Disease-causing helix-8 mutations reprogram parathyroid hormone receptor-1 effector selectivity.

Helix-8 is an important determinant of G protein-coupled receptor (GPCR) effector selectivity, yet its functional relevance and contribution to human disease remain poorly understood. Two helix-8 mutations in parathyroid hormone receptor-1 (PTH1R), E4658.49K and E4698.53K, cause skeletal dysplasia, but their mechanistic effects on signaling were unclear. Using live-cell biosensors, structural modeling, and humanized knock-in mice, we show that these mutants reconfigure polar networks at the receptor's cytoplasmic face to reshape effector coupling. E465K shifts PTH1R signaling away from Gαs toward Gαq activation, causing a switch in G protein selectivity. E469K stabilizes βarrestin engagement and enhances GRK2 recruitment, to thereby competitively limit G protein signaling. E469K-PTH1R mice show impaired calcemic responses to PTH, which are restored by a βarrestin-deficient PTH analog. These findings establish helix-8 as a critical determinant of GPCR effector selectivity and reveal mutation-driven rewiring of receptor signaling as a mechanism underlying PTH1R-associated disease.