK), activating transcription factor 6 (ATF6), and inositol requiring enzyme 1 (IRE1). Activation of these ER stress sensors leads to transcriptional reprogramming of the cells. Recently, microRNAs (miRNAs), small noncoding RNAs that generally repress gene expression, have emerged as key regulators of ER homeostasis and important players in ERUPR-dependent signaling. Moreover, the miRNAs biogenesis machinery appears to also be regulated upon ER stress. Herein we extensively review the relationships existing between “canonical” ERUPR signaling, control of ER homeostasis, and miRNAs. We reveal an intricate signaling network that might confer specificity and selectivity to the ERUPR in tissue- or stress-dependent fashion. We discuss these issues in the context of the physiological and pathophysiological roles of ERUPR signaling.

Poplars are one of the woody plants that are very sensitive to water stress, which may reduce the productivity of fast- growing plantations. Poplars can exhibit several drought tolerance strategies that may impact productivity differently. Trees from two improved hybrids, Populus balsamifera × Populus trichocarpa Torr. & Gray (clone B × T) and P. balsamifera × Populus maximowiczii A. Henry (clone B × M), having P. balsamifera L. as a parent and trees from native and unimproved P. balsamifera were subjected to a 1-month drying cycle in a growth chamber and then rewatered. The unimproved and native B clone maintained higher stomatal conductance (gs) than the hybrids, and high photosynthetic activity and transpiration, even when soil water content was nearly zero. As a result, both instantaneous water use efficiency (WUE i) and leaf carbon isotope composition (δ 13 C) indicated that this clone was less affected by drought than both hybrids at maximal drought stress. However, this clone shed its leaves when the drought threshold was exceeded, which implied a greater loss of productivity. The B × M hybrid showed a relatively conservative response to water stress, with the greatest decrease in transpiring versus absorbing surface (total leaf area to root biomass ratio). This clone was also the only one to develop new leaves after rewatering, and its total biomass production was not significantly decreased by drought. Among the two hybrids, clone B × T was the most vigorous, with the greatest transpiration (Ei) and net CO2 assimilation (A) rates, allowing for high biomass production. However, it had a more risky strategy under drought conditions by keeping its stomata open and high Ei rates under moderate drought, resulting in a lower recovery rate after rewatering. The opposite drought response strategies of the two hybrids were reflected by clone B × T having lower WUEi values than clone B × M at maximal drought, with a very low Ψmin value of −3.2 MPa, despite closed stomata and stopped photosynthetic activity. Positive linear relationships between A and gs for the three hybrids indicated strong stomatal control of photosynthesis. Moreover, the three poplar clones showed anisohydric behaviour for stomatal control and their use under long-term drought should be of interest, especially the B × M clone.

Steatohepatitis is a cause of hepatocellular carcinoma development; however, the underlying mechanisms are poorly defined. In this issue of Cancer Cell, Nakagawa and colleagues demonstrate that activation of endoplasmic reticulum stress signaling is instrumental in the development of steatohepatitis and synergizes with proinflammatory pathways to promote hepatocarcinogenesis.

The start of university life demands an adaptation period, which can sometimes have an impact on the mental health of students. A total of 233 first year psychology students answered a questionnaire based on a Beck Depression Inventory, Coping Inventory of Stressful Situations, Social Support Questionnaire and Hassles Scale. The results showed 41 per cent of those surveyed suffered of depressive symptoms and that daily hassles encountered during the first year of university could be considered a relevant risk factor for depression. This study makes it possible to observe that not all coping strategies work equally well. Only task-centered, active coping strategies were effective. With many students having to cope with leaving home, social support also plays a fundamental role. In fact, 70 per cent of those interviewed had left their hometown in order to study. The results of this study suggest that depressive symptoms should be considered a significant problem particularly in first year students. Copyright © 2006 John Wiley & Sons, Ltd.

