Cell cycle progression of Saccharomyces cerevisiae cells was monitored in continuous cultures limited for glucose or nitrogen. The G1 cell cycle phase, before initiation of DNA replication, did not exclusively expand when growth rate decreased. Especially during nitrogen limitation, non-G1 phases expanded almost as much as G1. In addition, cell size remained constant as a function of growth rate. These results contrast with current views that growth requirements are met before initiation of DNA replication, and suggest that distinct nutrient limitations differentially impinge on cell cycle progression. Therefore, multiple mechanisms are hypothesized to regulate the coordination of cell growth and cell division. Genetic interactions were identified between the dose-dependent cell-cycle regulator 2 (DCR2) phosphatase and genes involving in secretion/unfolded protein response pathway, including IRE1, through a genome-wide dominant negative genetic approach. Accumulation of unfolded proteins in the endoplasmic reticulum triggers the unfolded protein response (UPR). How the UPR is downregulated is not well understood. Inositol requirement 1 (IRE1) is an endoplasmic reticulum transmembrane UPR sensor in Saccharomyces cerevisiae. When the UPR is triggered, Ire1p is autophosphorylated, on Ser 840 and Ser 841, inducing the cytosolic endonuclease activity of Ire1p, thereby initiating the splicing and translational de-repression of HAC1 mRNA. Homologous to Atf/Creb1 (Hac1p) activates UPR transcription. We found that that Dcr2p phosphatase functionally and physically interacts with Ire1p. Overexpression of DCR2, but not of a catalytically inactive DCR2 allele, significantly delays HAC1 splicing and sensitizes cells to the UPR. Furthermore, Dcr2p physically interacts in vivo with Ire1p-S840E, S841E, which mimics phosphorylated Ire1p, and Dcr2p dephosphorylates Ire1p in vitro. Our results are consistent with de-phosphorylation of Ire1p being a mechanism for antagonizing UPR signaling.

Les bactéries intracellulaires Wolbachia ont développé une vaste gamme d’interactions symbiotiques, du parasitisme reproductif au mutualisme chez les arthropodes terrestres et les nématodes filaires, devenant ainsi les endosymbiotes les plus répandus sur terre. Bien qu’elles se développent lentement dans les cultures cellulaires d’insectes pour lesquelles les marqueurs sont limités et qu’elles ne sont génétiquement pas manipulables, il existe un intéret croissant de déchiffrer leur mode de vie intracellulaire pour 2 raisons. Premièrement, Wolbachia intervient dans le développement et la transmission des arbovirus et deuxièmement, les filarioses lymphatiques sont traitables grâce à la susceptibilité des Wolbachia qui infectent les nématodes filaires aux antibiotiques. Au début de ce projet, j’ai infecté 2 lignées cellulaires de Drosophila melanogaster qui sont transcriptomiquement divergentes par une même souche de Wolbachia pouvant naturellement infecter Drosophila melanogaster. J’ai utilisé ces 2 lignées cellulaires qui sont différentiellement permissive à l’infection pour explorer l’interaction de Wolbachia avec le réticulum endoplasmique. Les observations par microscopie à fluorescence en temps réel et par microscopie électronique prouvent que cet organite est une source de membranes pour Wolbachia et possiblement, une source de nutriments. Pourtant, les analyses d’expression génique et les approches d’immunofluorescence démontrent que Wolbachia n’induit ni un stress au niveau du réticulum endoplasmique ni une protéolyse via la voie de signalisation ERAD suggérant dès lors, que Wolbachia subvertissent d’autres mécanismes pour assurer leur besoin en acides aminés. Au cours de ce projet, j’ai commencé à mettre en place une technique pour transformer Wolbachia par biolistique. La validation de cette technique de transformation a ouvert la voie vers l’optimisation de la procédure de sélection des transformants pour enfin pouvoir génétiquement manipuler Wolbachia. / The intracellular bacteria Wolbachia have developed a wide range of symbiotic interactions, from being opportunistic reproductive parasites to mutualists with terrestrial arthropods and filarial nematode species, making them the most common endosymbionts on earth. The discovery that they interfere with arboviruses development and transmission by mosquito vectors and that filarial diseases can be cured by targeting Wolbachia, have created a strong interest in deciphering the mechanisms underlying their intracellular lifestyle. However, being obligate intracellular endosymbionts, Wolbachia remain genetically intractable. They grow slowly in insect cell cultures, for which markers are limited. Despite these obstacles, and to limit cell line-specific phenotypes, I chose to infect 2 Drosophila melanogaster cell lines presenting different sets of expressed genes, with a unique Wolbachia strain, naturally hosted by Drosophila melanogaster. Using these 2 cell lines that are differently permissive to the infection, I explored the interaction of Wolbachia with the endoplasmic reticulum (ER). Through fluorescence time-lapse confocal and electron microscopy observations, I provide strong evidence that this organelle is the source of membrane for Wolbachia, and possibly a source of nutrients. However, gene expression analyses and immunofluorescence approaches demonstrate that Wolbachia do not induce ER stress nor an increased ERAD- induced proteolysis, suggesting; unlike previously reported, that Wolbachia salvage amino acids by other subversion mechanisms. Additionally, I pioneered biolistic bombardement of Wolbachia-infected cells and the validation of this transformation technique has paved the way towards optimization of transformant selection steps and ultimately to the genetic engineering of Wolbachia.

