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Eneko Urizar"Sensing 7TM receptor function : biophysics of GPCR receptor activation".

Abstract :

A major obstacle to understanding the functional importance of dimerization between Class A G protein-coupled receptors (GPCRs) has been the methodological limitation in achieving control of the identity of the components comprising the signaling unit. We have developed physical and functional complementation assays that enable such control and illustrate it for the human dopamine D2 receptor (D2R). The minimal signaling unit, two receptors and a single G protein, is maximally activated by agonist binding to a single protomer, which suggests an asymmetrical activated dimer. Whereas inverse agonist binding to the second protomer enhances signaling, agonist binding to the second protomer blunts signaling. Ligand-independent constitutive activation of the second protomer also inhibits signaling. Thus, a given GPCR heterodimer function can be modulated by the activity state of the second protomer, which may be altered in pathological states. Our novel methodology also makes possible the characterization of signaling from a defined heterodimer, enabling the identification of modulators of heterodimer function. After validation of the system in vitro, we are currently translating the system in Drosophila melanogaster and use flies as an in vivo neuronal expression system to study both the assembly and the signaling of receptor homomers and heteromers. The novel imaging tools which we have available for dissecting D2R signaling will be broadly applicable to other GPCRs. Once we are capable of detecting ligand-induced FRET changes, reflecting conformational changes within the G protein heterotrimer, we will express the biosensors in primary cortical cultures and ultimately knock them into the mouse genome. Our larger goal is to develop broad-based methods for imaging the downstream signaling pathways, both G protein dependent and independent, for a variety of GPCRs.

Invité par Laurent Groc

Selected publications

Dopamine D2 receptors form higher order oligomers at physiological expression levels. Guo W, Urizar E, Kralikova M, Mobarec JC, Shi L, Filizola M, Javitch JA.
EMBO J. 2008 Sep 3;27(17):2293-304.