KT3.2 and KT3.3, Two Novel Human Two-Pore K⁺Channels Closely Related to TASK-1

Abstract

We report the cloning of human KT3.2 and KT3.3 new members of the two-pore K(+) channel (KT) family. Based on amino acid sequence and phylogenetic analysis, KT3.2, KT3.3, and TASK-1 constitute a subfamily within the KT channel mammalian family. When Xenopus oocytes were injected with KT3.2 cRNA, the resting membrane potential was brought close to the potassium equilibrium potential. At low extracellular K(+) concentrations, two-electrode voltage-clamp recordings revealed the expression of predominantly outward currents. With high extracellular K(+) (98 mM), the current-voltage relationship exhibited weak outward rectification. Measurement of reversal potentials at different [K(+)](o) revealed a slope of 48 mV per 10-fold change in K(+) concentration as expected for a K(+)-selective channel. Unlike TASK-1, which is highly sensitive to changes of pH in the physiological range, KT3.2 currents were relatively insensitive to changes in intracellular or extracellular pH within this range due to a shift in the pH dependency of KT3.2 of 1 pH unit in the acidic direction. On the other hand, the phorbol ester phorbol 12-myristate 13-acetate (PMA), which does not affect TASK-1, produces strong inhibition of KT3.2 currents. Human KT3.2 mRNA expression was most prevalent in the cerebellum. In rat, KT3.2 is exclusively expressed in the brain, but it has a wide distribution within this organ. High levels of expression were found in the cerebellum, medulla, and thalamic nuclei. The hippocampus has a nonhomogeneous distribution, expressing at highest levels in the lateral posterior and inferior portions. Medium expression levels were found in neocortex. The KT3.2 gene is located at chromosome 8q24 1-3, and the KT3.3 gene maps to chromosome 20q13.1.

Motoneurons Express Heteromeric TWIK-Related Acid-Sensitive K+(TASK) Channels Containing TASK-1 (KCNK3) and TASK-3 (KCNK9) Subunits

“…Bottom, Immunohistochemistry was used to detect TASK-3 distribution in sections of rat brain and adrenal gland. TASK-3-like immunoreactivity was detected in a distribution pattern that was strikingly similar to previous TASK-3 in situ hybridization reports Talley et al, 2001;Vega-Saenz de Miera et al, 2001;Bayliss et al, 2003). Especially strong labeling was seen in cell bodies from the following regions (clockwise from top left): hypoglossal motor nucleus (HMs), locus ceruleus (LC), adrenal gland, anterodorsal thalamic nucleus (ThAD), dorsal raphe nucleus (RDo), and striatum.…”

“…1 A) and in the basal forebrain (data not shown). One other nucleus of note was the anterodorsal thalamic nucleus, which also contains particularly high levels of TASK-3 mRNA Talley et al, 2001;Vega-Saenz de Miera et al, 2001). Outside the CNS, TASK-3 immunoreactivity was extremely high in the adrenal gland, again as expected given earlier functional and RNA expression studies (Czirjak and Enyedi, 2002b;Bayliss et al, 2003); in the adrenal gland, immunostaining was readily apparent in the medulla and was extremely intense in the outer cortex, particularly in the zona glomerulosa .…”

“…6 A). In motoneurons and in other neuronal cell groups in which both channels are expressed Talley et al, 2001;Vega-Saenz de Miera et al, 2001), the potential to assemble channels in homomeric or heteromeric configurations provides a mechanism for increasing diversity of leak K ϩ channel properties. For example, changes in the relative expression of TASK-1 and TASK-3, as happens in cerebellar granule neurons through development (Han et al, 2002;Lauritzen et al, 2003), could serve to favor formation of different combinations of homomeric or heteromeric TASK channels and thereby regulate the pH range over which cells respond.…”

“…At that time, which preceded the cloning of TASK-3, we attributed the native motoneuronal current to TASK-1 because the pK for inhibition was ϳ7.4, essentially identical to that of TASK-1 Talley et al, 2000). However, in light of current evidence for strong expression of TASK-3 in motoneurons Talley et al, 2001;Vega-Saenz de Miera et al, 2001), we suspected that those subunits could contribute to native TASKlike motoneuronal currents, perhaps paired with TASK-1 in heterodimeric channels. In the present study, we used biochemical and electrophysiological approaches in a heterologous expression system to demonstrate formation of heterodimeric TASK channels and to characterize their properties.…”

“…This combinatorial potential provides a mechanism for additional diversity in K ϩ channel properties within individual cells (Wei et al, 1990;Krapivinsky et al, 1995;Wang et al, 1998), but evidence in support of native heterodimeric channels of the KCNK family is scarce (Kang et al, 2003). In the case of TASK-1 and TASK-3, heterodimeric channels have been detected in heterologous expression systems (Czirjak and Enyedi, 2002a;Kang et al, 2003;Lauritzen et al, 2003; but see Karschin et al, 2001;Pei et al, 2003), and a substantial overlap of subunit expression in many areas of the CNS provides a substrate for their coassembly in vivo Talley et al, 2001;Vega-Saenz de Miera et al, 2001). In cerebellar granule neurons that express TASK-1 and TASK-3, coassociation of TASK channels into functional native heterodimers was demonstrated by taking advantage of the unique single-channel properties and ruthenium red sensitivity defined for recombinant linked heterodimeric TASK-1/TASK-3 channels (Kang et al, 2003).…”