A unique amino acid of the Drosophila GABA receptor with influence on drug sensitivity by two mechanisms.

Zhang, H G; ffrench-Constant, Richard H.; Jackson, Meyer B.

doi:10.1113/jphysiol.1994.sp020278

Cited by 151 publications

(136 citation statements)

References 40 publications

Supporting

Mentioning

122

Contrasting

Order By: Relevance

“…In the present experiments, the substitution of glutamine or asparagine for the native serine may have influenced a state transition that altered picrotoxin inhibition of the channel, resulting in the use-facilitated blockade. Several studies have suggested that picrotoxin may inhibit GABA A receptors by stabilization of a desensitized state (11,12,31). We observed no evidence of this at the whole-cell level, but alterations in kinetic transitions can only be definitively observed with single channel recordings.…”

Section: Discussioncontrasting

confidence: 74%

“…An alternative explanation is that it may be part of a pivotal signal transduction point through which the effects of many of these drugs converge (13). Indeed, conformational changes corresponding to different functional states are greatly influenced by mutations within TMII (13,31,(42)(43). In the present experiments, the substitution of glutamine or asparagine for the native serine may have influenced a state transition that altered picrotoxin inhibition of the channel, resulting in the use-facilitated blockade.…”

Section: Discussionmentioning

confidence: 71%

See 1 more Smart Citation

Identification of a Novel Residue within the Second Transmembrane Domain That Confers Use-facilitated Block by Picrotoxin in Glycine α1 Receptors

Dibas

Gonzales

Das

et al. 2002

Journal of Biological Chemistry

View full text Add to dashboard Cite

The central nervous system convulsant picrotoxin (PTX) inhibits GABA A and glutamate-gated Cl ؊ channels in a use-facilitated fashion, whereas PTX inhibition of glycine and GABA C receptors displays little or no use-facilitated block. We have identified a residue in the extracellular aspect of the second transmembrane domain that converted picrotoxin inhibition of glycine ␣1 receptors from non-use-facilitated to usefacilitated. In wild type ␣1 receptors, PTX inhibited glycine-gated Cl ؊ current in a competitive manner and had equivalent effects on peak and steady-state currents, confirming a lack of use-facilitated block. Mutation of the second transmembrane domain 15-serine to glutamine (␣1(S15Q) receptors) converted the mechanism of PTX blockade from competitive to non-competitive. However, more notable was the fact that in ␣1(S15Q) receptors, PTX had insignificant effects on peak current amplitude and dramatically enhanced current decay kinetics. Similar results were found in ␣1(S15N) receptors. The reciprocal mutation in the ␤2 subunit of ␣1␤2 GABA A receptors (␣1␤2(N15S) receptors) decreased the magnitude of use-facilitated PTX inhibition. Our results implicate a specific amino acid at the extracellular aspect of the ion channel in determining use-facilitated characteristics of picrotoxin blockade. Moreover, the data are consistent with the suggestion that picrotoxin may interact with two domains in ligand-gated anion channels.Glycine receptors belong to a superfamily of ligand-gated chloride channels that include GABA A 1 receptors, GABA C receptors, and glutamate-gated chloride channels (1). In native tissue, glycine receptors exist as either ␣ homomers or ␣␤ heteromers (1). They comprise five subunits (usually three ␣ subunits and two ␤ subunits) arranged asymmetrically around the ion pore. Each subunit is made up of a large extracellular N-terminal region, four transmembrane domains (TM), and a large cytoplasmic domain; TMII forms the channel lumen (2). Glycine receptors are targets of therapeutics such as anesthetics as well as toxins like the central nervous system convulsant picrotoxin (1).Picrotoxin inhibits all known anionic ligand-gated Cl Ϫ channels (3-5). The mechanism of action and the exact location of picrotoxin binding are still unknown (6 -12). However, several studies have indicated that TMII is the probable site for picrotoxin action (6, 13-22) (Fig. 1). For example, the TMII of the glycine ␤ subunit was found to be responsible for conferring resistance to picrotoxin in heteromeric glycine ␣ n ␤ receptors (n ϭ 1-3) (6). Subsequent work has defined the existence of a phenylalanine residue at the 6Ј position of the TMII glycine ␤ subunit in conferring insensitivity to picrotoxin (16). In addition, other TMII residues (2Ј and 19Ј) have also been implicated directly or indirectly in the mechanism by which picrotoxin inhibits these channels (13, 15,16). The mutations at positions 2Ј and 19Ј have been shown to affect the type of the inhibition (competitive versus non-competitive) by picrotoxin ...

