Identification of a Critical Motif Responsible for Gating of Kir2.3 Channel by Intracellular Protons

Abstract

Protons are involved in gating Kir2.3. To identify the molecular motif in the Kir2.3 channel protein that is responsible for this process, experiments were performed using wild-type and mutated Kir2.3 and Kir2.1. CO 2 and low pH i strongly inhibited wild-type Kir2.3 but not Kir2.1 in whole cell voltage clamp and excised inside-out patches. This CO 2 /pH sensitivity was completely eliminated in a mutant Kir2.3 in which the N terminus was substituted with that in Kir2.1, whereas a similar replacement of its C terminus had no effect. Site-specific mutations of all titratable residues in the N terminus, however, did not change the CO 2 /pH sensitivity. Using several chimeras generated systematically in the N terminus, a 10-residue motif near the M1 region was identified in which only three amino acids are different between Kir2.3 and Kir2.1. Mutations of these residues, especially Thr 53 , dramatically reduced the pH sensitivity of Kir2.3. Introducing these residues or even a single threonine to the corresponding positions of Kir2.1 made the mutant channel pH-sensitive. Thus, a critical motif responsible for gating Kir2.3 by protons was identified in the N terminus, which contained about 10 residues centered by Thr 53 .The modulation of membrane excitability is a significant cellular property, which not only provides cells with an important strategy for responding to their external stimuli but also enables them to monitor their internal environment and intermediary metabolism (1). Inward rectifier K ϩ channels play a part in this process in which several intracellular molecules such as nucleotides and protons are involved (2, 3). Whereas the concentration of protons is normally controlled by several buffering and feedback-controlling systems, pH can fall beyond its normal levels under certain pathophysiological conditions. For instance, CO 2 retention, or hypercapnia, can cause a reduction in intra-and extracellular pH leading to respiratory acidosis (4).It is known that CO 2 sensing, which plays an important role in the feedback regulation of CO 2 and pH homeostasis in mammalian systems, is carried out by chemoreceptors, especially the central chemoreceptors (5-8). Through these chemoreceptors, a high PCO 2 level stimulates respiratory neuronal networks in the brain stem and enhances respiratory motor output. During this process, the information of PCO 2 levels received by these chemoreceptor cells may first be conveyed to the change in their membrane excitability and then passed to the respiratory neuronal networks through chemical or electrical synaptic transmissions (9, 10). Hence, the change in membrane excitability constitutes an important step in the CO 2 sensing.The alteration in membrane excitability with hypercapnia is known to be mediated by specific ion channels. CO 2 has been shown to enhance the firing activity of locus ceruleus neurons by suppressing a proton-and polyamine-sensitive inward rectifying K ϩ current (11). Our previous studies have shown that CO 2 induces a depolarization in Lymnaea snail ...

Differential Sensitivity of Inward Rectifier K+Channels to Metabolic Inhibitors

“…It has also been reported that K ir 2.3 is more sensitive than K ir 2.1 to decreased intracellular pH (14). Therefore the sensitivity of K ir 2.3 to metabolic inhibitors compared with K ir 2.1 is consistent with a decrease in intracellular pH caused by metabolic inhibition.…”

Kir2 potassium channels in rat striatum are strategically localized to control basal ganglia function

“…/2000COHEN, N.A. 1996;ZHU, G. 1999;HUANG, C.-L. 1998;COULTER, K.L. 1995;NAMBA, N. 1996; 25 CHEN, H. 1998 + = effect, -= no effect, (r) = rat, (h) = human KARSCHIN, C. 1999;KARSCHIN, C. 1996;FALK, T. 199529 TÖPERT, C. 1998TÖPERT, C. 2000 So far, only few studies investigated the distribution of Kir proteins in the mammalian brain by using specific antibodies 30 .…”

The K + channel K IR 2.1 functions in tandem with proton influx to mediate sour taste transduction

“…Notably, K IR 2.1 is much less sensitive to intracellular acidification than other inward rectifiers, such as K IR 2.3 and K IR 1.1 (35). Nonetheless, K IR 2.1 can be inhibited at a sufficiently low intracellular pH (≤5.5; our data and ref.…”

“…Nonetheless, K IR 2.1 can be inhibited at a sufficiently low intracellular pH (≤5.5; our data and ref. 35). Moreover, we find that K IR 2.1 can be blocked in whole-cell recording by weak acids that activate sour taste cells.…”

“…Sensitivity to intracellular acidification is not a well-documented feature of K IR 2.1 as it is for other K IR and K 2P channels (35,36). We therefore tested whether heterologously expressed K IR 2.1 channels could be blocked by intracellular acidification.…”