Obesity and Type 2 Diabetes are now considered as worldwide epidemics. To prevent and/or to cure these pathologies, the regulatory mechanisms of biological processes involved in their development need to be characterized. Among these processes, endogenous glucose production (EGP) is a crucial function for glucose homeostasis. Indeed, increased hepatic glucose production (HGP) has been associated with the settle of type 2 diabetes. On the contrary, intestinal glucose production (IGP) induces beneficial effects in energy and glucose homeostasis. Indeed, in the portal vein, the sensing of glucose produced by the intestine induces a signal to the brain. The portal glucose signal produced by IGP protects against the development of obesity and type 2 diabetes by decreasing food intake and HGP and by activating energy expenditure. Tissue-specific regulations of both HGP and IGP are thus crucial processes to decipher. EGP is restricted to the liver, kidney and intestine since these organs are the only organs expressing the key enzyme glucose-6-phosphatase (G6Pase). Glucose-6 phosphatase catalyzes the production of glucose, which is then exported out of the cell by the facilitated transporter Glut2 and by a membrane-based pathway, which still needs to be fully deciphered. This project aims at characterizing this vesicular glucose transport pathway and demonstrating the role of G6Pase in such a glucose transport process. I will first identify the components of the vesicular pathway of glucose export in the liver. Our preliminary data suggested that caveolin-1 (Cav1: a key component of caveolae), and dynamins (Dyn: necessary for the fission of vesicles) are involved in glucose export. Based on in vivo experiments performed in transgenic mouse models and in vitro experiments performed in hepatocytes, I will further characterize how these proteins participate in glucose transport. The importance of the vesicular glucose transport pathway compared with the glucose transport by Glut2 will be then assessed. We recently demonstrated that G6Pase activity is regulated by a membrane-based mechanism. I will then question the involvement of G6Pase in this glucose export pathway. Particularly, I will analyze the interaction of G6Pase with the components of the vesicular glucose transport pathway identified previously. I will then specify whether the vesicular glucose transport pathway is deregulated in the context of type 2 diabetes. Finally, I will extend this study to other gluconeogenic organs (i.e the kidney and intestine). In addition to molecular studies, I will focus on intestinal glucose absorption to firmly demonstrate that G6Pase may directly participate to a glucose transport function. G6Pase might participate in intestinal glucose absorption in the absence of Glut2. I will thus assess whether the suppression of G6Pase specifically in the intestine (I.G6pc-/-) decreases intestinal glucose absorption. Thus, my project aims at identifying a new function of G6Pase linking its capacity of glucose production to vesicular glucose export. These results may possibly lead to revisit the paradigms in glucose homeostasis. In addition, deciphering the regulations of this new function, and particularly tissue-specific regulations, should provide the bases for original strategies targeting EGP to prevent and/or cure type 2 diabetes.