Abstract
The ATP-dependent potassium (KATP) channel in the pancreatic beta-cell is a complex of two proteins, the pore-forming Kir6.2 and the sulfonylurea receptor type 1 (SUR1). Both subunits are required for functional KATP channels because expression of Kir6.2 alone does not result in measurable currents. However, truncation of the last 26 or 36 amino acids of the C terminus of Kir6.2 enables functional expression of the pore-forming protein in the absence of SUR1. Thus, by using the truncated form of Kir6.2, expressed in the absence and presence of SUR1, it has been shown that the site at which ATP mediates channel inhibition is likely to be situated on Kir6.2. We have now examined the effects of long chain acyl-CoA (LC-CoA) esters on the C-terminally truncated mouse Kir6.2Delta365-390 (Kir6. 2DeltaC26) in inside-out patches isolated from Xenopus laevis oocytes. LC-CoA esters, saturated (C14:0, C16:0) and unsaturated (C18:1), increased Kir6.2DeltaC26 currents, whereas short and medium chain CoA esters (C3:0, C8:0, C12:0) were unable to affect channel activity. The LC-CoA esters were also able to counteract the blocking effect of ATP on Kir6.2DeltaC26. The stimulatory effect of the esters could be explained by the induction of a prolonged open state of Kir6.2DeltaC26. In the presence of the esters, channel open time was increased approximately 3-fold, which is identical to what was obtained in the native mouse KATP channel. Coexpression of SUR1 together with Kir6.2DeltaC26 did not further increase the ability of LC-CoA esters to stimulate channel activity. We conclude that Kir6.2 is the primary target for LC-CoA esters to activate the KATP channel and that the esters are likely to induce a conformational change by a direct interference with the pore-forming subunit, leading to openings of long duration.
Highlights
The ATP-sensitive potassium channel (KATP1 channel) couples glucose metabolism to electrical activity and insulin secretion in the pancreatic -cell
In oocytes injected with 50 nl H2O, no effect of long chain acyl-CoA (LC-CoA) esters on channel activity was obtained (Fig. 1D), showing that the stimulatory effect of the esters cannot be explained by activation of endogenous channels present in the oocyte
Our earlier studies have shown that LC-CoA esters are potent activators of KATP channel activity in the mouse pancreatic -cell and that CoA esters interact at a binding site different from that of ADP
Summary
The ATP-sensitive potassium channel (KATP1 channel) couples glucose metabolism to electrical activity and insulin secretion in the pancreatic -cell. Stimulation of the -cell with glucose leads to an increased ATP/ADP ratio resulting in inhibition of KATP channel activity, an effect which induces depolarization and activation of voltage-dependent Ca2ϩ channels, thereby triggering insulin secretion. The effects of the esters appear to be specific for the KATP channel, in that at least two other potassium channels are unaffected by the esters, the large-conductance Kϩ channel and the 8 pS Kϩ channel described in the -cell (2) This indicates that the KATP channel is a sensitive target for LC-CoA esters, and because of their ability to reverse the blocking effect of ATP, the esters may counteract glucose-induced depolarization of the -cell membrane potential. The stimulatory effect was achieved by saturated and unsaturated LC-CoA esters and characterized by openings of long duration
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