The Drosophila gene fs(2)cup interacts with otu to define a cytoplasmic pathway required for the structure and function of germ-line chromosomes.

The Drosophila ovarian tumor gene (otu) encodes cytoplasmic proteins that are required in germ-line cells for cyst formation, nurse cell chromosome structure and egg maturation. We have analyzed a gene, fs(2)cup, that participates in many of the same processes and interacts with otu genetically. Both nurse cell and oocyte chromosomes require cup to attain a normal morphology. In addition, the gene is needed for the oocyte to grow normally by taking up materials transported from the nurse cells. The gene encodes a 1132-amino-acid protein containing a putative membrane-spanning domain. Cup protein (but not cup RNA) is transported selectively into the oocyte in germarial cysts, like the p104 Otu protein. It is strongly associated with large structures in the cytoplasm and perinuclear region of nurse cells and, like Otu, moves to the periphery of these cells in stages 9-10. Moreover, cup mutations dominantly disrupt meiotic chromosome segregation. We propose that cup, otu and another interacting gene, fs(2)B, take part in a common cytoplasmic pathway with multiple functions during oogenesis.

Multiple response elements in the Sex-lethal early promoter ensure its female-specific expression pattern.

The choice of sexual identity in somatic tissues of the fruit fly Drosophila melanogaster is determined early in embryogenesis by the X-chromosome-to-autosome (X/A) ratio. The system that signals the X/A ratio selects the sexual development pathway by determining the activity state of the binary switch Sex-lethal (Sxl). In 2X/2A animals, the X/A signalling system turns the Sxl gene on, ultimately activating an RNA-splicing autoregulatory feedback loop which serves to maintain the female state during the remainder of development. In 1X/2A animals, this autoregulatory feedback loop is not activated and the male state is subsequently maintained by the default splicing machinery. In the studies reported here, we have examined how the X/A signalling system controls the initial choice of sexual identity through its action on a special early embryonic Sxl promoter, Sxl-Pe. We show that in the early embryo, the activity of Sxl-Pe is controlled in a highly dose-sensitive fashion by the genes on the X chromosome that function as numerator elements and by genes located on the autosomes that function as denominator elements. Functional dissection of Sxl-Pe indicates that activating the promoter in females requires the cumulative action of multiple numerator genes which appear to exert their effects through reiterated cis-acting target sites in the promoter. Conversely, maintaining the promoter silent in males requires the repressive activities of denominator genes, and at least one of the denominator genes also appears to function through target sequences within the promoter.

The primary sex determination signal of Drosophila acts at the level of transcription.

For Drosophila, the choice between male and female development is made by the switch gene, Sxl, in response to the X:A ratio. Once Sxl is turned on in females, it actively maintains the determined state, independent of the X:A signal, by a positive autoregulatory feedback loop in which Sxl proteins direct the female-specific splicing of Sxl transcripts. In this paper we have investigated the mechanism controlling pathway initiation. Our results suggest a two-step model for the initial activation of Sxl in females. In the first step, a special class of Sxl mRNAs is expressed in female embryos from an early promoter that responds to the genes signaling the X:A ratio. The proteins produced from these early mRNAs then initiate the autoregulatory loop by directing the female-specific processing of transcripts from the late Sxl promoter.

The Drosophila female-specific sex-determination gene, Sex-lethal, has stage-, tissue-, and sex-specific RNAs suggesting multiple modes of regulation.

For proper sexual development of females, the Sex-lethal (Sxl) gene must be activated early in development and remain on during the rest of the life cycle. Conversely, in males, Sxl must remain functionally off through development. Here, we show that the Sxl transcription unit spans a DNA segment of greater than 20 kb and encodes at least 10 distinct, but overlapping, RNA species. These RNAs range in size from 4.4 to 1.7 kb and exhibit sex, stage, and tissue specificity. Six RNAs, three female specific and three male specific, are first detected by midembryogenesis and persist through the adult stage: Their expression reflects the on/off regulation of Sxl's activity at the level of sex-specific alternate splicing. Four Sxl RNAs are found in adult females. Two of these RNAs are dependent on the presence of a functional germ line and may be relevant to Sxl's role in adult germ-line development. All four are present in unfertilized eggs. Finally, three Sxl RNAs are found only transiently during very early embryogenesis; we suggest that the expression of these RNAs may reflect an early regulation of Sxl at the level of transcription and that these transcripts are involved in the initial selection of the Sxl activity state in response to the primary sex-determination signal, the X/A ratio.