Author:

Keywords:

Alzheimer's disease, GPR3

Abstract:

The orphan G protein–coupled receptor (GPCR) GPR3 regulates activity of the γ-secretase complex in the absence of an effect on Notch proteolysis, providing a potential therapeutic target for Alzheimer’s disease (AD). However, given the vast resources required to develop and evaluate any new therapy for AD and the multiple failures involved in translational research, demonstration of the pathophysiological relevance of research findings in multiple disease-relevant models and understanding the mechanism of action are necessary prior to the initiation of costly drug development programs. The first part of the project involved evaluation of the physiological consequence of the loss of Gpr3 in four AD transgenic mouse models, including two that contain the humanized murine Aβ sequence and express similar amyloid precursor protein (APP) levels as wild-type mice, thereby reducing potential artificial phenotypes. Findings reveal that genetic deletion of Gpr3 reduced amyloid pathology in all of the AD mouse models and alleviated the cognitive deficits in APP/PS1 transgenic mice. Additional three-dimensional visualization and analysis of the amyloid plaque burden provided accurate information on the amyloid load, distribution, and volume in the structurally intact adult mouse brain. Analysis of 10 different regions in healthy human postmortem brain tissue indicate that GPR3 expression is stable during aging. However, two cohorts of human AD postmortem brain tissue samples show a correlation between elevated GPR3 and AD progression. Collectively, these studies provide significant evidence that GPR3 mediates the amyloidogenic proteolysis of APP in four AD transgenic mouse models as well as the physiological processing of APP in wild-type mice and suggest that GPR3 is a potential therapeutic target for AD drug development. The second part of the project focused on understanding the mechanism of the GPR3-mediated effect on Aβ generation. The results reveal that the phosphorylation status of GPR3 regulates β-arrestin 2 recruitment and Aβ generation. Mutagenesis of putative phosphorylation sites in GPR3 suggest that a specific phosphorylation barcode regulates the interaction between GPR3 and β-arrestin 2 and the subsequent Aβ generation. Genetic deletion of specific G protein–coupled receptor kinases (GRKs), which are involved in phosphorylation of GPCRs, in human embryonic kidney (HEK) cells led to a reduction in Aβ generation, suggesting that GRK-dependent phosphorylation of GPR3 is involved in the modulation of Aβ generation. Together, these studies demonstrate that the phosphorylation status of GPR3 is important for β-arrestin 2 recruitment and Aβ generation, and the process may involve preferential phosphorylation of GPR3 at the cytoplasmic tail by specific GRKs.