The sodium leak channel NALCN (Na + leak channel non-selective) is the pore-forming subunit of the NALCN channelosome, formed together with the auxiliary proteins UNC79, UNC80, and FAM155A. This channelosome generates a depolarizing sodium current in numerous mammalian tissues, especially neurons, and is hence acting as a key regulator of cellular excitability. Several studies have suggested that NALCN function is regulated by G protein-coupled receptors (GPCRs), but the underlying molecular mechanisms of this regulation are poorly understood. Here, we investigate this regulation of NALCN by expressing the NALCN channelosome in Xenopus laevis oocytes together with various GPCRs and G protein subunits, using two-electrode voltage-clamp (TEVC) to evaluate its function. We further complement this with biochemical methods, using pulldowns and western blots to probe possible direct interactions between proteins. We find that co-expression of the inhibitory G protein α subunits Gα i , and Gα o leads to a strong reduction in NALCN currents, in line with previous reports suggesting NALCN inhibition by G i/o -coupled GPCRs. By contrast, co-expression of the stimulatory Gα s subunit strongly increases NALCN currents, a regulatory mechanism that had not yet been described. Using multiple complementary approaches, we demonstrate that this regulation by Gα s involves the C terminus of NALCN, whereas regulation by Gα o does not. Furthermore, pharmacological manipulation of the adenylyl cyclase, the common downstream target of Gα s and Gα i/o , does not affect NALCN function. Instead, pulldowns indicate a possible direct interaction between NALCN and Gα s . Overall, our data demonstrate that NALCN function is regulated by different G protein α subunits. This molecular mechanism of regulation further emphasizes the crucial role of NALCN in cellular excitability and may open new approaches for pharmacological manipulation.
Harms et al. (Sun,) studied this question.