Newswise – Irvine, Calif., April 6, 2022 – In a new study at the University of California, Irvine, researchers have revealed the impact of native lipids on rhodopsin signaling and regeneration, potentially ushering in a new paradigm for drug discovery targeting coupled receptors to G proteins (GPCR).

GPCRs are cell surface receptors that respond to a variety of stimuli to activate signaling pathways across cell membranes. All GPCRs are membrane-bound and have rarely been studied in their native membrane environment. Recent advances have yielded atomic structures of key intermediates and roles for lipids in mediating signaling. However, capturing signaling events from a wild-type receptor in real time, across a native membrane to its downstream effectors, has remained elusive until now. These receptors represent by far the largest class of drug targets, and a large number of approved drugs modulate their functions.

In this new study published today in Natureentitled “Capturing a Rhodopsin Receptor Signaling Cascade Through a Native Membranethe researchers, using mass spectrometry, probed the archetypal class A GPCR, rhodopsin, directly in fragments of native disc membranes. They monitored the real-time photoconversion of dark-adapted rhodopsin to opsin, delineating stepwise retinal isomerization and retinal-opsin adduct hydrolysis, further finding that the reaction is significantly slower. in its natural membrane environment than in artificial detergent micelles.

“Human diseases, ranging from cancer to cardiovascular disease to blindness, are all strongly impacted by the function of GPCRs. In addition to the quantitative analysis of the signaling function, this new technology has, for the first time, allowed the direct detection of new potential targets of therapeutic interest for the visual system, within native membranes. I am confident that similar work will be done on many other GPCR systems,” explained Krzysztof Palczewski, PhDDonald Bren Professor of Ophthalmology at the UCI School of Medicine and co-corresponding author.

Considering lipids ejected with rhodopsin from membrane fragments in the mass spectrometer, researchers were able to demonstrate that opsin can be regenerated in membranes through photoisomerized retinal-lipid conjugates, and obtained evidence for increased association rhodopsin with the long chain unsaturated phosphatidylcholine. during signal transduction.

The team also captured secondary steps in the signaling cascade following rhodopsin activation. Monitoring transducin (Gt) light activation and guanosine diphosphate (GDP) dissociation to generate an intermediate apo trimeric G protein, they observed Gt_a.GTP subunits interacting with phosphodiesterase 6 (PDE6), found in cone and rod photoreceptor cells, which hydrolyzes the second messenger molecule cyclic guanosine monophosphate (cGMP).

“By applying compounds targeting rhodopsin, we have shown how they stimulate or attenuate signaling via the rhodopsin-opsin and transducin signaling pathways,” Palczewski said. “Using instantaneous light flashes, synchronized with recordings on a mass spectrometer, we were able to capture the signaling cascade and demonstrate the roles of lipids and ligands in rhodopsin signaling. This work highlights new opportunities for drug discovery in native environments and may lead to a new way to study the pharmacology of membrane-bound receptors.

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