The Dominant Cold-SensitiveOut-ColdMutants ofDrosophila melanogasterHave Novel Missense Mutations in the Voltage-Gated Sodium Channel Geneparalytic

Abstract

Here we report the molecular characterization of Out-cold (Ocd) mutants of Drosophila melanogaster, which produce a dominant, X-linked, cold-sensitive paralytic phenotype. From its initial 1.5-Mb cytological location within 13F1-16A2, P-element and SNP mapping reduced the Ocd critical region to ,100 kb and to six candidate genes: hangover, CG9947, CG4420, eIF2a, Rbp2, and paralytic (para). Complementation testing with para null mutations strongly suggests Ocd and para are allelic, as does gene rescue of Ocd semilethality with a wild-type para transgene. Pesticide resistance and electrophysiological phenotypes of Ocd mutants support this conclusion. The para gene encodes a voltage-gated sodium channel. Sequencing the Ocd lines revealed mutations within highly conserved regions of the para coding sequence, in the transmembrane segment S6 of domain III (I1545M and T1551I), and in the linker between domains III and IV (G1571R), the location of the channel inactivation gate. The G1571R mutation is of particular interest as mutations of the orthologous residue (G1306) in the human skeletal muscle sodium channel gene SCN4A are associated with cases of periodic paralysis and myotonia, including the human cold-sensitive disorder paramyotonia congenita. The mechanisms by which sodium channel mutations cause cold sensitivity are not well understood. Therefore, in the absence of suitable vertebrate models, Ocd provides a system in which genetic, molecular, physiological, and behavioral tools can be exploited to determine mechanisms underlying sodium channel periodic paralyses. T HE molecular mechanisms underlying cold sensitivity in metazoans are not well understood, not least because relatively few studies have addressed this question. A few cold-sensitive mutations have been described in Drosophila melanogaster (Siddiqi and Benzer 1976;Sondergaard 1980;Lilly et al. 1994), but again the precise mechanisms by which these specific mutations lead to cold sensitivity have not been elucidated. The objective of this study was to identify the biological processes underlying cold sensitivity.The dominant cold-sensitive paralytic Out-cold (Ocd) mutations of D. melanogaster were originally isolated in a screen for X-linked male-lethal mutations (Sondergaard 1975(Sondergaard , 1979. Further analyses of the male-lethal stocks identified seven lines in which heterozygous mutant females exhibited a dominant cold-sensitive paralysis.

Mutation in the RmβAOR gene is associated with amitraz resistance in the cattle tick Rhipicephalus microplus

“…Such interaction has been found in mutant mosquitos, Culex pipiens, bearing the resistance gene Ace R , in which survival costs in these insects during winter were estimated to be in the order of 70% (22). In Drosophila melanogaster, mutations in the para sodium channel gene have been shown to cause cold-induced paralysis (23). In our study cold weather conditions were encountered between the Winter 2004 and Winter 2005 collection time points.…”

Footprints of positive selection associated with a mutation ( N1575Y ) in the voltage-gated sodium channel of Anopheles gambiae

“…In varroa mites, an L-P change at 1770 (L1596P housefly numbering) has been demonstrated to effect sensitivity to fluvalinate (a pyrethroid) when expressed in a Xenopus system (26). A G-R mutation at position 1575 in the inactivation particle (position 1559 housefly numbering) also underlies a cold-sensitive phenotype in Drosophila (27). Interestingly, the G1575R mutation occurs as a double mutation with I1545M (1533 in Musca) and the Drosophila Ocd strain carrying these two mutations are 1000-fold more resistant to DDT than the Oregon-R strain, the progenitor of Ocd (27) suggesting that one or both of these mutations are strongly involved in DDT resistance.…”

“…A G-R mutation at position 1575 in the inactivation particle (position 1559 housefly numbering) also underlies a cold-sensitive phenotype in Drosophila (27). Interestingly, the G1575R mutation occurs as a double mutation with I1545M (1533 in Musca) and the Drosophila Ocd strain carrying these two mutations are 1000-fold more resistant to DDT than the Oregon-R strain, the progenitor of Ocd (27) suggesting that one or both of these mutations are strongly involved in DDT resistance. I1545M lies in exon 30 and this was covered by our sequencing for the haplotype analysis.…”

Disruption of Endocytosis with the Dynamin Mutantshibirets1Suppresses Seizures inDrosophila

“…For example, in mice and humans, missense mutations of the Cacna1a calcium channel have been linked to absence seizures, while in Drosophila, mutations in the homologous cac TS2 channel result in heat-activated seizures but are also a suppressor of bang-sensitive mutants (Zamponi et al 2009). Additionally, there are different para alleles in Drosophila that can cause either bang sensitivity, seizure suppression, heat-activated seizure sensitivity, cold sensitivity, or temperature-sensitive paralysis, all extremes of either hyperor hypoactivity of the sodium channel (Lindsay et al 2008;Parker et al 2011b;Sun et al 2012;Schutte et al 2014).…”