Objective
G protein coupled receptors (GPCRs) are membrane proteins, which mediate a number of physiological signal transduction processes. Currently, about 60% of known pharmacological targets belong to the GPCR superfamily. Rhodopsin is a member of this super family of receptors, which initiates visual transduction. This signalling process comprises biochemical pathways underlying the complex sensorial phenomenon of vision. Activation of rhodopsin by light is caused by the cis-trans summarisation of its 11-cis-retinal chromophore upon light absorption. This initial ultra fast chemical event promotes a conformational change in rhodopsin, which allows it to bind and to activate a G protein called Gt, or transducin. Activation of Gt, in turn, is followed by a cascade of enzymatic reactions resulting in hyperpolarization of photoreceptor cell membranes. Much attention has been devoted to this activation mechanism, while the inactivation mechanism of photoactivated rhodopsin by phosphorylation by its kinase, GRK-1, is still less understood. In particular, the structural determinants in the cytoplasmic domain of rhodopsin responsible for GRK-1 binding and activation are still not determined in detail; the same is true for the domains and specific amino acid residues of GRK-1 responsible for interaction with and phosphorylation of rhodopsin. Furthermore, kinetic information about the receptor-kinase enzymatic reaction is still lacking. This knowledge is crucial for understanding the rhodopsin inactivation process and the role of the receptor-kinase interactions (as well as other involved key players like Ca2+) in retinal degenerations, particularly Retinitis Pigmentosa. Rhodopsin phosphorylation by its kinase GRK-1 is also a critical step in the desensitization process and plays a role in the fine regulation of the physiological process of light adaptation.
In the present project, key amino acid residues and structural elements in the cytoplasmic regions of rhodopsin (including membrane boundaries) crucial for rhodopsin phosphorylation by and interaction with GRK-1 will be elucidated. To this aim, rhodopsin mutants, including those related to the retinal degenerative disease Retinitis Pigmentosa, in these domains would be produced. The rhodopsin mutants will be structurally and biochemically characterized, and the levels of their phosphorylation by GRK-1 will be compared to reveal amino acid residues in rhodopsin crucial for its recognition by GRK-1. The phenomenon of constitutive phosphorylation of rhodopsin will also be investigated. This will be followed by a detailed study of the different kinetic parameters, with the aim to obtain a complete mechanistic description of the receptor-kinase interaction (both binding and dissociation), will be undertaken. This will be done using a novel spectroscopic technique, surface plasmon resonance spectroscopy (SPR-spectroscopy). In these experiments, both non-phosphorylated and phosphorylated rhodopsin of a wild type.
Programme(s)
Multi-annual funding programmes that define the EU’s priorities for research and innovation.
Multi-annual funding programmes that define the EU’s priorities for research and innovation.
Topic(s)
Calls for proposals are divided into topics. A topic defines a specific subject or area for which applicants can submit proposals. The description of a topic comprises its specific scope and the expected impact of the funded project.
Calls for proposals are divided into topics. A topic defines a specific subject or area for which applicants can submit proposals. The description of a topic comprises its specific scope and the expected impact of the funded project.
Call for proposal
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Procedure for inviting applicants to submit project proposals, with the aim of receiving EU funding.
Funding Scheme
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Funding scheme (or “Type of Action”) inside a programme with common features. It specifies: the scope of what is funded; the reimbursement rate; specific evaluation criteria to qualify for funding; and the use of simplified forms of costs like lump sums.
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Coordinator
08193 Bellaterra (Barcelona)
Spain
The total costs incurred by this organisation to participate in the project, including direct and indirect costs. This amount is a subset of the overall project budget.