Transformer (tra) is the second gene of a regulatory cascade based on RNA splicing that determines sex in Drosophila melanogaster. Splicing of tra transcripts is regulated by the master gene Sex lethal and tra itself regulates splicing of the transcriptional regulator doublesex (dsx). We present the isolation and characterization of Botra, the olive fruit fly Bactrocera oleae orthologue to the Drosophila gene transformer. As in Drosophila, Botra transcripts are spliced in a sex-specific manner so that only females encode a functional polypeptide of 422 amino acids, whereas males encode presumably nonfunctional peptide(s). The identification of multiple TRA/TRA-2 binding sites within the Botra male-specific exons, suggests an autoregulation mechanism of tra, through TRA/TRA2 activities. The fundamental role of the TRA protein in sex determination of Bactrocera was investigated by RNA interference, where the introduction of Botra dsRNA into embryos resulted in complete transformation of XX flies into fertile males.

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We hybridized cloned DNA segments to salivary gland polytene chromosomes of the medfly, Ceratitis capitata, and thus established molecular markers for 24 sites on 6 out of 10 autosomal arms. An additional marker identified a medfly repetitive element that hybridizes to approximately 100 autosomal sites as well as a granular network that is thought to represent the X chromosome. Some of the markers correspond to 9 characterized transcription units, while 17 remain anonymous; at least 3 of the latter are restriction fragment length polymorphism (RFLP) markers. The characterized transcription units document that chromosomal arm 5L of C. capitata is homologous to the Drosophila melanogaster X chromosome, in agreement with previous inferences based on the extensive conservation of linkage groups in Diptera.

Two recessive female-sterile mutants, K451 and K1214, disrupt chorion formation by causing underproduction of all major chorion proteins. We present evidence that this effect is due to underaccumulation of the chorion mRNAs and that, in turn, this is caused by a substantial reduction in the level of chorion gene amplification. The mutants are X-linked but located at two sites far from the chorion gene cluster at 7F1-2; their effect is even more pronounced on the third chromosome chorion gene cluster, and thus the wild type gene must act in trans. The time course of amplification in mutant and wild-type follicles is documented.Formation of the chorion in Drosophila melanogaster is a precisely regulated developmental program that takes place in the late stages of oogenesis. During a period of only 5 hr, the various chorion layers are successively laid down by the highly specialized follicular epithelium that envelops the oocyte. Approximately 20 proteins have thus far been identified as chorion components, of which 6 are quite abundant (1, 2). The "major" genes encoding these abundant proteins are tightly clustered into two distinct chromosomal loci. The X chromosome locus at 7F1-2 includes two major chorion genes, s36-1 and s38-J, which are expressed in the early stages of choriogenesis, plus several additional minor genes (refs. 3-5; unpublished observations). The other four major chorion genes, sl5-i, sf6-1, sl8-1, and sf9-1, which are expressed late in choriogenesis albeit not in a strictly coordinate fashion, are clustered on the third chromosome at 66D12-15 (5-8).Both of these loci are differentially replicated in the follicular cells (9). Tissue-specific amplification begins by stage 9, at least 5 hr prior to the onset of choriogenesis, and has the effect of increasing the copy number of the major genes at a time when rapid chorion protein synthesis is required. Gene amplification is apparently needed for production of the normal, massive amounts of chorion proteins, although the expression of individual genes is subject to further temporal and quantitative regulation (8). In the chromosome each chorion gene cluster resides in a 50-to 100-kilobase (kb) amplification domain; maximal amplification occurs in the center of the domain, where the chorion genes are located (5). In the mutant, ocelliless, a small inversion with a breakpoint within the X chromosome cluster disrupts the amplification of that cluster (10). Apparently as a result, the s36-1 and s38-1 proteins are underproduced, an abnormal chorion is formed, and the embryos fail to develop (3, 11). In summary, amplification is itself a developmentally regulated phenomenon, an important subroutine within the overall developmental program of choriogenesis.We have begun a genetic analysis in an effort to dissect and identify the various steps in the program of choriogenesis. cis-acting mutants (like ocelliless) that map in the vicinity of the chorion genes can be expected to help identify regulatory DNA sequences, such as the origins of amplificat...

Male-specific serum proteins (MSSPs) are low molecular weight proteins which accumulate in high amounts in the haemolymph of adult males of the medfly Ceratitis capitata. By screening an expression library with anti-MSSP antibodies, we have isolated and determined the nucleotide sequence of a cDNA clone coding for one of the male-specific polypeptides (MSSP-alpha). The MSSP-alpha mRNA encodes a polypeptide of 144 amino acids with a secretory signal sequence of sixteen amino acids. Southern analysis indicated that there are multiple copies of MSSP genes in the medfly genome. Northern analysis showed that the MSSP mRNAs are synthesized only in adult males. The accumulation pattern of these mRNAs during development suggests that the expression of the MSSP genes is developmentally regulated at both transcriptional and translational levels. The predicted peptide sequence of MSSP-alpha shows significant similarity to a group of pheromone- and general odourant-binding proteins of insects.