On the other hand, gain-of-function mutations in NCA-1, referred to as nca(gf) henceforth, lead to exaggerated body bending termed coiling ( Yeh et al., 2008). The in vivo physiological properties of these invertebrate CHIR-99021 manufacturer channels remain to be determined. However, genetic studies of the behavioral phenotypes of C. elegans ( Humphrey et al., 2007; Jospin et al., 2007; Yeh et al., 2008) and Drosophila ( Humphrey et al., 2007)

have led to the identification of UNC-79 and UNC-80, two conserved auxiliary subunits of this new channel. Multiple auxiliary subunits of sequence-related cation channels, such as the voltage-gated calcium channels (VGCCs), promote the stabilization and membrane localization of the channel, and/or modulate channel gating and kinetics ( Catterall, 2000b; Simms and Zamponi, 2012). Despite bearing no sequence similarity to known cation channel auxiliary subunits, UNC-79 and UNC-80 exert similar effects on the expression and localization of the NCA channel ( Jospin et al., 2007; Yeh et al.,

2008), and mUNC-80 couples the NALCN channel conductivity with an intracellular signaling cascade ( Lu et al., 2010). In C. elegans, the loss of either UNC-79 or UNC-80 suppresses and reverts the coiler phenotype exhibited by nca(gf) PD0332991 cell line to that of fainters ( Yeh et al., 2008). unc-79 and unc-80 mutants exhibit a fainter phenotype identical to that of nca(lf) mutants. The loss of either UNC-79 or UNC-80 causes a reduced localization of NCAs along the axon. UNC-79 and UNC-80 also localize along the axon, but only in Adenylyl cyclase the presence of NCAs, implicating their copresence in a channel complex ( Jospin et al., 2007; Yeh et al., 2008). Indeed, mouse mUNC-79 and mUNC-80 coimmunoprecipitated with NALCN (

Lu et al., 2010). Identifying genetic suppressors of nca(gf) therefore effectively reveals subunits or effectors of this new channel. Through genetic suppressor screens for nca(gf), we identified another recessive, loss-of-function suppressor, nlf-1, that rescues the coiler phenotype exhibited by nca(gf) animals. Below, we present molecular, biochemical, electrophysiological, calcium imaging and behavioral analyses on nlf-1 and nca that demonstrate (1) NCA contributes to a Na+ leak current in C. elegans neurons; (2) NLF-1 is an ER resident protein that specifically promotes axon delivery of the NCA Na+ leak channel; (3) NCA/NLF-1-mediated Na+ leak current maintains the RMP and potentiates the activity of premotor interneurons to sustain C. elegans’ rhythmic locomotion; (4) a mouse homolog mNLF-1 is functionally conserved with NLF-1 in vivo, and physically interacts with the mammalian Na+ leak channel NALCN in vitro. We isolated a recessive, loss-of-function mutation allele (hp428) of the nlf-1 gene that suppresses the behavioral phenotypes of nca(gf) mutants.