Au cours de ce travail, nous nous sommes intéressés à l'impact de l'exposition à la ciclosporine A (CsA) sur la signalisation angiogénique dans le carcinome à cellules rénales (renal cell carcinoma – RCC) qui représente la deuxième cause de cancer chez les patients transplantés rénaux. Nous avons examiné in vitro l'impact de l'exposition à la CsA sur la réponse UPR (Unfolded Protein Response) et la régulation des protéines sécrétées en portant un intérêt particulier à la régulation du VEGF (Vascular Endothelial Growth factor). Nous confirmons l'effet de la CsA sur la protéostase et montrons que l'activation de l'UPR par la CsA, conduisant à une augmentation de la production de VEGF en hypoxie, pourrait participer à l'agressivité des tumeurs. Nous proposons de rechercher certains biomarqueurs de l'UPR chez les patients ayant développé un RCC post-transplantation afin d'examiner de façon plus approfondie l'altération de la protéostase et la régulation de l'UPR dans ce contexte. Dans le domaine de la médecine personnalisée, d'autres approches comme la pharmacogénétique sont désormais utilisées dans la pratique médicale. Dans ce contexte, nous avons évalué l'intérêt du génotypage du VEGF dans une cohorte de patients transplantés rénaux. Nous montrons que le polymorphisme VEGF 936 C>T est associé de façon significative au risque de développer un RCC post-transplantation. Même s'il est évident que des études supplémentaires doivent être menées, nos résultats indiquent que le génotypage de VEGF 936 C>T pourrait être envisagé pour améliorer la gestion des traitements immunosuppresseurs chez les patients identifiés comme étant à risque de développer un RCC post-transplantation / In this work, we investigated the impact of cyclosporine A (CsA) exposure on angiogenic signalling in renal cell carcinoma (RCC), the second mostly observed cancer in renal transplanted patients. We examined in vitro the effect of CsA exposure on the Unfolded Protein Response (UPR) and the regulation of secreted proteins with a focus on VEGF (Vascular Endothelial Growth Factor) regulation. We confirm the effect of CsA on proteostasis and we show that the activation of UPR by CsA, leading to an increased VEGF hypoxic expression, could contribute to the aggressiveness of tumours. We propose to investigate a list of candidate UPR biomarkers in patients who have developed a post-transplant RCC in order to confirm the alteration of proteostasis and the UPR activation in this context. In the field of personalized medicine, other approaches such as pharmacogenetics are now used in medical practice. In this context, we evaluated the VEGF genotyping in a cohort of renal transplanted patients. We show that VEGF 936 C>T is significantly associated with the risk of developing a post-transplant RCC. Although it is evident that additional studies need to be conducted, our results indicate that VEGF 936 C>T genotypes might be useful to classify patients according to their post-transplant RCC risk in order to improve immunosuppressive drugs management

MCDS is an autosomal dominant disorder, with a mild dwarfed phenotype and is caused by mutations in collagen X. The majority of the mutations identified so far are localized almost exclusively within the NC1 domain, which is responsible for trimerization of the collagen X protein. Little is known about the onset of MCDS, but recently, up-regulation of ER stress has been suggested as an important mechanism promoting the MCDS phenotype. Several studies have shown that the mutated collagen X protein is retained within the ER triggering the UPR, which has proved to be the key pathway responsible for the pathogenesis of the MCDS phenotype. In order to study the consequences of the expressing the MCDS-causing COL10A1p.N617K mutation at the molecular level, we selected HeLa cells as an appropriate cell line for the characterisation of the UPR response, by showing that the three branches of the UPR can be activated by ER stress inducing conditions in a similar manner to that seen in vivo in the MCDS growth plate. Importantly we have also shown that HeLa cells can be transduced with the collagen X cDNA constructs and will express, fold and secrete collagen X into the supernatant.Having established the cellular model for MCDS studies we demonstrated for the first time direct evidence for the retention of mutant collagen X within the ER. Moreover, we demonstrated that the mutant collagen X was degraded via a proteasomal pathway. Nevertheless, the level of ER stress induced by expression of mutant collagen X, based on BiP induction at the protein level, was disappointingly low. We therefore directly compared the level of ER stress induced by the COL10A1p.N617K mutation with that of the chondrodysplasias-causing MATN3p.V194D mutation. The ER stress induced by the matrillin mutation was far greater than that caused by the mutant collagen X. We showed that general protein synthesis was reduced in cells expressing either of the mutant proteins, most likely by the mechanism associated with the phosphorylation of eIF2alpha. Moreover, we showed the mutant matrilin-3 protein was also retained specifically in the ER. However, we could find no evidence for either proteasomal or autophagic/lysosomal degradation of mutant matrilin 3.We tested a broad range of ER stress-relieving compounds on cells expressing mutant