Intra-cellular VHH targeting the FSHR and LHR
Résumé
Intracellular variable fragments from heavy-chain antibody from camelids (intra-VHH) have been successfully used as chaperones to solve the 3D structure of active G protein-coupled receptors (GPCRs) bound to their cognate transducers. They may also provide a better understanding on the relationships between receptor conformation and signalling activity, but so far, this property has been explored only for four GPCRs, all binding peptide ligands. We report the isolation and characterization of iPRC1, the first intra-VHH recognizing the receptors of follicle-stimulating hormone (FSHR) and of luteinizing hormone/ choriogonadotropin (LHCGR). This intra-VHH recognizes the 3rd intracellular loop (ICL3) of the receptors and decreases the cAMP and steroid production upon hormone-binding. In addition, using a next generation sequencing approach after phage display selection on the FSH-activated FSHR, a specific anti-FSHR intra-VHH, iPRC2, was isolated. Like iPRC1, iPRC2 decreases the cAMP production in response to FSH, and in addition, its presence increases the FSHR accumulation in the early endosomes, while decreasing its recycling rate. Therefore, iPRC2 has the ability to reroute the intracellular trafficking of a GPCR. Altogether, these intra-VHHs provide new tools to help completing structure/activity studies performed so far on this receptor.