show abstract

Section: Discussioncontrasting

confidence: 74%

Section: Discussionmentioning

confidence: 71%

Identification of a Novel Residue within the Second Transmembrane Domain That Confers Use-facilitated Block by Picrotoxin in Glycine α1 Receptors

Dibas

Gonzales

Das

et al. 2002

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…Importantly, the CBZ effect on RDL desensitization was completely blocked by the A302S mutation, which has a potent effect on desensitization without drug (ref. 23 and Fig. 6d).…”

Section: Resultsmentioning

confidence: 81%

“…Similar to mammalian GABA A receptors, RDL channels mediate fast inhibitory neurotransmission 21 and are expressed in the CNS 22 . Notably, the mutation that causes the insecticide resistance phenotype (A302S) 20 specifically decreases the rate of RDL desensitization with little or no effect on other channel properties 23 . As a consequence, the mutant receptor has a longer single channel open duration and, therefore, increased channel current, at least under certain conditions (see below).…”

mentioning

confidence: 99%

Modulation of GABAA receptor desensitization uncouples sleep onset and maintenance in Drosophila

et al. 2008

View full text Add to dashboard Cite

Many lines of evidence indicate that GABA and GABA A receptors make important contributions to human sleep regulation. Pharmacological manipulation of these receptors has differential effects on sleep onset and sleep maintenance insomnia. Here we show that sleep is regulated by GABA in Drosophila and that a mutant GABA A receptor, Rdl A302S , specifically decreases sleep latency. The drug carbamazepine (CBZ) has the opposite effect on sleep; it increases sleep latency as well as decreasing sleep. Behavioral and physiological experiments indicated that Rdl A302S mutant flies are resistant to the effects of CBZ on sleep latency and that mutant RDL A302S channels are resistant to the effects of CBZ on desensitization, respectively. These results suggest that this biophysical property of the channel, specifically channel desensitization, underlies the regulation of sleep latency in flies. These experiments uncouple the regulation of sleep latency from that of sleep duration and suggest that the kinetics of GABA A receptor signaling dictate sleep latency.Insomnia is the most common sleep problem and affects approximately one third of the adult American population 1 . Patients with insomnia are generally subdivided into three categories: sleep onset insomnia, sleep maintenance insomnia and terminal insomnia (early-morning awakening coupled with an inability to return to sleep) 2 . The biological basis for these insomnia classifications remains unknown. Nonetheless, a single class of drugs, agonistic modulators of GABA A receptors, effectively ameliorates these diverse symptoms 2,3 . GABA A receptors are a family of pentameric ligand-gated Cl -channels 4 and are a major source of inhibitory currents throughout the CNS 5,6 . These receptors are also an important target for pharmacologic treatment of many other neurological disorders in addition to sleep 7 .The fruit fly Drosophila melanogaster is an ideal model for dissecting the relationships between molecules and behaviors, as well as between different sleep states 8,9 . As in mammals, it has been shown that the sleep-like state of Drosophila is associated with reduced sensory responsiveness and reduced brain activity 10,11 , and is subject to both circadian and homeostatic regulation 12,13 . Researchers have also identified a number of genes 14,15 , Notably, the mutation that causes the insecticide resistance phenotype (A302S) 20 specifically decreases the rate of RDL desensitization with little or no effect on other channel properties 23 . As a consequence, the mutant receptor has a longer single channel open duration and, therefore, increased channel current, at least under certain conditions (see below). Because of these characteristics, and because this mutation does not have obvious effects on health or viability, we decided to establish the importance of GABAergic transmission to sleep in flies and to examine the effects of the Rdl mutation.Interestingly, flies with this mutant GABA A receptor subunit slept more, primarily because of decreased sleep lat...