collagen X and matrilin 3. Carbamazepine, which was previously shown to reduce ER stress in alpha1-antitripsin deficiency, reduced ER stress in cells expressing the mutant collagen X (but not matrilin 3) by way of enhanced proteasomal degradation of the retained protein. This drug should now be tested in vivo against the MCDS mouse to determine its capacity to reduce disease severity.The results presented within this thesis have contributed to the understanding of how cells deal with mutant collagen X and matrilin-3 proteins. We have identified a potential therapeutic compound that may be of use in the treatment of MCDS. Furthermore, the data presented support the concept that generic approaches to relieving ER stress may not be suitable for treating a broad range of diseases. Treatments may need to be tailored not only in a gene-specific manner but also may need to be tailored to address the differing consequences of different mutations in the same gene.

Osteosarcoma (OS) is the most common primary malignant bone tumour in humans and dogs. Although medicine has made dramatic progress in treating osteosarcoma by surgery, with chemotherapy given before and after surgery, drug resistance and highly metastatic spread are often responsible for the failure of current therapies. Thus, more effective therapeutic approaches for treating osteosarcoma are needed. Previous results from our laboratory and others had shown that the basic-leucine zipper (bLZip) containing transcription factor, Zhangfei/CREBZF is a potent inhibitor of a variety of other transcription factors and has a dramatic effect on the growth of several cancer cell lines, including dog OS and human medulloblastoma cells. The objective of the studies described in this thesis was to determine the molecular mechanisms by which Zhangfei exerts its effect on dog and human OS cells. Several stressors in the microenvironment of cancer cells directly or indirectly perturb the endoplasmic reticulum (ER), which then activates the Unfolded Protein Response (UPR). The UPR modulates the effects of stress and allows tumours to survive, develop, metastasize and escape therapy. The UPR is regulated by three bLZip transcription factors—ATF6, ATF4 and Xbp1s. Since Zhangfei inhibits Luman/CREB3, a bLZip structurally similar to and closely related to ATF6 and ATF4, I initially focused my efforts on this pathway. I hypothesized that Zhangfei interacts with UPR-related bLZip transcription factors and inhibits their ability to activate the UPR signaling pathways, thereby suppressing the growth of cancer cells and increasing their susceptibility to ER stressors. To test this hypothesis, we monitored cell growth as well as levels of UPR gene transcripts and proteins in several dog and human osteosarcoma cell lines infected with adenovirus vectors expressing Zhangfei, and studied the interactions between Zhangfei and the UPR-mediator, Xbp1s. The results showed that the ectopic expression of Zhangfei in cell lines derived from dog osteosarcomas potently suppressed cell growth and inhibited their ability to activate the UPR. Further studies demonstrated that Zhangfei inhibited the UPR, at least partially, by binding to Xbp1s and suppressing its ability to activate transcription from a promoter containing unfolded protein response elements (UPRE). The leucine zipper of Zhangfei was required for this interaction, which led to the subsequent proteasomal degradation of Xbp1s. However, we also found that the effects of Zhangfei were not universal. While Zhangfei had a profound effect on the growth and UPR in some OS cell lines, it either had only a partial effect, or no effect on others. This suggested that susceptibility (or resistance) to Zhangfei may be an inherent property of OS cell lines. Since the suppressive effects of Zhangfei were not universal, and it had no obvious effects on untransformed cells and some cancer cell lines, I proposed that Zhangfei mediates its effect on cell growth and the UPR through an intermediary that is either not induced or is defective in cells that are unaffected by Zhangfei. I found that this intermediary was the tumour suppressor protein p53. The inhibitory effects of Zhangfei were only observed in the wild-type p53 expressing OS cell line U2OS while Zhangfei had no effect on the p53-null OS cell line MG63. In cells with functional p53, the ectopic expression of Zhangfei caused it to displace the ubiquitin ligase mdm2 and stabilize p53. Suppression of p53 by siRNA partially inhibited the effects of Zhangfei on the UPR and cell growth. In contrast, OS cells lacking functional p53 could be made to respond to Zhangfei if they were transfected to express wild-type p53. These results explain why Zhangfei has a profound effect on some cancer cells while having no obvious effect on others. I also characterized the interaction of Zhangfei and p53 by mapping the interacting domains on both proteins, showing that the bLZip domain of Zhangfei and the N-terminal transactivation domain (NTD) of p53 were required for their interactions. My findings reveal the profoundly inhibitory effects of Zhangfei on OS growth and the UPR, a stress-response known to promote tumour survival. I also show how Zhangfei may exert its effects. My work suggests an alternative modality for the therapy of certain types of OS, and perhaps other tumours with functional p53.

