The effect of BFPP mutations on trafficking, signaling and function of ADGRG1/GPR56 receptor

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2022-9-12

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Demir, Nil

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Bilateral frontoparietal polymicrogyria (BFPP) is a hereditary brain abnormality characterized by defects on the brain cortex surface in the form of smaller (micro-) and multiplexed (poly-) gyri and sulci. More than 20 mutations in the ADGRG1/GPR56 gene have contributed to BFPP in various clinical studies, including in Turkish patients. ADGRG1/GPR56 receptor belongs to the adhesion G protein-coupled receptor family (aGPCR). Even though some of these mutations were found to reduce cell surface expression of the receptor, structural properties and molecular mechanisms of BFPP mutations that affect ADGRG1/GPR56 function are yet unknown. The study aims to (i) visualize the cell surface expression of tagged ADGRG1/GPR56 receptors carrying BFPP mutations using a laser scanning confocal microscope, (ii) investigate the effects of BFPP mutations on the receptor and Gα12 interactions, and (iii) investigate the effects of BFPP mutations on the receptor and β-arrestin interactions using Bioluminescence Resonance Energy Transfer (BRET) biosensors in live cells. Herein, eight missense BFPP mutations were introduced on ADGRG1/GPR56 receptor by using site-directed mutagenesis. The results showed mutant receptors had decreased cell surface expression compared to the wild-type. Gβ/ɣ and GRK, based BRET biosensors were used to investigate the coupling of ADGRG1/GPR56 with Gα12. C346S, W349S, and L640R mutations decreased the interaction of the receptor with Gα12 with respect to the wild-type. Arrestin-based BRET biosensor was used to investigate β-arrestin recruitment following receptor activation. All mutations tested in this study showed decreased beta-arrestin recruitment to the plasma membrane compared to the wild-type.

Subject Keywords

BFPP, BRET, GPR56, ADGRG1, Adhesion GPCR

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https://hdl.handle.net/11511/99566

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Graduate School of Natural and Applied Sciences, Thesis

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Citation Formats

N. Demir, “The effect of BFPP mutations on trafficking, signaling and function of ADGRG1/GPR56 receptor,” M.S. - Master of Science, Middle East Technical University, 2022.