Project 11 - Ligand-biased G-protein activation by CaSR

Host Institution: Stichting VU-VUMC
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Hypothesis: CaSR is believed to activate three different G-proteins in a ligand-combination dependent manner. So far, this has been indirectly inferred from downstream signalling activation, such as ERK, cAMP, and intracellular calcium oscillations. A second hypothesis is that the intracellular signalling state can (de-)sensitize CaSR for extracellular signals, for instance via its phosphorylation by PKC.

Objectives: To study ligand-biased activation of G_q and G_i by the CaSR as function of Ca²⁺_o, and allosteric modifiers at the level of single cells using G-protein sensors (G_q and G_i) to track receptor decisions at the membrane (AIM 1). These experiments will be carried out as function of the modulation of intracellular signalling processes, such as PKC and PP2A that are known to be involved in the phosphorylation of CaSR, to investigate whether the cellular signalling state can (de-)sensitize the receptor (AIM 2). To reach those objectives, ESR_VU will perform experiments using G_q & G_i FRET sensors using the same cell lines as ESR_UM. Mathematical modelling of CaSR signalling based on the results of ESR_VU, ESR_UM, ESR_MUW1 will be done to integrate data.