Research Article: Osteopontin activates the diabetes-associated potassium channel TALK-1 in pancreatic β-cells

Date Published: April 12, 2017

Publisher: Public Library of Science

Author(s): Matthew T. Dickerson, Nicholas C. Vierra, Sarah C. Milian, Prasanna K. Dadi, David A. Jacobson, Bernard Attali.

http://doi.org/10.1371/journal.pone.0175069

Abstract

Glucose-stimulated insulin secretion (GSIS) relies on β-cell Ca2+ influx, which is modulated by the two-pore-domain K+ (K2P) channel, TALK-1. A gain-of-function polymorphism in KCNK16, the gene encoding TALK-1, increases risk for developing type-2 diabetes. While TALK-1 serves an important role in modulating GSIS, the regulatory mechanism(s) that control β-cell TALK-1 channels are unknown. Therefore, we employed a membrane-specific yeast two-hybrid (MYTH) assay to identify TALK-1-interacting proteins in human islets, which will assist in determining signaling modalities that modulate TALK-1 function. Twenty-one proteins from a human islet cDNA library interacted with TALK-1. Some of these interactions increased TALK-1 activity, including intracellular osteopontin (iOPN). Intracellular OPN is highly expressed in β-cells and is upregulated under pre-diabetic conditions to help maintain normal β-cell function; however, the functional role of iOPN in β-cells is poorly understood. We found that iOPN colocalized with TALK-1 in pancreatic sections and coimmunoprecipitated with human islet TALK-1 channels. As human β-cells express two K+ channel-forming variants of TALK-1, regulation of these TALK-1 variants by iOPN was assessed. At physiological voltages iOPN activated TALK-1 transcript variant 3 channels but not TALK-1 transcript variant 2 channels. Activation of TALK-1 channels by iOPN also hyperpolarized resting membrane potential (Vm) in HEK293 cells and in primary mouse β-cells. Intracellular OPN was also knocked down in β-cells to test its effect on β-cell TALK-1 channel activity. Reducing β-cell iOPN significantly decreased TALK-1 K+ currents and increased glucose-stimulated Ca2+ influx. Importantly, iOPN did not affect the function of other K2P channels or alter Ca2+ influx into TALK-1 deficient β-cells. These results reveal the first protein interactions with the TALK-1 channel and found that an interaction with iOPN increased β-cell TALK-1 K+ currents. The TALK-1/iOPN complex caused Vm hyperpolarization and reduced β-cell glucose-stimulated Ca2+ influx, which is predicted to inhibit GSIS.

Partial Text

Glucose-stimulated insulin secretion (GSIS) from pancreatic β-cells is critical to the maintenance of glucose homeostasis. This process requires pancreatic β-cell Ca2+ influx through voltage dependent calcium channels (VDCCs), which are activated by plasma membrane depolarization. Two-pore-domain potassium (K2P) channels are important to this process due to their role in regulating beta-cell membrane potential (Vm) [1–3]. For example, the TWIK-related alkaline pH-activated K2P (TALK)-1 channel and the TWIK-related acid-sensitive K2P (TASK)-1 channel hyperpolarize β-cell Vm [4–6]. These K2P channels help to set the basal beta-cell Vm with their continuous K+ flux at all physiological Vm [7–9]. However, the conductance of K2P channels is significantly less than that of the ATP sensitive K+ channel (KATP), the primary K+ channel responsible for setting the beta-cell Vm under low glucose conditions [8, 10, 11]. Thus, the activity of beta-cell K2P channels would be expected to play a more prominent role in controlling the Vm during glucose stimulation and KATP inhibition. Moreover, the small conductance of beta-cell K2P channels stabilizes the glucose induced plateau potential from which action potentials fire at a Vm that is optimal for activation of VDCCs [12, 13]. While K2P channels play an important role in regulating beta-cell Vm and Ca2+ influx, the mechanism(s) controlling beta-cell K2P channels have not been elucidated.

TALK-1 channel modulation of β-cell Vm regulates Ca2+ influx and second phase GSIS [6]. A coding sequence polymorphism that results in a GOF of TALK-1 channels is also responsible for an increased risk of developing type 2 diabetes (T2D) [19, 20]. However, the mechanisms that influence beta-cell TALK-1 channel activity have not been determined. Here we utilized an unbiased MYTH assay to identify TALK-1 channel-interacting proteins from human islets, which enabled us to identify the first protein interactions with TALK-1 channels. These interacting partners of TALK-1 will be useful in identifying β-cell regulatory complexes that control TALK-1 channel activity and contribute to islet function.

 

Source:

http://doi.org/10.1371/journal.pone.0175069

 

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