Doctor of Philosophy / Biochemistry and Molecular Biophysics Interdepartmental Program / Gerald R. Reeck / Armet is a bifunctional protein that is apparently universally distributed among multicellular animal species, vertebrate and invertebrate alike. A member of the Unfolded Protein Response, (UPR) Armet promotes survival in cells that are under endoplasmic-reticulum (ER) stress. I have carried out biophysical studies on human Armet looking for compounds that bind to Armet and hence could reduce its anti-apoptotic function, thus potentially joining the growing class of pro-apoptotic drugs. Performed primarily with 1H-15N HSQC NMR, ligand studies showed that approximately 60 of the 158 residues are potentially involved with binding. The 60 residues are distributed throughout both domains and the linker suggesting multi-domain interaction with the ligand. Circular dichroism studies showed heat denaturation in a two-step unfolding process with independent unfolding of both domains of Armet with Tm values near 68°C and 83 C with the C-terminal domain unfolding first, as verified by 1H-15N HSQC NMR measurements. I also provide the first identification of UPR transcripts in pea aphids, Acyrthosiphon pisum, the genetic model among aphids. I measured transcript abundance with hope of finding future transcriptional targets for pest mitigation. I identified 74 putative pea aphid UPR components, and all but three of the components have higher transcript levels in aphids feeding on plants than those that fed on diets. This activated UPR state is attributed to the need for saliva proteins for plant feeding. Because aphids are agriculturally significant pests, and saliva is pivotal to their feeding on host plants, genes that encode saliva proteins may be targets for pest mitigation. Here I have sought the aphid’s saliva proteome by combining results obtained in several laboratories by proteomic and transcriptomic approaches on several aphid species. With these data I constructed a tentative saliva proteome for the pea aphid by compiling, collating, and annotating the data from several laboratories. I used RNA-seq to verify the transcripts in pea aphid salivary glands, thus expanding the proposed saliva proteome from approximately 50 components to around 130 components, I found that transcripts of saliva proteins are upregulated during plant feeding compared to diet feeding.

Indiana University-Purdue University Indianapolis (IUPUI) / The unfolded protein response (UPR) is important for cell adaptation to accumulation of unfolded proteins in the endoplasmic reticulum (ER). A central UPR sensor of ER stress is PKR- like ER Kinase (PERK), which phosphorylates eIF2 to reduce global translation and help mitigate ER stress. While this is a survival mechanism that serves to save the cell from catastrophic events during ER stress, PERK can also be activated in cancer cells due to genetic changes and exposure to stresses inherent in the tumor micro-environment. Published reports have indicated that PERK is activated in cancer cells in response to hypoxia, nutrient deprivation, matrix detachment, and increased protein load by oncogene activation to facilitate cell survival. The UPR features PERK and another ER stress sensory protein, IRE1α, which also regulates the dynamic assembly of the actin cytoskeleton; loss of either PERK or IRE1α functions decrease cell migration activity. We hypothesized that PERK is required for anchorage-independent survival of the cancer cell line HCT116 and that PERK is essential for cell migration. Consistent with these premises, inhibition of PERK using pharmacological inhibitors GSK2656157 and LY-4 in suspended cells showed reduced growth. Furthermore, PERK-deficient cells showed reduced migration in transwell migration assays as compared to their wild type counterpart. These results suggest that PERK facilitates anchorage-independent growth of cancer cells and cell migration.

