elegans Unc-79 and Unc-80, respectively). First, mutations in Unc79 and Unc80 in Drosophila, C. elegans, and the mouse have essentially identical phenotypes to those of Nalcn mutants. Second, mutation in one of these genes also affects the protein levels and/or localization of the others ( Humphrey et al., 2007, Jospin et al., 2007, Nakayama et al., 2006, Pierce-Shimomura et al., 2008 and Yeh et al., 2008). Defining evidence that UNC79 and UNC80 form a physical complex with NALCN came from coimmunoprecipitation experiments in mouse brain showing that antibody

against any one of the three could bring BKM120 cell line down the others ( Lu et al., 2009 and Lu et al., 2010). Whether there are other core subunits in this complex awaits the purification of the protein complex and a determination of the subunit composition. UNC79 and UNC80 are well conserved among animals but share no obvious sequence similarity with any other protein with known function (Humphrey et al., 2007, Jospin et al., 2007,

Lu et al., 2009 and Lu et al., 2010). Both are large proteins (2654 aa and 3326 aa, respectively, in humans), larger than any known auxiliary subunit in the 24-TM channel family. Despite their size, there are no obviously identifiable Selleck Pfizer Licensed Compound Library protein domains in UNC79 and UNC80. The lack of sequence similarity among the auxiliary subunits of the NALCN, NaVs, CaVs, and CatSper suggest that, unlike the pore-forming subunits, the auxiliary subunits evolved independently. In heterologous expression systems, UNC80 and NALCN have been shown to interact (Lu et al., 2009 and Lu et al., 2010). UNC79 and UNC80 also associate with each other, and UNC79 requires the presence of UNC80 to associate with NALCN. These data suggest a NALCN complex model whereby UNC80 serves as a bridge between UNC79 and NALCN (Figure 4). In Unc79 knockout mouse brain, UNC80 protein is also undetectable, but NALCN is present. Unc79 mutant neurons also retain the NALCN-dependent basal Na+ leak current ( Lu et al., 2010). Unlike in wild-type neurons, however the current in the mutant is not regulated by G protein-coupled receptors (GPCRs; see

below). However, the GPCR regulation of NALCN can be restored by overexpression of UNC80 in the Unc79 knockout, suggesting that UNC80 is not required for the basal function of NALCN, but is required for the regulation of the channel ( Lu et al., 2010). Consistent with this idea, NALCN alone forms a leak channel when transfected into HEK293 cells, but its regulation by several GPCRs requires the cotransfection of UNC80 ( Lu et al., 2010). The function of UNC79 is less clear. The whole-cell basal Na+ leak is of a similar size in neurons cultured from the wild-type and in neurons from the Unc79 knockout mouse, and overexpression of UNC80 in an Unc79 null background restores the regulation of NALCN, suggesting that this channel’s biophysical properties in mouse brain, as they are currently understood, do not require UNC79 ( Lu et al., 2010).