Background: KV10.1 is a potassium channel expressed in the brain and important for non-neural tumorigenesis.Results: An interaction between the C terminus of KV10.1 and the proline-rich domain of cortactin stabilizes the channel at the plasma membrane.Conclusion: Cortactin interacts with KV10.1 and controls surface expression of the channel.Significance: Our findings provide a functional and mechanistic link between the functions of two oncology-relevant proteins.

Ionotropic, AMPA-type glutamate receptors (GluRs) critically shape excitatory synaptic signals in the CNS. Ligand binding induces conformational changes in the glutamate-binding domain of the receptors that are converted into opening of the channel pore via three short linker sequences, a process referred to as gating. Although crystallization of the glutamate-binding domain and structural models of the ion pore advanced our understanding of ligand-binding dynamics and pore movements, the allosteric coupling of both events by the short linkers has not been described in detail. To study the role of the linkers in gating GluR1, we transplanted them between different GluRs and examined the electrophysiological properties of the resulting chimeric receptors in Xenopus laevis oocytes and HEK293 cells. We found that all three linkers decisively affect receptor functionality, agonist potency, and desensitization. One linker chimera was nondesensitizing and exhibited strongly increased agonist potencies, while fluxing ions even in the absence of agonist, similar to properties reported for the GluR1 lurcher mutation. Combining this new lurcher-like linker chimera with the original lurcher mutation allowed us to reassess the effect of lurcher on GluR1 gating properties. The observed differential but interdependent influence of linker and lurcher mutations on receptor properties suggests that the linkers are part of a fine-tuned structural element that normally stabilizes the closed ion pore. We propose that lurcher-like mutations act by disrupting this element such that ligand-induced conformational changes are not necessarily required to gate the channel.Key words: glutamate; AMPA receptor; gating; lurcher; linker; desensitization IntroductionIonotropic glutamate receptors (GluRs) are responsible for the majority of excitatory neurotransmission in the mammalian brain . Each GluR subunit harbors an extracellular N terminus, an intracellular C terminus, three transmembrane domains (TMDs; A-C), and a hairpin loop between TMDs A and B (see Fig. 1 B). TMD B and the hairpin loop are the major channel-lining domains and together with TMD A are inserted between two discontinuous stretches of sequence, S1 and S2, which make up the ligand-binding domain (LBD).Gating in GluRs encompasses the conformational change induced by ligand binding and its subsequent translation into pore opening. This movement is propagated via three short linkers of 9 -17 aa that connect the LBD with the TMDs, presumably forming the outer vestibule of the ion channel (Beck et al., 1999). Inclusion of these linkers in the crystal structure of the LBD was precluded by their flexibility (Armstrong et al., 1998), and structural determinants of gating that reside within them still remain completely unknown. So far it has only been reported that mutations in the linker preceding TMD A (linker A) (Stern-Bach et al., 1998), as well as a point mutation in the linker following TMD B (linker B), attenuate AMPA receptor (AMPAR) desensitization .We transplanted all th...

Although crucial for their correct function, the mechanisms controlling surface expression of ion channels are poorly understood. In the case of the voltage-gated potassium channel KV10.1, this is determinant not only for its physiological function in brain, but also for its pathophysiology in tumors and possible use as a therapeutic target. The Golgi resident protein PIST binds several membrane proteins, thereby modulating their expression. Here we describe a PDZ domain-mediated interaction of KV10.1 and PIST, which enhances surface levels of KV10.1. The functional, but not the physical interaction of both proteins is dependent on the coiled-coil and PDZ domains of PIST; insertion of eight amino acids in the coiled-coil domain to render the neural form of PIST (nPIST) and the corresponding short isoform in an as-of-yet unknown form abolishes the effect. In addition, two new isoforms of PIST (sPIST and nsPIST) lacking nearly the complete PDZ domain were cloned and shown to be ubiquitously expressed. PIST and KV10.1 co-precipitate from native and expression systems. nPIST also showed interaction, but did not alter the functional expression of the channel. We could not document physical interaction between KV10.1 and sPIST, but it reduced KV10.1 functional expression in a dominant-negative manner. nsPIST showed weak physical interaction and no functional effect on KV10.1. We propose these isoforms to work as modulators of PIST function via regulating the binding on interaction partners.