Glycine at the third position of TM3 determines the action of fluralaner
on insect and rat GABA receptor
Abstract
BACKGROUND AND PURPOSE Fluralaner is a novel isoxazoline insecticide
with broad insect spectrum, and mainly acts on the insect GABA receptor
with unique binding action, but its molecular interaction with insect
GABA receptor has not been deeply identified on molecular level
according to its selectivity between target (insect) and non-target
(mammal) organisms. EXPERIMENTAL APPOACH The potential binding residues
(I258T and L275I in TM1; V288I, M298N, G303N and A304S in TM2; G3’M/S,
A327S, G336N, M338I and A339F in TM3; M473V and I477D in TM4) were
predicted by SYBYL-X 2.1 software, and verified respectively by the
site-directed mutagenesis and two-electrode voltage clamp (TEVC)
technique. KEY RESULTS In the 11 predicted amino acids, the G3’M has the
strongest ability to reduce the sensitivity of recombinant rice stem
borer RDL homomeric channel to fluralaner. Compared with the wild-type
(WT)-RDL, the G3’M mutation almost completely abolish the binding of
fluralaner and avermectin, but not fipronil on recombinant homomeric
channel of RDL from several orders of insects in vitro. In addition, the
M3’G on rat Mus musculus β2 improved the sensitivity of recombinant
heteromeric Mmα1β2-M3’G channel to fluralaner. Our results demonstrated
that the glycine at the third position of TM3 determines the action of
fluralaner and should be the binding site of fluralaner with RDL.
CONCLUSION AND IMPLICATIONS These results would contribute to
understanding the molecular interaction of fluralaner with RDL homomeric
channel and may be used to guide future modification of isoxazolines to
achieve highly selective control of pests with minimal effects on
non-targeted organisms.