Le réticulum endoplasmique (RE) joue un rôle majeur dans la conformation des protéines. L’accumulation de protéines non- ou mal-conformées dans le RE induit un stress qui peut être résolu par la réponse aux protéines mal-conformées (UPR). Un stress chronique du RE entraine une apoptose dépendante de l’UPR et se traduit par un déséquilibre de l’homéostasie tissulaire. Bien que l’apoptose dépendante d’un stress du RE soit observée et à l’origine d’un grand nombre de maladies humaines, les mécanismes pro-apoptotiques ainsi que ceux favorisant l’homéostasie tissulaire en réponse à un stress chronique du RE restent à ce jour méconnus. Cette thèse apporte une meilleure compréhension de ces mécanismes grâce à un nouveau modèle d’induction de stress du RE chez la drosophile basé sur la surexpression de la préséniline. L’apoptose observée dans ce modèle dépend d’une répression au moins transcriptionnelle du gène anti-apoptotique diap1 par la branche PERK/ATF4 de l’UPR, alors que les voies pro-apoptotiques classiquement impliquées dans l’apoptose en réponse à un stress du RE chez les mammifères ne semblent pas être impliquées. Par ailleurs, la branche PERK/ATF4 active la voie JNK par l’intermédiaire de la petite GTPase Rac1 et de la MAP3K Slipper qui sont activées dans les cellules apoptotiques. Cette activation aboutit à l’expression de Dilp8, un peptide ressemblant à l’insuline qui cause un retard de développement et permet ainsi de remplacer partiellement les cellules éliminées par apoptose. Dans notre modèle, les mécanismes classiquement décrits dans le maintien de l’homéostasie tissulaire chez la drosophile tels que la prolifération compensatoire ou la réparation des tissus ne semblent pas avoir de rôle majeur. Ces résultats établissent, une nouvelle voie qui participe à l’homéostasie tissulaire dans un nouveau modèle de stress chronique du RE / The Endoplasmic Reticulum (ER) plays a major role in protein folding. The accumulation of unfolded proteins in the ER induces a stress which can be resolved by the Unfolded Protein Response (UPR). The chronicity of ER-stress leads to UPR-induced apoptosis and in turn to an unbalance of tissue homeostasis. Although ER stress-dependent apoptosis is observed in a great number of devastating human diseases, how cells activate apoptosis and promote tissue homeostasis after chronic ER-stress remains poorly understood. During my thesis we have established of a novel model of chronic ER-stress using the Drosophila wing imaginal disc as a model system. We have validated that Presenilin (Psn) overexpression induces chronic ER-stress in Drosophila associated to a PERK/ATF4-dependent apoptosis requiring the down-regulation of the anti-apoptotic diap1 gene. Interestingly, the classical pro-apoptotic pathways described in mammals do not seem implicated in Psn-overexpression-dependent apoptosis. PERK/ATF4 also activated the JNK pathway through the small GTPase Rac1 and the MAP3K Slipper activation in apoptotic cells, leading to the expression of Dilp8. This insulin-like peptide caused a developmental delay, which partially allowed the replacement of apoptotic cells. The other mechanisms involved in tissue homeostasis in Drosophila, i.e. compensatory homeostasis and wound healing, do not seem to have a major role in our model. These results establish a new pathway that participates in tissue homeostasis thanks to a novel chronic Drosophila ER stress model

Les signalisations oncogéniques induisent une consommation accrue de glucose qui n'est que partiellement satisfaite par le microenvironnement. Pour s'adapter et survivre à ce stress métabolique, les cellules malignes mettent en jeu des mécanismes qui restent mal compris. Nos travaux montrent que cette limitation en glucose a pour principale conséquence de déclencher une apoptose via la voie de signalisation PERK-CHOP de la réponse à un stress du réticulum endoplasmique (SRE), nommée Unfolded Protein Response (UPR). Nous avons découvert que le RE est capable de sentir la carence en glucose via la diminution de la disponibilité en UDP Nacétylglucosamine produit par la voie des hexosamines. La délétion du facteur pro-Apoptotique CHOP dans un modèle de cancer spontané du poumon induit par KrasG12V chez la souris augmente l'incidence tumorale, confirmant que le SRE constitue un mécanisme cellulaire de sauvegarde anti-Tumoral. Nous montrons également que le franchissement de cette barrière implique l'atténuation sélective de la voie PERK-CHOP par la protéine chaperon p58IPK, qui permet aux cellules de bénéficier en retour des effets protecteurs