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Mediation of cardiac glycoside insensitivity in the monarch butterfly (Danaus plexippus): Role of an amino acid substitution in the ouabain binding site of Na+,K+-ATPase

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Abstract

The Monarch butterfly (Danaus plexippus) sequesters cardiac glycosides (CG) for its chemical defense against predators. Larvae and adults of this butterfly are insensitive towards dietary cardiac glycosides, whereas other Lepidoptera are sensitive and intoxicated by ouabain. Ouabain inhibits Na+,K+-ATPase by binding to its α-subunit. We have amplified and cloned the DNA-sequence encoding the respective ouabain binding site. Instead of the amino acid asparagine at position 122 in ouabain-sensitive insects, the Monarch has a histidine in the putative ouabain binding site, which consists of 12 amino acids. Starting with the CG-sensitive Na+,K+-ATPase gene fromDrosophila, we converted pos. 122 to a histidine residue as inDanaus plexippus by site-directed mutagenesis. Human embryonic kidney cells (HEK) (which are sensitive to ouabain) were transfected with the mutated Na+,K+-ATPase gene in a pSVDF-expression vector and showed a transient expression of the mutatedDrosophila Na+,K+-ATPase. When treated with ouabain, the transfected cells tolerated ouabain at a concentration of 50 mM, whereas untransformed controls or controls transfected with the unmutatedDrosophila gene, showed a substantial mortality. This result implies that the asparagine to histidine exchange contributes to ouabain insensitivity in the Monarch. In two other CG-sequestering insects, e.g.,Danaus gilippus andSyntomeida epilais, the pattern of amino acid substitution differed, indicating that the Monarch has acquired this mutation independently during evolution.

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  1. Institut für Pharmazeutische Biologie, Universität Heidelberg, Im Neuenheimer Feld 364, D-69120, Heidelberg, Germany

    F. Holzinger & M. Wink

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Holzinger, F., Wink, M. Mediation of cardiac glycoside insensitivity in the monarch butterfly (Danaus plexippus): Role of an amino acid substitution in the ouabain binding site of Na+,K+-ATPase. J Chem Ecol 22, 1921–1937 (1996). https://doi.org/10.1007/BF02028512

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