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The True Identity of GPR30/GPER : Ligand Discovery and Yeast-Based Functional Testing
Location: 34
Mentor: Dr. Daniel Isom
G-protein estrogen receptor (GPER), also known as GPR30, has long been proposed as a direct estrogen receptor based on its reported activation by 17β-estradiol (E2). However, recent structural and functional studies show that E2 does not bind GPR30. Instead, Lys05 and other positively charged ligands were identified as the true GPR30 activators. To investigate signaling and verify newly proposed ligands, specialized human-yeast chimeras were CRISPR-edited to express GPR30 across 10 Gα subunits. A high-throughput yeast-based GPCR assay (DCyFIR) was employed, which showed that GPR30 did not couple with any of the subunits. None of the tested ligands—including E2—elicited a functional response. To better understand GPR30’s structural and evolutionary context, a comparative bioinformatic analysis was performed. BLAST and Foldseek searches identified candidates based on homology and structure respectively. Phylogenetic analysis revealed that GPR30 is evolutionarily distant, branching off earlier than other GPCRs in the dataset. Structural analysis revealed strong similarity between GPR30 and receptors typically involved in opioid and peptide/hormone-like signaling, despite distant sequence homology. This suggests that small, charged peptide ligands are likely physiological activators of GPR30, rather than a rigid, hydrophobic steroid like E2. The integration of GPR30 into the DCyFIR system and the subsequent verification of new ligands is crucial in determining the role of GPR30 signaling in humans and the development of therapeutics. Further testing in human cells is necessary to assess whether the ligands are truly nonfunctional or just unable to signal in DCyFIR.
