Duplication of the Rdl GABA receptor subunit gene in an insecticide-resistant aphid, Myzus persicae.
Resistance to cyclodiene insecticides is associated with replacements of a single amino acid (alanine 302) in a gamma-aminobutyric acid (GABA) receptor subunit encoded by the single-copy gene Resistance to dieldrin (Rdl). Alanine 302 is predicted to reside within the second membrane-spanning region of the Rdl receptor, a region that is thought to line the integral chloride ion channel pore. In all cyclodiene-resistant insects studied to date, this same alanine residue is replaced either by a serine, or, in some resistant strains of Drosophila simulans, a glycine residue. Therefore, individuals can carry only two different Rdl alleles. In contrast, here we report the presence of up to four different Rdl-like alleles in individual clones of the green peach aphid, Myzus persicae. In addition to the wild-type copy of Rdl gene (encoding A302 or allele A), M. persicae carries three other alleles with the following amino acid replacements: A302-->Glycine (allele G), A302-->SerineTCG (allele S) and A302-->SerineAGT (allele S'). Evidence from direct nucleotide sequencing and Single Stranded Conformational Polymorphism (SSCP) analysis shows that at least three of these different Rdl alleles (i.e. A, G and S) are commonly present in individual aphids or aphid clones. Southern analysis using allele-specific probes and analysis of sequences downstream of the exon containing the resistance-associated mutation confirm the presence of two independent Rdl-like loci in M. persicae. One locus carries the susceptible alanine (A) and/or resistant glycine (G) allele while the other carries the two serine alleles (S or S'). Whereas resistance levels are correlated with the glycine replacement, the S allele was present in all aphid clones, regardless of their resistance status. These results suggest that target site insensitivity is associated with replacements at the first (A/G) but not the second (S/S') locus. Phylogenetic analysis of nucleotide sequences indicates that both putative aphid Rdl loci are monophyletic with respect to other insect Rdl genes and may have arisen through a recent gene duplication event. The implications of this duplication with respect to insecticide resistance and insect GABA receptor subunit diversity are discussed.
Anthony
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N
Department of Entomology and Center for Neuroscience, University of Wisconsin-Madison, 53706, USA.
Unruh
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T
Ganser
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ffrench-Constant
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R
D000409:Alanine; D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D001042:Aphids / Q000187:drug effects / Q000235:genetics*; D001483:Base Sequence; D015139:Blotting, Southern; D029721:Drosophila Proteins; D004726:Endosulfan / Q000494:pharmacology; D020440:Gene Duplication; D007305:Insecticide Resistance / Q000235:genetics*; D007438:Introns; D008969:Molecular Sequence Data; D009154:Mutation; D010802:Phylogeny; D011963:Receptors, GABA-A / Q000235:genetics*; D017422:Sequence Analysis, DNA
D029721:Drosophila Proteins; C082030:Rdl protein, Drosophila; D011963:Receptors, GABA-A; D000409:Alanine; D004726:Endosulfan