Two-pore domain potassium channels (K2P) control neuronal excitability and play a central role in the establishment and maintenance of the resting membrane potential of almost all animal cells. In C. elegans, 47 genes code for K2P channel subunits but only four have been studied so far. We have found that heterologous expression of the uncharacterized K2P UNC-58 in Xenopus oocytes causes a strong depolarisation of the resting membrane potential, which is not consistent with the activity of a potassium-selective channel. In fact, ionic replacement experiments demonstrate that UNC-58 is highly permeable to sodium, which has never been reported in a two-pore domain potassium channel.
unc-58 is expressed in body wall muscles, and A, B and D-type motoneurons.
unc-58 null mutants are loopy on solid media and have a significantly decreased thrashing rate in liquid. Conversely, the dominant mutant
unc-58(
e665) displays a distinctive phenotype, whereby worms flip rapidly around their body axis, but are unable to move forward or backward. To identify factors that directly control the number, activity or distribution of K2P channels in vivo, we have performed a forward genetic screen for suppressors of
unc-58(
e665). Among 133 mutants, we have found six alleles of
unc-44/ankyrin and
unc-70/ beta -spectrin. In C. elegans, null mutations in
unc-70 and neuronal
unc-44 isoforms cause axonal fragility and strong axon guidance defects, respectively. Strikingly, these phenotypes are not observed in mutants from our screen. Using
zif-1/ZF1-mediated tissue-specific depletion, we show that
unc-58(
e665) paralysis is due to the activity of the channel in A and B motoneurons, but not in body wall muscles or D-type neurons. Therefore, UNC-44 and UNC-70 must be required for UNC-58 function in these neurons. In addition to its role in motoneurons, we find that
unc-44/ankyrin regulates UNC-58 trafficking to the cell surface in body wall muscles. In vertebrates, ankyrin binding to phosphatidylinositol 3-phosphate (PtdIns(3)P) is required for microtubule-dependent cargo transport. In an
unc-44 allele from our screen, a serine to leucine mutation near the PtdIns(3)P binding pocket is sufficient to disrupt UNC-58 localisation, and to relocate the channel to an intracellular compartment. This phenotype is specific for UNC-58, since two other muscle K2P channels, TWK-18 and SUP-9, are unaffected. Surprisingly, our beta -spectrin mutants do not alter the distribution of UNC-58, suggesting that ankyrin functions independently of beta -spectrin in muscle cells. Taken together, our data highlight new roles for
unc-58,
unc-44/ankyrin and
unc-70/ beta -spectrin in the control of neuromuscular function in C. elegans.