The bioluminescence resonance energy transfer (BRET) approach involves resonance energy transfer between a light-emitting enzyme and fluorescent acceptors. The major advantage of this technique over biochemical methods is that protein-protein interactions (PPI) can be monitored without disrupting the natural environment, frequently altered by detergents and membrane preparations. Thus, it is considered as one of the most versatile technique for studying molecular interactions in living cells at “physiological” expression levels. BRET analysis has been applied to study many transmembrane receptor classes including G-protein coupled receptors (GPCR). It is well established that these receptors may function as dimeric/oligomeric forms and interact with multiple effectors to transduce the signal. Therefore, they are considered as attractive targets to identify PPI modulators. In this review, we present an overview of the different BRET systems developed up to now and their relevance to identify inhibitors/modulators of protein–protein interaction. Then, we introduce the different classes of agents that have been recently developed to target PPI, and provide some examples illustrating the use of BRET-based assays to identify and characterize innovative PPI modulators in the field of GPCRs biology. Finally, we discuss the main advantages and the limits of BRET approach to characterize PPI modulators.

Serotonin (5-HT) is known as a potent immune cell modulator in autoimmune diseases and should be protective in the pathogenesis of multiple sclerosis (MS). Nevertheless, there is limited knowledge about receptors involved in 5-HT effects as well as induced mechanisms. Among 5-HT receptors, the 5-HT₇ receptor is able to activate naïve T cells and influence the inflammatory response; however, its involvement in the disease has never been studied so far. In this study, we collected blood sample from three groups: acute relapsing MS patients (ARMS), natalizumab-treated MS patients (NTZ), and control subjects. We investigated the 5-HT₇ expression on circulating lymphocytes and evaluated the effects of its activation on cytokine production with peripheral blood mononuclear cell (PBMC) cultures. We found a significant increase in the 5-HT₇ surface expression on T lymphocytes and on the different CD4⁺ T cell subsets exclusively in NTZ-treated patients. We also showed that the selective agonist 5-carboxamidotryptamine (5-CT)-induced 5-HT₇R activation significantly promotes the production of IL-10, a potent immunosuppressive cytokine in PBMCs. This study provides for the first time a dysregulation of 5-HT₇ expression in NTZ-MS patients and its ability to promote IL-10 release, suggesting its protective role. These findings strengthen the evidence that 5-HT₇ may play a role in the immuno-protective mechanisms of NTZ in MS disease and could be considered as an interesting therapeutic target in MS.

BackgroundInterleukin (IL)-33 is expressed in healthy brain and plays a pivotal role in several neuropathologies, as protective or contributing to the development of cerebral diseases associated with cognitive impairments. However, the role of IL-33 in the brain is poorly understood, raising the question of its involvement in immunoregulatory mechanisms.MethodsWe induced a local IL-33 release by intra-hippocampal injection of recombinant mouse IL-33 (rmIL-33) into C57BL/6J (WT) and IL-1ab deficient mice. Chronic minocycline administration was done and cognitive functions were examined trough short- and long-term memory assays. Hippocampal inflammatory responses were investigated by RT-qPCR. The microglia activation was assessed using immunohistological staining and Fluorescence-activated cell sorting. ResultsWe showed that IL-33 administration in mice led to a long-term memory defect associated with an increase of inflammatory markers in hippocampus while, minocycline administration limited the inflammatory response. Quantitative assessment of glial cell activation in situ was performed and demonstrated an increase of proximal intersections per radius in each part of hippocampus. Moreover, rmIL-33 significantly promoted the outgrowth of microglial processes. Fluorescence-activated cell sorting performed on isolated microglia, revealed an overexpression of IL-1β, 48h post-rmIL-33 administration. This microglial reactivity was closely related to the onset of cognitive disturbance. Finally, we demonstrated that IL-1ab deficient mice were resistant to cognitive disorders after intra-hippocampal IL-33 injection. ConclusionThus, hippocampal IL-33 induced an inflammatory state, including IL-1β overexpression by microglia cells, being causative of the cognitive impairment. These results highlight the pathological role for IL-33 in the central nervous system, independently of a specific neuropathological model.