Two female-specific DSX proteins are encoded by the sex-specific transcripts of dsx, and are required for female sexual differentiation in two wild silkmoth species, Antheraea assama and Antheraea mylitta (Lepidoptera, Saturniidae)

• Published in: Insect biochemistry and molecular biology Vol. 40; no. 9; pp. 672 - 682
• Main Authors: Shukla, J.N., Nagaraju, J.
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01-09-2010
• Subjects: Alternative Splicing; Animals; Antheraea; Antheraea assama; Antheraea mylitta; Conserved Sequence; doublesex gene; Exons; Female; females; gender differences; Gene Dosage; gene expression; gene expression regulation; gene silencing; genes; genome; genomics; Insect Proteins - genetics; Insect Proteins - metabolism; Insect Proteins - physiology; males; messenger RNA; molecular sequence data; moths; Moths - genetics; Moths - metabolism; Moths - physiology; OD1 domain; OD2 domain; open reading frames; RNA Interference; RNA, Messenger - metabolism; RNA, Messenger - physiology; Sequence Alignment; sequence analysis; Sequence Analysis, RNA; sex determination; Sex Determination Processes; transcription (genetics); transcriptome; transcriptomics; Alternative splicing; OD2 domain; Antheraea assama; Antheraea mylitta; OD1 domain
• ISSN: 0965-1748; 1879-0240
• DOI: 10.1016/j.ibmb.2010.06.008

doublesex (
dsx) is the bottom most gene of the sex-determination cascade of
Drosophila melanogaster. The pre-mRNA of
dsx splices to produce male- and female-specific transcripts which code for the male- and female-specific proteins, respectively.
dsx homologues have been characterized from different (many in Diptera, two in Hypmenoptera and only one in Lepidoptera) insect species. Sex-specific splice forms of
dsx pre-mRNA in all these species code for one male- and one female-specific DSX proteins, which regulate the downstream target genes responsible for sex-specific characters. In the present study we have cloned and characterized the
dsx homologues from two saturniid silkmoths,
Antheraea assama and
Antheraea mylitta. The divergence time between Saturniidae and Bombycidae to which the domesticated silkworm,
Bombyx mori belongs is estimated to be around 160.9 MY. Interestingly, the
dsx pre-mRNA of these wild silkmoths sex-specifically splices to generate multiple splice variants. On the basis of their open reading frame (ORF) and conceptual translation, two female-specific (DSX
F1 and DSX
F2) and one male-specific (DSX
M) proteins could be inferred, in both the moths. Presence or absence of a 15 bp stretch within the ORF of the two groups of female-specific transcripts resulted in the production of two distinct female-specific DSX proteins. The sex-specific DSX proteins have common amino-terminal sequence but sex-specific carboxy termini. The two female-specific DSX proteins (DSX
F1 and DSX
F2) share common DNA binding domain (DM domain) and oligomerization domain (OD domain) and differ only at their extreme C-termini by 21aa. Functional analysis of
dsx transcripts in
A. assama by
dsRNA mediated knock-down resulted in complete abolition of expression of
vitellogenin and
hexamerin genes, the direct targets of the DSX proteins, irregular differentiation of gonads, and drastic reduction in fecundity and hatchability. Together, these results suggest the involvement of both the female-specific DSX proteins in the process of female sexual differentiation. Further, conservation of the 4th exon sequence, especially the PESS sequence responsible for the sex-specific splicing of
Bmdsx in the female-specific transcripts of
Aadsx and
Amydsx, indicated the existence of a common mechanism of sex-specific splicing of
dsx homologues in silkmoths. To our knowledge this is the first report of existence of multiple splice forms of
dsx pre-mRNA encoding two female-specific DSX proteins.
► The study reports doublesex (dsx) homologues of the sex determination cascade from two wild saturniid silkmoths,
Antheraea mylitta and
Antheraea assama. ► The results suggest that unlike in other insects, in silkmoths dsx premRNA splices sex-specifically to generate multiple splice-variants. ► On the basis of ORFand coceptual translation, two female specific (DSXF1 and DSXF2) and one male-specific (DSXM) could be inferred in both the moths. ► The two female-specific DSX proteins differ at their extreme C-termini by 21aa. ► RNAi analysis suggests that both the female DSX proteins are essential for female sexual differentiation.