Novel sodium channel gene mutations in Blattella germanica reduce the sensitivity of expressed channels to deltamethrin

Abstract

Pyrethroid insecticides alter the normal gating of voltage-gated sodium channels in the nervous system. Three sodium channel mutations (E434K, C764R, L993F) were recently identified in pyrethroid resistant German cockroach populations. In this report, we show that the L993F mutation decreased sodium channel sensitivity to the pyrethroid, deltamethrin, by five-fold in Xenopus oocytes. In contrast, neither E434K nor C764R alone decreased channel sensitivity to deltamethrin. However, E434K or C764R combined with L993F reduced deltamethrin sensitivity by 100-fold. Furthermore, concomitant presence of all three mutations (KRF) reduced channel sensitivity to deltamethrin by 500-fold. None of the mutations significantly affected channel gating. However, sodium current amplitudes from the mutant sodium channel carrying either E434K or C764R alone were much reduced compared to those of the wild-type channel or the channel carrying the double or triple mutations (KF, RF and KRF). These results indicated that evolution of sodium channel insensitivity in the German cockroach is achieved by sequential selection of a primary mutation L993F and two secondary mutations E434K and C764R, and concomitant presence of all three mutations dramatically reduced sodium channel sensitivity to deltamethrin.

Three mutations identified in the voltage-sensitive sodium channel α-subunit gene of permethrin-resistant human head lice reduce the permethrin sensitivity of house fly Vssc1 sodium channels expressed in Xenopus oocytes

“…Assays with channels containing the M827I and L932I mutations, either singly or in combination, also exhibited acceleration of the rate of tail current decay that was correlated with the degree of resistance afforded to 200 μM permethrin. Both the reduction in permethrin sensitivity and the acceleration of tail current decay observed with channels containing the M827I and L932I mutations are qualitatively similar the to the previously-characterized effects of the V410M and L1014F mutations on the permethrin sensitivity of D. melanogaster Para/tipE sodium channels (Lee et al, 2000a;Feng et al, 1995;Smith et al, 1997;Warmke et al, 1997;Tan et al, 2002b;Smith et al, 1998;Zhao et al, 2000;Vais et al, 2000;Lee et al, 1999b) and the cismethrin sensitivity of Vssc1 WT /tipE sodium channels (Lee et al, 2000a;Feng et al, 1995;Smith et al, 1997;Warmke et al, 1997;Tan et al, 2002b;Smith et al, 1998;Zhao et al, 2000;Vais et al, 2000;Lee et al, 1999b).…”

“…Instead, cypermethrin causes profound use-dependent modification of sodium currents with more persistent tail currents than those induced by cismethrin (Smith et al, 1998;Lee et al, 1999b). Studies with deltamethrin, a close structural relative of cypermethrin, and other insect sodium channels in the Xenopus oocyte expression system confirm the exclusively use-dependent action of these α-cyano pyrethroids (Vais et al, 2000;Tan et al, 2002b;Vais et al, 2003).…”

“…The decay constants for permethrin-induced tail currents did not vary with permethrin concentration, implying that the rate of tail current decay is an intrinsic property of each combination of pyrethroid and channel variant. Moreover, studies of the action of deltamethrin on pyrethroid resistance mutations in German cockroach sodium channels showed that resistance in several different mutated channel variants was not correlated with the acceleration of the decay of deltamethrin-induced tail currents (Tan et al, 2002b). Therefore, changes in the rate of tail current decay cannot be regarded as reliable indices of changes in the pyrethroid sensitivity of insect sodium channels.…”

“…Heterologous expression of insect voltage-sensitive sodium channels in Xenopus oocytes has been utilized to determine the functional characteristics and pharmacological significance of kdr-associated point mutations (Lee et al, 2000a;Feng et al, 1995;Smith et al, 1997;Warmke et al, 1997;Tan et al, 2002b;Smith et al, 1998;Zhao et al, 2000;Vais et al, 2000;Lee et al, 1999b). Studies with the wildtype house fly Vssc1 channel coexpressed either with the Drosophila melanogaster tipE auxiliary subunit (Vssc1 WT /tipE channels) or the house fly Vsscβ auxiliary subunit (Vssc1 WT /Vsscβ channels) in oocytes and with mutated Vssc1 variants carrying the kdr-associated L1014F, M918T and V410M mutations demonstrated that the single (L1014F) or double (L1014F and M918T) kdr-type mutations significantly reduced or abolished pyrethroid sensitivity (Smith et al, 1997;Lee et al, 1999b;Lee and Soderlund, 2001).…”

The Sophisticated Peptide Chemistry of Venomous Animals as a Source of Novel Insecticides Acting on Voltage-Gated Sodium Channels

“…Up to date, the full length cDNA clones for only 3 orthologous para genes have been identified: in the housefly Musca domestica (Vssc1), the German cockroach Blattella germanica (BgNa V ) and the mite Varroa destructor (VmNa V ) (Dong, 1997;Ingles et al, 1996;Wang et al, 2003). All 3 genes generate voltage-dependent sodium currents when heterologously expressed in oocytes (Du et al, 2009;Ingles et al, 1996;Tan et al, 2002b). A functional characterization of 20 BgNa V splice variants demonstrated that, similar to para, RNA editing and alternative splicing results in an astonishing diversity of Na V channels with distinct expression levels and varying kinetics of gating.…”