show abstract

“…Thus, the Ala 301 to Ser or Gly mutation in Rdl exhibits unique properties, providing high levels of dieldrin resistance without abolishing the role of the RDL receptor (12). Electrophysiological studies showed that the 301 replacement affects cyclodiene sensitivity by two mechanisms: inhibiting direct binding and allosterically modifying the Rdl receptor to disrupt the antagonist-favored conformation (13). These differences in channel properties have little effect on overall fitness in D. melanogaster other than temperature sensitivity.…”

mentioning

confidence: 99%

Gene duplication in the major insecticide target site,Rdl, inDrosophila melanogaster

Remnant

Good

Schmidt

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

The Resistance to Dieldrin gene, Rdl, encodes a GABA-gated chloride channel subunit that is targeted by cyclodiene and phenylpyrazole insecticides. The gene was first characterized in Drosophila melanogaster by genetic mapping of resistance to the cyclodiene dieldrin. The 4,000-fold resistance observed was due to a single amino acid replacement, Ala 301 to Ser. The equivalent change was subsequently identified in Rdl orthologs of a large range of resistant insect species. Here, we report identification of a duplication at the Rdl locus in D. melanogaster. The 113-kb duplication contains one WT copy of Rdl and a second copy with two point mutations: an Ala 301 to Ser resistance mutation and Met 360 to Ile replacement. Individuals with this duplication exhibit intermediate dieldrin resistance compared with single copy Ser 301 homozygotes, reduced temperature sensitivity, and altered RNA editing associated with the resistant allele. Ectopic recombination between Roo transposable elements is involved in generating this genomic rearrangement. The duplication phenotypes were confirmed by construction of a transgenic, artificial duplication integrating the 55.7-kb Rdl locus with a Ser 301 change into an Ala 301 background. Gene duplications can contribute significantly to the evolution of insecticide resistance, most commonly by increasing the amount of gene product produced. Here however, duplication of the Rdl target site creates permanent heterozygosity, providing unique potential for adaptive mutations to accrue in one copy, without abolishing the endogenous role of an essential gene.T he single point mutation in the Resistance to dieldrin (Rdl) gene represents one of the most significant cases of target site resistance to an insecticide yet observed. Cyclodiene resistance was reported in 62% of insecticide resistant species in the 1980s, following widespread use of cyclodiene insecticides, including dieldrin, which started in the 1950s (1). The nature of the genetic target, Rdl, was discovered after dieldrin was discontinued because of the widespread evolution of resistance in many species. Rdl was first discovered in Drosophila melanogaster using a positional cloning approach. High homology to human GABA receptors confirmed it was the first insect ligand-gated chloride channel subunit identified (2-4). A point mutation in the chloride channel pore-lining domain, replacing alanine 301 with serine, was present in all resistant D. melanogaster strains (5). This mutation provided 4,000-fold resistance when homozygous and lower levels of resistance in heterozygotes (2, 6). The homologous mutation was subsequently found in a large number of cyclodiene-resistant species from many insect orders (7-9), as well as a glycine replacement at the homologous site in some resistant strains of Drosophila simulans and other species (5, 10, 11).Characterization of deficiency lines and inversions in D. melanogaster showed that Rdl is an essential gene (3). Thus, the Ala 301 to Ser or Gly mutation in Rdl exhibits unique properties, p...

show abstract

A unique amino acid of the Drosophila GABA receptor with influence on drug sensitivity by two mechanisms.

Cited by 151 publications

References 40 publications

Identification of a Novel Residue within the Second Transmembrane Domain That Confers Use-facilitated Block by Picrotoxin in Glycine α1 Receptors

Identification of a Novel Residue within the Second Transmembrane Domain That Confers Use-facilitated Block by Picrotoxin in Glycine α1 Receptors

Modulation of GABAA receptor desensitization uncouples sleep onset and maintenance in Drosophila

Gene duplication in the major insecticide target site,Rdl, inDrosophila melanogaster

Contact Info

Product

Resources

About