Know more

Our use of cookies

Cookies are a set of data stored on a user’s device when the user browses a web site. The data is in a file containing an ID number, the name of the server which deposited it and, in some cases, an expiry date. We use cookies to record information about your visit, language of preference, and other parameters on the site in order to optimise your next visit and make the site even more useful to you.

To improve your experience, we use cookies to store certain browsing information and provide secure navigation, and to collect statistics with a view to improve the site’s features. For a complete list of the cookies we use, download “Ghostery”, a free plug-in for browsers which can detect, and, in some cases, block cookies.

Ghostery is available here for free:

You can also visit the CNIL web site for instructions on how to configure your browser to manage cookie storage on your device.

In the case of third-party advertising cookies, you can also visit the following site:, offered by digital advertising professionals within the European Digital Advertising Alliance (EDAA). From the site, you can deny or accept the cookies used by advertising professionals who are members.

It is also possible to block certain third-party cookies directly via publishers:

Cookie type

Means of blocking

Analytical and performance cookies

Google Analytics

Targeted advertising cookies


The following types of cookies may be used on our websites:

Mandatory cookies

Functional cookies

Social media and advertising cookies

These cookies are needed to ensure the proper functioning of the site and cannot be disabled. They help ensure a secure connection and the basic availability of our website.

These cookies allow us to analyse site use in order to measure and optimise performance. They allow us to store your sign-in information and display the different components of our website in a more coherent way.

These cookies are used by advertising agencies such as Google and by social media sites such as LinkedIn and Facebook. Among other things, they allow pages to be shared on social media, the posting of comments, and the publication (on our site or elsewhere) of ads that reflect your centres of interest.

Our EZPublish content management system (CMS) uses CAS and PHP session cookies and the New Relic cookie for monitoring purposes (IP, response times).

These cookies are deleted at the end of the browsing session (when you log off or close your browser window)

Our EZPublish content management system (CMS) uses the XiTi cookie to measure traffic. Our service provider is AT Internet. This company stores data (IPs, date and time of access, length of the visit and pages viewed) for six months.

Our EZPublish content management system (CMS) does not use this type of cookie.

For more information about the cookies we use, contact INRA’s Data Protection Officer by email at or by post at:

24, chemin de Borde Rouge –Auzeville – CS52627
31326 Castanet Tolosan CEDEX - France

Dernière mise à jour : Mai 2018

Menu Logo Principal logo Université Clermont Auvergne & associés

Human Nutrition Unit

Zone de texte éditable et éditée et rééditée

CARE-LUC mouse model

An innovative tool to monitor the Integrated Stress Response pathway in vivo

The Integrated Stress Response

Mammalian cells have evolved complex stress signaling pathways that are activated in response to a range of physiological changes and different pathological situations. These pathways initiate a wide array of adaptative mechanisms and ultimately, if necessary, programmed cell death. Such stresses commonly include cell extrinsic factors such as UV irradiation, hypoxia, viral infection and deprivation of nutrients and also intrinsic stresses such as endoplasmic reticulum (ER) stress. Reversible phosphorylation on serine 51 of the α subunit of eukaryotic translation initiation factor 2 (eIF2α) is a highly conserved regulatory event activated in response to diverse stresses. In mammals, four protein kinases are known to couple distinct upstream stress signals to eIF2α phosphorylation.


GCN2 kinase (General Control Non-depressible-2) is activated in response to amino acid deprivation by binding to uncharged tRNAs via a histidyl-tRNA synthetase related domain and the PERK kinase (PKR-like ER kinase) by the accumulation of unfolded proteins leading to the endoplasmic reticulum stress (ER) (Figure 1). Regarding the PKR kinase (Protein Kinase double-stranded RNA-dependent), it is activated by double-stranded RNA (dsRNA) during viral infections and by several cytokines (interferon g, TNFα and IL-1β). Finally, HRI kinase (Heme Regulated Inhibitor) is mainly involved in erythrocyte differentiation during erythropoiesis and in cellular responses to intracellular bacterial pathogens. The phosphorylation of eIF2α elicits a severe decline in de novo protein synthesis as its primary consequence and is an important strategy in the cell’s armory against a large panel of stressful insults. Accompanying this global translational control, it also favors increased translation of a selected number of mRNAs containing short upstream open reading frames.

One of the proteins for which translation is increased is activating transcription factor 4 (ATF4), a member of the ATF subfamily of the basic leucine zipper (bZIP) transcription factor superfamily. ATF4 is a master regulator that triggers increased transcription by binding to C/EBP-ATF Response Element (CARE) sequences of a subset of specific target genes involved in metabolism, nutrient uptake, control of redox status, and regulation of apoptosis and autophagy. The idea that ATF4 is a common downstream target that integrates signaling from multiple eIF2α kinases has led to the eIF2α-ATF4 pathway being referred to as the integrated stress response (ISR). Importantly, the ISR has been shown to be involved in a number of human stress-related pathologies including cancer or neurodegenerative disorders.

