The transformer2 gene of the pumpkin fruit fly, Bactrocera tau (Walker), functions in sex determination, male fertility and testis development

Abstract

The insect transformer2 (tra2) gene has a prevalent role in cooperating with the sex-determining gene transformer (tra) to direct female differentiation. Here, we report the identification and characterization of Btau-tra2, the tra2 orthologue of the pumpkin fruit fly, Bactrocera tau, an invasive agricultural pest. The Btau-tra2 gene produces three transcript variants. However, only two transcripts can be examined; one is present at all developmental stages in the soma and germline of both sexes and the other one is specific to the embryo and the germline. Knocking down the function of Btau-tra2 produced a male-biased sex ratio and some intersexes. Consistent with a role in sex determination, the obtained intersexual and male sterility phenotypes express a mix of male and female splice variants of the tra and doublesex (dsx) orthologues, indicating that Btau-tra2 has a conserved splicing regulatory function and acts together with/upstream of tra and dsx. In addition, some males obtained from the knock down are fertile but their fertilities are extremely reduced. Moreover, almost all surviving RNA interference (RNAi) males harbour testes having some defects in their external morphologies. Most notably, the body size of a few surviving RNAi flies was two-to threefold increased with respect to the normal size. Our findings suggest that Btau-tra2 is involved in male fertility and may also have an unprecedented role in body size control besides its conserved role in sex determination.

Male-biased Adult Production of the Striped Fruit Fly, Zeugodacus scutellata, by Feeding dsRNA Specific to Transformer-2

“…In tephritid fruit flies, mRNA variants of Tra-2 have been reported. For example, three mRNA variants of Tra-2 have been identified in Z. tau, of which at least one is expressed in both sexes and the other is specific to the embryo or germline [21]. Although Zs-Tra2 mRNA variants were not assessed in our current study, its expression profile in males suggested the presence of variants of the Zs-Tra2 transcript.…”

“…In B. dorsalis, nearly 100% males were obtained with embryonic RNAi against Tra-2 [20]. In Z. tau, dsRNA injection to embryo has produced 72.4% males while the control has produced only 38.6% males [21]. This indicated an increase of 187.6% in male production.…”

“…They play crucial roles in the regulation of female-specific splicing of Dsx pre-mRNA. These include 12 Anastrepha species [37,38], Bactrocera oleae [39], B. tryoni, B. jarvisi [40], B. dorsalis [20], B. tau [21], and C. capitata [19]. The knocking down of Tra-2 expression in these tephritid fruit flies can interrupt the splicing pattern of Dsx pre-mRNA of females to differentiate females into male phenotypes [41,42].…”

“…When this RNAi technique against Tra-2 expression was applied to eggs of the Oriental fruit fly, Bactrocera dorsalis, it produced almost 100% males [20]. Similar approach has been applied to the pumpkin fruit fly (Z. tau), resulting in male-biased sex ratio and some intersexes [21]. However, egg injections of double-strand RNA are a practical limitation to produce males only in mass-rearing for SIT.…”

Identification and characterization of doublesex from the pumpkin fruit fly, Bactrocera tau (Diptera: Tephritidae)

“…The presence in unfertilized eggs of only the Btau-dsx F transcript sug-gests that Btau-dsx pre-mRNA may be a maternal substrate transferred to the eggs during oogenesis. Since we also detected the maternal transformer female-specifi c (Btautra F ) transcript (our unpublished data) and the maternal transformer2 (Btau-tra2) transcript (Thongsaiklaing et al, 2018) in early laid unfertilized and fertilized eggs, it is likely that in B. tau female embryos and in unfertilized eggs, the maternal Btau-dsx pre-mRNAs may be spliced into the female mode upon activation of the maternal Btau-TRA/Btau-TRA2 protein complex. In early fertilized eggs, the Btau-dsx M1 transcript was also detected, albeit at low levels, indicating that the Btau-dsx M1 transcript may exist only in male embryos.…”