title: Osteopontin activates the diabetes-associated potassium channel TALK-1 in pancreatic β-cells.
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.
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, Tennessee, United States of America.
authors.forename: Matthew T
authors.identifier: http://orcid.org/0000-0002-6187-3951
authors.initials: MT
authors.lastname: Dickerson
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, Tennessee, United States of America.
authors.forename: Nicholas C
authors.initials: NC
authors.lastname: Vierra
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, Tennessee, United States of America.
authors.forename: Sarah C
authors.initials: SC
authors.lastname: Milian
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, Tennessee, United States of America.
authors.forename: Prasanna K
authors.initials: PK
authors.lastname: Dadi
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, Tennessee, United States of America.
authors.forename: David A
authors.initials: DA
authors.lastname: Jacobson
mesh_terms: D000368:Aged; D000818:Animals; D020013:Calcium Signaling; D003924:Diabetes Mellitus, Type 2 / Q000378:metabolism / Q000473:pathology; D005260:Female; D005947:Glucose / Q000502:physiology; D057809:HEK293 Cells; D006801:Humans; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000378:metabolism*; D008564:Membrane Potentials; D018345:Mice, Knockout; D053495:Osteopontin / Q000502:physiology*; D011188:Potassium / Q000378:metabolism; D024683:Potassium Channels, Tandem Pore Domain / Q000378:metabolism*
chemical_list: D007328:Insulin; C477398:KCNK16 protein, human; D024683:Potassium Channels, Tandem Pore Domain; C506895:SPP1 protein, human; D053495:Osteopontin; D005947:Glucose; D011188:Potassium