MicroRNAs (miRNA) are generally described as negative regulators of gene expression. However, some evidence suggests that they may also play positive roles. As such, we reported that miR-1291 leads to a GPC3 mRNA expression increase in hepatoma cells through a 3′ untranslated region (UTR)-dependent mechanism. In the absence of any direct interaction between miR-1291 and GPC3 mRNA, we hypothesized that miR-1291 could act by silencing a negative regulator of GPC3 mRNA expression. Based on in silico predictions and experimental validation, we demonstrate herein that miR-1291 represses the expression of the mRNA encoding the endoplasmic reticulum (ER)-resident stress sensor IRE1α by interacting with a specific site located in the 5′ UTR. Moreover, we show, in vitro and in cultured cells, that IRE1α cleaves GPC3 mRNA at a 3′ UTR consensus site independently of ER stress, thereby prompting GPC3 mRNA degradation. Finally, we show that the expression of a miR-1291-resistant form of IRE1α abrogates the positive effects of miR-1291 on GPC3 mRNA expression. Collectively, our data demonstrate that miR-1291 is a biologically relevant regulator of GPC3 expression in hepatoma cells and acts through silencing of the ER stress sensor IRE1α.

International audience; Hepatocellular carcinoma (HCC) is the major primary liver cancer. Glypican-3 (GPC3), one of the most abnormally expressed genes in HCC, participates in liver carcinogenesis. Based on data showing that GPC3 expression is posttranscriptionally altered in HCC cells compared to primary hepatocytes, we investigated the implication of microRNAs (miRNAs) in GPC3 overexpression and HCC. To identify GPC3-regulating miRNAs, we developed a dual-fluorescence FunREG (functional, integrated, and quantitative method to measure posttranscriptional regulations) system that allowed us to screen a library of 876 individual miRNAs. Expression of candidate miRNAs and that of GPC3 messenger RNA (mRNA) was measured in 21 nontumoral liver and 112 HCC samples. We then characterized the phenotypic consequences of modulating expression of one candidate miRNA in HuH7 cells and deciphered the molecular mechanism by which this miRNA controls the posttranscriptional regulation of GPC3. We identified five miRNAs targeting GPC3 3'-untranslated region (UTR) and regulating its expression about the 876 tested. Whereas miR-96 and its paralog miR-1271 repressed GPC3 expression, miR-129-1-3p, miR-1291, and miR-1303 had an inducible effect. We report that miR-1271 expression is down-regulated in HCC tumor samples and inversely correlates with GPC3 mRNA expression in a particular subgroup of HCC. We also report that miR-1271 inhibits the growth of HCC cells in a GPC3-dependent manner and induces cell death.CONCLUSION:Using a functional screen, we found that miR-96, miR-129-1-3p, miR-1271, miR-1291, and miR-1303 differentially control GPC3 expression in HCC cells. In a subgroup of HCC, the up-regulation of GPC3 was associated with a concomitant down-regulation of its repressor miR-1271. Therefore, we propose that GPC3 overexpression and its associated oncogenic effects are linked to the down-regulation of miR-1271 in HCC.

Introduction:Glioblastoma multiforme (GBM) is the most lethal form of glioma with an overall survival at 5 years nearly null (< 5%). Increasing evidences point towards the RNase activity of IRE1 as a central player in GBM development, particularly in cancer cell invasion, tumor vascularization and recruitment of inflammatory or immune cells. Indeed, IRE1 RNase promotes XBP1 mRNA splicing, a well-described cytoprotective transcription factor that is required for cancer cell survival and this activity also contributes to the degradation of mRNA and microRNA, an activity called RIDD. Recent studies unraveled the presence of somatic mutations on the IRE1 gene in GBM that could play a driver role but without providing any functional information.Results:Here, we have sequenced IRE1 exons in 23 human GBMs and identified a new somatic mutation on the IRE1 gene. Interestingly, using biochemical approaches and an orthotopic tumor graft mouse model we show that this variant displays an altered IRE1 RNase activity. IRE1-dependent gene expression pattern alterations lead to major functional consequences in terms of tumor phenotype including (i) a magnified induction of the inflammatory and pro-angiogenic pathways based on transcriptome signatures from human tumors, (ii) an increase of the aggressiveness/infiltration potential of U87 derived tumors in mice and (iii) the modulation of immune cells recruitment to the tumor.Conclusions:This study provides the first evidence that the selectivity of IRE1 RNase can be modulated naturally and unravels the first mechanistic example of how a somatic mutation in the IRE gene can provide adaptive advantages to glioblastoma cells by reprogramming the tumor stroma.

