Human FSHR Stable Cell Line-HEK293

Here, researchers first assessed the potential of FSHR to activate steroidogenesis and proapoptotic signaling, potentially linked to cAMP production. Intracellular cAMP accumulation depends on FSHR-Gαs protein coupling, which increases continuously with increasing receptor expression levels. They demonstrated this using a BRET biosensor labeled with Gαs and FSHR. Results showed a leftward shift in the BRET saturation curve at the maximum concentration of the receptor-encoding plasmid used to transfect HEK293 cells (HEK293/FSHR). This suggests that receptor-G protein binding affinity is higher when FSHR is highly expressed (Figure 1A). The receptor expression-dependent association with Gαs was not observed with other G proteins reported to be FSHR intracellular interactors, such as Gαi and Gαq (Figures 1B-1E). Although Gαs may compete with other interacting proteins for binding to small amounts of FSHR, its potential to reduce cell viability (Figure 1F) and mediate basal and FSH-induced cAMP activation is FSHR concentration-dependent (Figure 1G). Increased intracellular cAMP is detrimental to the viability of FSHR-expressing cells. Indeed, treatment of FSHR-overexpressing HEK293 cells or primary human granulosa cells with 8-br-cAMP induced a concentration-dependent decrease in cell viability (Figures 1H and 1I). Interestingly, the decrease in granulosa cell viability induced by the cAMP analog was inhibited by 50 pg/ml E2 (Figure 1I), suggesting the potential for crosstalk between opposing intracellular effects mediated by gonadotropins and estrogens. Indeed, in the absence of E2, FSH treatment resulted in a decrease in FSHR-overexpressing HEK293 cell viability (Figures 1J and 1K).

Figure 1. Excessive FSHR Expression Levels Negatively Impact HEK293 Cell Viability. (Casarini L, et al., 2020)