Localization of TREK-2 K + channel domains that regulate channel kinetics and sensitivity to pressure, fatty acids and pH i

Abstract

TREK-2, a member of the tandem-pore K+ channel family, is activated by membrane stretch, unsaturated free fatty acids and acidic conditions, and exhibits unique open channel kinetics. To identify the regions responsible for these properties, we studied the role of the cytoplasmic regions of TREK-2. Deletion of the N-terminus had no effect on any aspect of TREK-2 function. Deletion of the C-terminus or its substitution with that of TASK-3 abolished the sensitivity to free fatty acids and intracellular pH (pHi), and reduced the sensitivity to pressure. The regions that allow activation by free fatty acids and low pHi were localized to 25- and 10-amino-acid domains, respectively, close to the fourth transmembrane segment. Substitution of KKTKEE, a charged region near the proximal C-terminus, with uncharged amino acids produced little change in TREK-2 function; however, its deletion abolished sensitivity to fatty acids and low pH, indicating that this region is structurally very important. The TREK-2 C-terminus was also found to be critical for its channel opening in bursts as well as for its increased basal activity. Thus, the C-terminus endows TREK-2 with unique channel kinetics and the ability to be gated by free fatty acids and low pHi, and with increased mechanosensitivity.

Heterodimerization within the TREK channel subfamily produces a diverse family of highly regulated potassium channels

“…pH is an especially prominent regulator of these channels. Intracellular acidification stimulates both TREK1 and TREK2 (8,9), whereas TRAAK is stimulated by intracellular alkalinization (10). In contrast, extracellular acidification is able to inhibit TREK1 and TRAAK, but activates TREK2 (11).…”

The Inhibition of TREK2 Channel by an Oxidizing Agent, 5,5'-dithio-bis (2-nitrobenzoic acid), via Interaction with the C-terminus Distal to the 353rd Amino Acid

“…The C-terminal domain has been known to be critical for TREK1 and TREK2 activation, resulting from membrane stretching, temperature and intracellular acidosis (Honore et al, 2002;Kim et al, 2001b;Maingret et al, 2000;Maingret et al, 1999;Patel et al, 1998). On the other hand, the C-terminus of TRAAK is not important for mediating fatty acid sensitivity (Kim et al, 2001a).…”

“…PCR was used to generate deletions and chimeras in TREK2 and TASK3, as described previously (Kim et al, 2001b). The point mutation was performed using a site-directed mutation kit (Stratagene, La Jolla, CA).…”

“…Inhibition of TREK1 by hypoxia requires the C-terminus of the channel (Miller et al, 2005), and structural modification of the channel can also lead to hypoxic effects. Indeed, the C-terminal domains of TREK1 and TREK2 may modulate electrophysiological properties that are critical for channel activation, resulting from membrane stretching, temperature changes and intracellular acidosis (Honore et al, 2002;Kim et al, 2001b;Maingret et al, 2000;Maingret et al, 1999;Patel et al, 1998).…”

“…It has earlier been suggested that truncation of the TREK2 C-terminal end can reduce mechanosensitivity, arachidonic acid sensitivity, and intracellular hydrogen sensitivity (Bang et al, 2000;Kim et al, 2001b). Therefore, we compared the effects of intracellular acidic pH on wild type TREK2 and TREK2 (1-353)/TASK3C chimera in the presence or absence of DTNB (Fig.…”