Drug-induced liver injury (DILI) limits the development and utilization of numerous therapeutic compounds, and consequently presents major challenges to the pharmaceutical industry and clinical medicine1, 2. Acetaminophen (APAP) containing compounds are among the most frequently prescribed drugs, and also the most common cause of DILI3. Here we describe a pharmacological strategy that targets gap junction communication to prevent amplification of fulminant hepatic failure and APAP-induced hepatotoxicity. We report that connexin 32 (Cx32), a key hepatic gap junction protein, is an essential mediator of DILI by showing that mice deficient in Cx32 are protected against liver damage, acute inflammation, and death. We identified a small molecule inhibitor of Cx32 as a novel hepatoprotectant that achieves the same result in wildtype mice when coadministered with known hepatotoxic drugs. These findings demonstrate that gap junction inhibition is an effective therapy for limiting DILI, and suggest a novel pharmaceutical strategy to improve drug safety.

International audience; Besides the fact that miR-96 and miR-182 belong to the miR-182/183 cluster, their seed region (UUGGCA, nucleotides 2-7) is identical suggesting potential common properties in mRNA target recognition and cellular functions. Here, we used the mRNA encoding Glypican-3, a heparan-sulfate proteoglycan, as a model target as its short 3' untranslated region is predicted to contain one miR-96/182 site, and assessed whether it is post-transcriptionally regulated by these two microRNAs. We found that miR-96 downregulated GPC3 expression by targeting its mRNA 3'-untranslated region and interacting with the predicted site. This downregulatory effect was due to an increased mRNA degradation and depended on Argonaute-2. Despite its seed similarity with miR-96, miR-182 was unable to regulate GPC3. This differential regulation was confirmed on two other targets, FOXO1 and FN1. By site-directed mutagenesis, we demonstrated that the miRNA nucleotide 8, immediately downstream the UUGGCA seed, plays a critical role in target recognition by miR-96 and miR-182. Our data suggest that because of a base difference at miRNA position 8, these two microRNAs control a completely different set of genes and therefore are functionally independent.

An increasing number of arguments, including altered microRNA expression, support the idea that post-transcriptional deregulation participates in gene disturbances found in diseased tissues. To evaluate this hypothesis, we developed a method which facilitates post-transcriptional investigations in a wide range of human cells and experimental conditions. This method, called FunREG (functional, integrated and quantitative method to measure post-transcriptional regulation), connects lentiviral transduction with a fluorescent reporter system and quantitative PCR. Using FunREG, we efficiently measured post-transcriptional regulation mediated either by selected RNA sequences or regulatory factors (microRNAs), and then evaluated the contribution of mRNA decay and translation efficiency in the observed regulation. We demonstrated the existence of gene-specific post-transcriptional deregulation in liver tumour cells, and also reported a molecular link between a transcript variant abrogating HDAC6 (histone deacetylase 6) regulation by miR-433 and a rare familial genetic disease. Because FunREG is sensitive, quantitative and easy to use, many applications can be envisioned in fundamental and pathophysiological research.

Objectives CT-scan screening programs for lung cancer detection have been proposed in high-risk subjects, and more recently in former asbestos-exposed subjects. However, to date no data are available on psychological impact of such programs. The aim of this study is to examine the risk factors of psychological distress at baseline of a CT-scan screening program among asbestos-exposed subjects. Methods The Asbestos Post-Exposure Survey (APEXS) was carried out in France between October 2003 and December 2005 in order to screen asbestos-related diseases by CT-scan. Volunteers underwent self-administered questionnaires including an asbestos exposure assessment and, for a large sub-sample, a validated psychological distress scale. Non-exposed subjects were used as reference group. Results At baseline, a significant higher level of distress was observed in exposed subjects (n = 3,122) relative to the reference group (n = 486) after adjustment on age, sex, and tobacco status. This distress is associated independently with the self-perception of (i) intensity of asbestos exposure and (ii) the risk of current or future disease related to the asbestos exposure. The perception of the cancer risk related to asbestos seems to play a fundamental role in this psychological distress. Conclusion In this study, asbestos-exposed subjects experienced a higher significant cancer distress than previously described in literature. These findings may be of potential public health importance. First, the impact of such occupational exposures on quality of life of patients who suffer from cancer related to these exposures has to be appraised. Secondly, the assessment of psychological impact of CT-scan screening programs among asbestos-exposed subjects is also required. Am. J. Ind. Med. 52:288–296, 2009. © 2009 Wiley-Liss, Inc.

