Regulation of Adhesion G Protein-Coupled Receptor ADGRG1 (GPR56) by Receptor Activity-Modifying Proteins 公开

Gao, Anqi (2017)

Permanent URL:


G protein-coupled receptors (GPCRs) are a diverse superfamily of transmembrane proteins that transmit signals from the extracellular side of cells into the cytoplasm. Adhesion GPCRs are a family of receptors that are characterized by long N-termini containing adhesion-like motifs. ADGRG1 (also known as GPR56 or G1) is an adhesion GPCR that is involved in brain development. Mutations to this receptor cause a neurodevelopmental disorder known as bilateral frontoparietal polymicrogyria (BFPP). The purpose of this project was to examine receptor activity-modifying proteins (RAMPs) might regulate the activity of G1. We found that G1 physically associates with RAMP1 and RAMP3, and that a truncated constitutively-active version of G1 (ΔNT-G1) associates with RAMP1 and RAMP3 even more robustly. Co-expression with RAMPs had no significant effect on the signaling activity of G1 or ΔNT-G1 when receptor signaling to NFAT luciferase was assessed. Conversely, co-expression with RAMP3 sharply decreased ΔNT-G1 signaling to SRF luciferase, revealing a differential effect of RAMP3 on distinct signaling readouts. These findings represent the first description of RAMP interactions with adhesion GPCRs and provide a novel mechanism by which G1 activity may be regulated.

Table of Contents

Table of Contents

Abbreviations 1

I. Introduction

- Cell to cell communication 2

- G protein-coupled receptors 2

- Adhesion G protein-coupled receptors 4

- G protein-coupled receptor 56 (ADGRG1) 5

- Bilateral frontoparietal polymicrogyria (BFPP) 5

- Roles of ADGRG1 in the immune system and cancer 6

- Receptor activity-modifying proteins (RAMPs) 8

- Connections between G protein-coupled receptors and RAMPs 9

II. Materials and Methods

- Constructs 10

- Cell culture 10

- Co-Immunoprecipitation of G1 and ΔNT-G1with HA-RAMP1-3 10

- Evaluation of the effects of G1 and ΔNT-G1 interactions with HA-RAMP1-3 11

on signaling through G1 via NFAT and SRF-luciferase gene reporter assays

- Evaluation of G1, ΔNT-G1, and HA-RAMP3 surface expression via biotinylation 12

- Western blot 13

- Stripping of Western blot membranes 14

- Statistical analysis 15

III. Results

- G1 physically interacts with RAMP1 and RAMP3 in HEK-293T cells 15

- NFAT-luciferase assay 16

- SRF-luciferase assay 17

- Interactions between G1 and ΔNT-G1 with RAMP3 decreases surface expression 18

of G1 and RAMP3 increases surface expression of ΔNT-G1

- Interactions between G1 and ΔNT-G1 with RAMP3 increases RAMP3 surface expression 18

IV. Discussion and Conclusions 19

VI. Figures

- Figure 1 24

- Figure 2 25

- Figure 3 26

- Figure 4 27

- Figure 5 28

- Figure 6 29

- Figure 7 30

- Figure 8 31

VII. References 32

About this Honors Thesis

Rights statement
  • Permission granted by the author to include this thesis or dissertation in this repository. All rights reserved by the author. Please contact the author for information regarding the reproduction and use of this thesis or dissertation.
  • English
Research field
Committee Chair / Thesis Advisor
Committee Members

Primary PDF

Supplemental Files