Biased GPCR signaling by the native parathyroid hormone–related protein 1 to 141 relative to its N-terminal fragment 1 to 36

The parathyroid hormone (PTH)–related protein (PTHrP) is indispensable for the development of mammary glands, placental calcium ion transport, tooth eruption, bone formation and bone remodeling, and causes hypercalcemia in patients with malignancy. Although mature forms of PTHrP in the body consist of splice variants of 139, 141, and 173 amino acids, our current understanding on how endogenous PTHrP transduces signals through its cognate G-protein coupled receptor (GPCR), the PTH type 1 receptor (PTHR), is largely derived from studies done with its N-terminal fragment, PTHrP_1-36. Here, we demonstrate using various fluorescence imaging approaches at the single cell level to measure kinetics of (i) receptor activation, (ii) receptor signaling via Gs and Gq, and (iii) receptor internalization and recycling that the native PTHrP_1-141 displays biased agonist signaling properties that are not mimicked by PTHrP_1-36. Although PTHrP_1–36 induces transient cAMP production, acute intracellular Ca²⁺ (iCa²⁺) release and β-arrestin recruitment mediated by ligand–PTHR interactions at the plasma membrane, PTHrP_1-141 triggers sustained cAMP signaling from the plasma membrane and fails to stimulate iCa²⁺ release and recruit β-arrestin. Furthermore, we show that the molecular basis for biased signaling differences between PTHrP_1-36 and properties of native PTHrP_1-141 are caused by the stabilization of a singular PTHR conformation and PTHrP_1-141 sensitivity to heparin, a sulfated glycosaminoglycan. Taken together, our results contribute to a better understanding of the biased signaling process of a native protein hormone acting in conjunction with a GPCR.