International audience; The effects of Phytophthora cinnamomi root damage on water relations of chestnut (Castanea sativa) saplings were investigated. The relationship between root damage severity and impact on water relations was studied using a four compartment split-root system. Saplings were submitted to five inoculation levels and two watering conditions: normal (well-watered) or restricted watering. In well-watered saplings, stomatal conductance and whole plant transpiration rate were negatively correlated to the proportion of necrotic roots. Nevertheless, plant hydraulic conductance and leaf water potential were only affected above 90% necrotic roots. Interaction between root damage and water restriction was difficult to assess, since soil moisture decreased only slightly in infested compartments. However, saplings under restricted watering displayed lower stomatal conductance and transpiration values, regardless of root damage severity. Furthermore, the threshold of root damage leading to a decrease in leaf water potential was lower under restricted watering than under normal watering.; Effets de dégâts racinaires variables causés par Phytophthora cinnamomi sur le fonctionnement hydrique de jeunes châtaigniers. Les effets des dégâts racinaires causés par Phytophthora cinnamomi sur le fonctionnement hydrique de jeunes châtaigniers (Castanea sativa) ont été étudiés. Afin de relier les effets observés à la sévérité des dégâts racinaires, un système de "split-root " à quatre compartiments a été utilisé. Les arbres ont été soumis à cinq niveaux d'inoculation et à deux régimes hydriques (arrosage normal ou arrosage réduit). Chez les arbres normalement arrosés, une diminution linéaire de la conductance stomatique et de la transpiration avec la proportion de racines nécrosées a été observée. Cependant, la conductance hydraulique de l'arbre et le potentiel hydrique foliaire n'ont été affectés qu'à partir de 90 % de racines nécrosées. L'interaction entre les dégâts racinaires et la restriction en eau a été difficile à évaluer car l'humidité du sol a faiblement baissé dans les compartiments inoculés. Cependant, les arbres soumis a une restriction en eau avaient une conductance stomatique et une transpiration faibles quelle que soit la proportion de racines nécrosées. De plus, le seuil de dégâts racinaires entraînant une baisse de potentiel hydrique foliaire était plus faible chez les arbres soumis à une restriction en eau que chez les arbres normalement arrosés.

Background The aim of this study was to assess the psychological consequences of HRCT scan screening in retired asbestos-exposed workers. Methods A HRCT-scan screening program for asbestos-related diseases was carried out in four regions of France. At baseline (T1), subjects filled in self-administered occupational questionnaires. In two of the regions, subjects also received a validated psychological scale, namely the psychological consequences questionnaire (PCQ). The physician was required to provide the subject with the results of the HRCT scan at a final visit. A second assessment of psychological consequences was performed 6 months after the HRCT-scan examination (T2). PCQ scores were compared quantitatively (t-test, general linear model) and qualitatively (chi²-test, logistic regression) to screening results. Multivariate analyses were adjusted for gender, age, smoking, asbestos exposure and counseling. Results Among the 832 subjects included in this psychological impact study, HRCT-scan screening was associated with a significant increase of the psychological score 6 months after the examination relative to baseline values (8.31 to 10.08, p < 0.0001, t-test). This increase concerned patients with an abnormal HRCT-scan result, regardless of the abnormalities, but also patients with normal HRCT-scans after adjustment for age, gender, smoking status, asbestos exposure and counseling visit. The greatest increase was observed for pleural plaques (+3.60; 95%CI [+2.15;+5.06]), which are benign lesions. Detection of isolated pulmonary nodules was also associated with a less marked but nevertheless significant increase of distress (+1.88; 95%CI [+0.34;+3.42]). However, analyses based on logistic regressions only showed a close to significant increase of the proportion of subjects with abnormal PCQ scores at T2 for patients with asbestosis (OR = 1.92; 95%CI [0.97-3.81]) or with two or more diseases (OR = 2.04; 95%CI [0.95-4.37]). Conclusion This study suggests that HRCT-scan screening may be associated with increased distress in asbestos-exposed subjects. If confirmed, these results may have consequences for HRCT-scan screening recommendations.