des autres voies d'un UPR devenu chronique. Ces résultats révèlent une dualité fonctionnelle pour le stress du RE dans la tumorigenèse contrôlée, au moins pour partie, par la protéine p58IPK / During carcinogenesis, oncogene activation induces high glucose avidity that outstrips the microenvironment supply until angiogenesis occurs. How malignant cells cope with this potentially lethal metabolic stress remains poorly understood. We found that oncogene-Driven glucose shortage triggers apoptosis through the PERK-CHOP pathway of the endoplasmic reticulum (ER) unfolded protein response (UPR). Deletion of the pro-Apoptotic UPR effector CHOP in a mouse model of KrasG12V induced lung cancer increases tumour incidence, strongly supporting the notion that ER stress serves as a barrier to malignancy. Overcoming this barrier requires the selective attenuation of the PERK-CHOP arm of the UPR by the molecular chaperone p58IPK. Furthermore, p58IPK-Mediated adaptive response enables cells to benefit from the protective features of chronic UPR. Altogether, these results show that ER stress activation and p58IPK expression control the fate of malignant cells facing glucose shortage

Le réticulum endoplasmique rugueux (RE), assure la synthèse, le repliement et la maturation des protéines de la voie de sécrétion. Les altérations des fonctions physiologiques du RE, entrainent l’accumulation de protéines mal repliées dans la lumière du RE, une condition appelée stress RE. En réponse au stress RE, un mécanisme compensatoire adaptatif appelé Unfolded Protein Response (UPR) est activé afin de restaurer l’homéostasie du RE et de permettre la survie de la cellule. Dans le cas d’un stress RE sévère ou prolongé, les altérations ne pouvant plus être compensées, la cellule est éliminée par apoptose contribuant ainsi au développement de pathologies cardiaques. Le but des recherches actuelles sur le stress RE en physiopathologie cardiaque n’est pas d’inhiber la réponse au stress RE, mais plutôt de la moduler afin de limiter l’apoptose des cardiomyocytes et de protéger le cœur. Dans ce contexte, nous avons mis en évidence que le stress RE induit une modification importante de l’architecture des cardiomyocytes associée à une altération de la fonction mitochondriale. De plus, nous avons montré que SIRT1, une désacétylase dépendante du NAD+, inhibe l’apoptose mitochondriale induite par un stress RE en limitant spécifiquement l’activation de la voie PERK de la réponse UPR via la désacétylation du facteur eIF2á sur la lysine K143. Enfin, nos résultats indiquent que SIRT1 protège les cardiomyocytes de l’apoptose induite par le stress RE en favorisant la mitophagie, via une activation de la voie de signalisation eEF2K/eEF2. Ces résultats montrent que SIRT1 est impliquée dans la régulation de la réponse autophagique et apoptotique des cardiomyocytes au stress RE et suggèrent que cette désacétylase serait une cible thérapeutique intéressante pour limiter le développement des pathologies cardiaques liées au stress RE. / The endoplasmic reticulum (ER) functions to properly synthesize, fold and process secreted and transmembrane proteins. Impairment of ER function induces an accumulation of misfolded proteins in the ER lumen, a condition termed ER stress. In response to ER stress, an adaptive compensatory mechanism called Unfolded Protein Response (UPR) is activated to restore ER homeostasis and promote cell survival. In the case of severe or prolonged ER stress, homeostasis cannot be restored and the cell is eliminated by apoptosis contributing to the development of cardiac pathologies. Currently, cardiac therapy based on ER stress modulation to conserve beneficial adaptations and to avoid cardiomyocyte apoptosis is viewed as a promising avenue towards effective therapies of ER stress-associated cardiac diseases.In this context, we demonstrated that ER stress induces architectural modifications and alterations of the mitochondrial function in cardiomyocytes. Furthermore, we showed that SIRT1, a NAD+-dependent deacetylase, inhibits mitochondrial apoptosis by modulating the activation of the PERK pathway of the UPR through deacetylation of the translation initiation factor eIF2á on lysine K143. Our results also indicate that SIRT1 protects cardiomyocyte from ER stress-induced apoptosis by activating mitophagy through eEF2K/eEF2 pathway. Collectively, these data demonstrate that SIRT1 regulates ER stress-induced autophagy and apoptosis in the heart and suggest that this deacetylase may be a therapeutic target to protect the heart against ER stress-induced injury.