Abstract
BackgroundThe doublesex gene controls somatic sexual differentiation of many metazoan species, including the malaria mosquito Anopheles gambiae and the dengue and yellow fever vector Aedes aegypti (Diptera: Culicidae). As in other studied dipteran dsx homologs, the gene maintains functionality via evolutionarily conserved protein domains and sex-specific alternative splicing. The upstream factors that regulate splicing of dsx and the manner in which they do so however remain variable even among closely related organisms. As the induction of sex ratio biases is a central mode of action in many emerging molecular insecticides, it is imperative to elucidate as much of the sex determination pathway as possible in the mosquito disease vectors.ResultsHere we report the full-length gene sequence of the doublesex gene in Culex quinquefasciatus (Cxqdsx) and its male and female-specific isoforms. Cxqdsx maintains characteristics possibly derived in the Culicinae and present in the Aedes aegypti dsx gene (Aeadsx) such as gain of exon 3b and the presence of Rbp1 cis-regulatory binding sites, and also retains presumably ancestral attributes present in Anopheles gambiae such as maintenance of a singular female-specific exon 5. Unlike in Aedes aegypti, we find no evidence for intron gain in the female transcript(s), yet recover a second female isoform generated via selection of an alternate splice donor. Utilizing next-gen sequence (NGS) data, we complete the Aeadsx gene model and identify a putative core promoter region in both Aeadsx and Cxqdsx. Also utilizing NGS data, we construct a full-length gene sequence for the dsx homolog of the northern house mosquito Culex pipiens form pipiens (Cxpipdsx). Analysis of peptide evolutionary rates between Cxqdsx and Cxpipdsx (both members of the Culex pipiens complex) shows the male-specific portion of the transcript to have evolved rapidly with respect to female-specific and common regions.ConclusionsAs in other studied insects, doublesex maintains sex-specific splicing and conserved doublesex/mab-3 domains in the mosquitoes Culex quinquefasciatus and Cx. pipiens. The cis-regulated splicing of Cxqdsx does not appear to follow either currently described mosquito model (for An. gambiae and Ae. aegypti); each of the three mosquito genera exhibit evidence of unique cis-regulatory mechanisms. The male-specific dsx terminus exhibits rapid peptide evolutionary rates, even among closely related sibling species.Electronic supplementary materialThe online version of this article (doi:10.1186/s12862-015-0386-1) contains supplementary material, which is available to authorized users.
Highlights
The doublesex gene controls somatic sexual differentiation of many metazoan species, including the malaria mosquito Anopheles gambiae and the dengue and yellow fever vector Aedes aegypti (Diptera: Culicidae)
Sequencing and genome alignment revealed this was due to the presence of a 75 bp (25 amino acid) alternatively spliced in-frame intronic sequence within exon 2 that was present in some transcripts but spliced out of others (Fig. 3)
An equivalent 63 bp (21 amino acid) tract was reported from Aedes aegypti dsx gene (Aeadsx) and a 72 bp (24 amino acid) tract reported in Angdsx [18], this appears to be specific to the Culicidae and has not been reported from sequenced dsx transcripts in other taxa
Summary
The doublesex gene controls somatic sexual differentiation of many metazoan species, including the malaria mosquito Anopheles gambiae and the dengue and yellow fever vector Aedes aegypti (Diptera: Culicidae). Most animals direct sex-specific cell fate by function of the Doublesex/Mab-3 Related Transcription factor (DMRT) family of zinc-finger proteins [4, 5] and the genes they regulate. Within the insects, this process involves a genetic cascade first elucidated in the model fly Drosophila melanogaster [6] whereby a primary signal triggers sexspecific splicing of one or more regulatory factors which subsequently bind pre-mRNA of the conserved DMRT “major switch” gene, doublesex, and direct its sex-specific splicing, initiating development of male or female forms [7]. The red flour beetle Tribolium castaneum Tcdsx has been implicated in oocyte development including Vitellogenins and their associated receptors [16], while Lepidopteran dsx has been shown to influence expression of pheromone-binding proteins and hexamerin storage proteins [17]
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