Ginseng Gintonin Activates the Human Cardiac Delayed Rectifier K+ Channel: Involvement of Ca2+/Calmodulin Binding Sites

Abstract

Gintonin, a novel, ginseng-derived G protein-coupled lysophosphatidic acid (LPA) receptor ligand, elicits [Ca2+]i transients in neuronal and non-neuronal cells via pertussis toxin-sensitive and pertussis toxin-insensitive G proteins. The slowly activating delayed rectifier K+ (IKs) channel is a cardiac K+ channel composed of KCNQ1 and KCNE1 subunits. The C terminus of the KCNQ1 channel protein has two calmodulin-binding sites that are involved in regulating IKs channels. In this study, we investigated the molecular mechanisms of gintonin-mediated activation of human IKs channel activity by expressing human IKs channels in Xenopus oocytes. We found that gintonin enhances IKs channel currents in concentration- and voltage-dependent manners. The EC50 for the IKs channel was 0.05 ± 0.01 μg/ml. Gintonin-mediated activation of the IKs channels was blocked by an LPA1/3 receptor antagonist, an active phospholipase C inhibitor, an IP3 receptor antagonist, and the calcium chelator BAPTA. Gintonin-mediated activation of both the IKs channel was also blocked by the calmodulin (CaM) blocker calmidazolium. Mutations in the KCNQ1 [Ca2+]i/CaM-binding IQ motif sites (S373P, W392R, or R539W)blocked the action of gintonin on IKs channel. However, gintonin had no effect on hERG K+ channel activity. These results show that gintonin-mediated enhancement of IKs channel currents is achieved through binding of the [Ca2+]i/CaM complex to the C terminus of KCNQ1 subunit.

A brief method for preparation of gintonin-enriched fraction from ginseng

“…Gintonin induces intracellular Ca 2+ mobilization via the phospholipase C–IP 3 –Ca 2+ pathway in mammalian cells expressing endogenous LPA receptors [2] . Further studies demonstrated that gintonin-mediated transient mobilization of [Ca 2+ ] i is coupled with Ca 2+ -dependent in vitro regulations of ion channels, neurotransmitter release by enhancing synaptic transmission, and in vivo anti-Alzheimer's activity by activating nonamyloidogenic pathways [3] , [4] , [5] , [6] , [7] . We extracted ginseng with multiple procedures such as anion exchange chromatography and dialysis, yielding approximately 0.2% in the previous method ( Fig.…”

“…5 ). Because gintonin-mediated [Ca 2+ ] i transient via LPA receptors is linked to Ca 2+ -dependent diverse in vitro and in vivo effects [3] , [4] , [5] , [6] , [7] , [9] , [10] , the procedure for GEF preparation containing gintonin as an active ingredient could be applicable for mass production with a high yield ( Fig. 1 ).…”

“…The transient elevation of intracellular Ca 2+ via LPA receptor activation is a key mediator of diverse gintonin-mediated in vitro and in vivo effects. For example, gintonin regulates various Ca 2+ -dependent ion channels [3] , [4] , [5] and modulates N -methyl- d -aspartic acid and P2X 1 receptors via Ca 2+ -dependent signaling pathways [6] , [7] . Gintonin-mediated ion channels and receptor regulations are linked to the increase of gastrointestinal contractility by stimulation of pacemaker activity in the gastrointestinal system [8] and enhancement of synaptic transmission in hippocampal slices in the brain [9] .…”

Structure of ginseng major latex-like protein 151 and its proposed lysophosphatidic acid-binding mechanism

“…Gintonin from ginseng specifically activates G protein-coupled LPA receptors with high affinity in animal cells . In vitro studies show that gintonin regulates a variety of Ca 2+ -dependent ion channels such as BK Ca , Kv1.2 and KCNQ K + channels, and receptors such as NMDA and P2X 1 , via activation of endogenous LPA receptors (Shin et al, 2012;Lee et al, 2013;Choi et al, 2014). Other studies show that gintonin exhibits anti-Alzheimer's disease effects by activating the non-amyloidogenic pathway via LPA receptors, and produces antimetastatic effects by inhibiting autotoxin activity Hwang et al, 2013).…”

Genomics-Assisted Breeding

“…Resequencing data of 150 rice RILs with 0.02-0.05 × coverage were used for ultra-high resolution bin mapping (Huang et al 2009 ). However, resequencing approaches cost too much for plants with large genome sizes, such as pepper ) and ginseng (Choi et al 2014 ). It is also diffi cult to obtain accurate genotype data using low coverage resequencing data for complex duplicated plant genomes such as those found in Brassicaceae crops (Yang et al 2006 ;Liu et al 2014a , b ).…”