 The CARE-LUC mouse model

To visualize the activation of the ISR signaling pathway in response to stress at the whole animal, we generated a transgenic mouse line called CARE-LUC expressing the luciferase reporter gene (LUC) under control of CARE sequences able to bind ATF4 (Figure 2A). Luciferase expressed in the different tissues catalyzes the oxidation of luciferin injected into the animal. The light emitted in this reaction is proportional to the luciferase gene expression level. Its detection allows to visualize the activation of the ISR pathway according to stresses, physiopathological or pathological conditions in (i) the living animal by bioluminescence imaging, (ii) each organ or tissue by bioluminescence imaging and enzymatic assay and (iii) each cell by immunohistochemistry using an anti-luciferase antibody.

 The activation profile of the ISR pathway in tissues is specific for the inducer.


The CARE-LUC mouse model enabled to visualize the activation of the ISR pathway, through the kinase GCN2 in response to a nutritional stress resulting from the consumption of an essential amino acid-deficient diet such as lysine, or through the kinase PERK during ER stress consecutive of the injection of tunicamycin (Tu) and thapsigargin (Tha) (Figure 2B). The results of bioluminescence imaging in the mice abdomen confirm that the ISR pathway is activated by these stimuli. As expected, no activation is detected after consumption of a diet deficient in alanine, a non-essential amino acid that the body synthesizes. Subsequently, the analysis of luciferase activity in different tissues has enabled us to show that the activation profile of the ISR pathway will depend on the inducer (Figure 2C-D). Indeed, if the induction of this pathway is still observed in the liver, it is variable depending on the inducer in certain tissues such as kidney, brown adipose tissue (BAT) and muscle. We confirmed these results by measuring induction of expression of the ISR pathway target genes as Trb3, ATF3 or Xpot in these tissues. This difference in the activation profile cannot be explained by the presence or absence of GCN2 and PERK kinases that are both expressed in these tissues. Thus, the mechanisms governing tissue specificity of the induction of the ISR pathway remain to be identified.

 The ISR pathway is activated in a hepatic fibrosis model.

It is now established that the ISR pathway is activated in a large number of pathological conditions such as Alzheimer's disease, tumor growth, nephropathy or bacterial infection. However, the role of this pathway in the progression or the fight against these diseases needs to be better characterized. We conducted a first proof of concept experiment using the CARE-LUC mouse line to study the activation of ISR pathway in a pathological situation such as hepatic fibrosis. This type of fibrosis is a result of chronic liver injury, which is characterized by abnormally high accumulation of components of the extracellular matrix in the liver parenchyma. We have demonstrated a correlation between the appearance of an induced liver fibrosis and a chemical activation of the ISR pathway through the PERK kinase. Immunohistochemistry experiments have enabled to demonstrate the induction of both the luciferase and collagen-1 proteins in hepatocytes. Thus, these data confirmed the relevance of the CARE-LUC mouse model for investigating stress-related pathologies.

 Visualization of the activation of the ISR pathway in stressful situations leading to skeletal muscle atrophy.


A variety of stresses including starvation, muscle disuse, systemic illness and aging cause an alteration of skeletal muscle in a manner that promotes myofiber atrophy. The group of C. Adams has recently established that ATF4 plays a key role in age-related muscle weakness and atrophy and that this transcription factor is also required for skeletal muscle atrophy during starvation and immobilization. The mechanism by which ATF4 promotes muscle atrophy is complex because it regulates the expression of many genes. Three of those genes appear to be involved in reducing protein synthesis (4E-BP1) or in inducing skeletal muscle fiber atrophy (Gadd45a and p21). In skeletal muscle, upstream mechanisms that control ATF4 activity are not yet known and the role of eIF2α phosphorylation by specific eIF2α kinases remains to be demonstrated.

We sought to show the relevance of the reporter mouse model to study stress-related pathologies leading to skeletal muscle atrophy. CARE-LUC mice fasted for 48h show a significant increase of bioluminescence in their abdominal cavity (Figure 3A). To localize more precisely the origin of the luminescence, we excised and individually imaged skeletal muscles from control or fasted mice (Figure 3B). Tibialis and gastrocnemius displayed induction of bioluminescence in response to food deprivation. To confirm imaging data, we quantified luciferase activity in homogenates of muscle samples collected from control and fasted mice. In accordance with the results from noninvasive imaging, gastrocnemius and tibialis exhibited significant induction in luciferase activity under fasting conditions (Figure 3C). To confirm the correlation between skeletal muscle atrophy and the activation of the ISR pathway, we examined the mRNA level of several ATF4-dependent genes involved in reducing protein synthesis or in inducing skeletal muscle fiber atrophy and of the E3 ubiquitin ligase MuRF1, a well characterized atrophy associated transcript which is partially required for muscle atrophy. Figure 3D shows increased level of Gadd45a and 4E-BP1 and of MuRF1 under fasting conditions. Overall, these results highlight that the CARE-LUC mouse model represents an innovative tool for investigating the role of the ISR pathway in stress-related disorders leading to skeletal muscle atrophy.

For more information : Chaveroux et al, Science Signaling 2015.

ContactAlain BRUHAT