THE PROTEOSTASIS OF THE ENDOPLASMIC RETICULUM AND THE ACTIVATION OF THE UNFOLDED PROTEIN RESPONSE PATHWAY IN VIVO

ABSTRACT Background. The protein components of eukaryotic cells face acute and chronic challenges to normal folding, refolding and function owing to a constant barrage of physical, metabolic and environmental stresses. Eukaryotic protein homeostasis, or proteostasis, enables healthy cell adaptation during development and protects against aging and diseases. Proteostasis refers to controlling the concentration, conformation, binding interaction and location of individual proteins making up the proteome by readapting the innate biology of the cell, often through transcriptional and translational changes. The endoplasmic reticulum (ER) responds to the accumulation of unfolded proteins in its lumen (ER stress) by activating intracellular signal transduction pathways, cumulatively called the unfolded protein response (UPR). The UPR activation triggers an extensive transcriptional and translational response, which adjusts the ER protein folding capacity according to needs. As such, the UPR constitutes one of the signaling pathways that regulates the capacity and composition of the proteostasis network according to the changing of the ER folding capacity. Previous work. My initial work, described in chapter 1.2.1 shows that the level of cellular energy is important for protein folding and disaggregation and thus can affect folding and repair processes. The proteostasis network is not only highly adaptable, enabled by the influence of multiple cell stress signaling pathways, but also can be quite distinct in each cell type. In chapter 1.2.2, by using as a cellular model the plasma cells differentiation, I could underline the role of the UPR signaling and proteasomal degradation in orchestrating the architectural and functional changes of the cells and balancing the proteostasis capacity. Recent studies suggest that lack/decrease of O2 perturbs the ER homeostasis. Indeed the ER has emerged as a cellular compartment that depends on O2 for oxidative folding of secretory and transmembrane proteins and that mediates the O2 signaling that is important for the survival and function of hypoxic cells. In vitro studies have shown that hypoxia triggers the UPR. However, in vivo and in vitro situations are indeed likely to diverge substantially with respect to parameters such as metabolic activity, O2 utilization and cell division rates, features that are predicted to vary the cell sensitivity to ER stress. Thus, despite a recognized role for hypoxia on UPR, few data exist on the effects of hypoxia in various organs in vivo. Aims. My study includes three aims: First, we will test if the hypoxic stress in vivo acts as a modifier that affects the activation of specific branches of the UPR in different tissues. Second, to get a better insight into the role of the UPR during low oxygen availability in tissues, we will test whether the UPR activation depends from the severity of the hypoxic stress. Third, we aim at delineating signaling circuits that control the capacity and composition of the proteostasis network through transcriptional and post transcriptional mechanisms to balance the ER homeostasis Results. I analyzed the effect of the hypoxic stress on the proteostasis network. Changes in O2 levels alter the ability of the cells to handle the proteostasis load with some differences between the cells type studied. Hepatocytes and myocytes respond to hypoxia by increasing their degradation activity as to increase the proteostasis capacity. While the hepatocytes activate an UPR-dependent apoptosis and are able to balance between apoptotic death and protein synthesis, in the myocytes the protein synthesis remains sustained under low oxygen availability while the UPR –dependent apoptosis could not be detected. Conclusion. This studies underlined several features of the ER- proteostasis. First, the proteostasis network is adaptable and able to fine tune the UPR signaling pathway in response to stress. Second, different cells have varying proteostasis capacities reflected in the composition and concentrations of their proteostasis components. Third, within a given cell type, the proteostasis does not possess significant excess capability, rater it is finely tuned and offers just enough facility for the protein folding load. Therefore, by setting the proteostasis boundary as a threshold for generating folded and functional proteins, the proteostasis network can create and maintain functional proteins in response to the local environment.