In recent years there has been growing evidence that all organisms and the environment are exposed to hormone-like chemicals, known as endocrine disruptor chemicals (EDCs). These chemicals may alter the normal balance of endocrine systems and lead to adverse effects, as well as an increasing number of hormonal disorders in the human population or disturbed growth and reduced reproduction in the wildlife species. For some EDCs, there are documented health effects and restrictions on their use. However, for most of them, there is still no scientific evidence in this sense. In order to verify potential endocrine effects of a chemical in the full organism, we need to test it in appropriate model systems, as well as in the fruit fly, Drosophila melanogaster. Here we report detailed in vivo protocols to study endocrine disruption in Drosophila, addressing EDC effects on the fecundity/fertility, developmental timing, and lifespan of the fly. In the last few years, we used these Drosophila life traits to investigate the effects of exposure to 17-α-ethinylestradiol (EE2), bisphenol A (BPA), and bisphenol AF (BPA F). Altogether, these assays covered all Drosophila life stages and made it possible to evaluate endocrine disruption in all hormone-mediated processes. Fecundity/fertility and developmental timing assays were useful to measure the EDC impact on the fly reproductive performance and on developmental stages, respectively. Finally, the lifespan assay involved chronic EDC exposures to adults and measured their survivorship. However, these life traits can also be influenced by several experimental factors that had to be carefully controlled. So, in this work, we suggest a series of procedures we have optimized for the right outcome of these assays. These methods allow scientists to establish endocrine disruption for any EDC or for a mixture of different EDCs in Drosophila, although to identify the endocrine mechanism responsible for the effect, further essays could be needed.
The rDNA magnification process consists of a rapid and inheritable rDNA increase occurring in bobbed males: in a few generations the bb loci acquire the wild-type rDNA value and reach a bb+ phenotype.-We have analyzed the rDNA magnification process in the repair-recombination-deficient mutant mei9, both at the phenotypical and rDNA content levels. In mei9 bb double mutants the recovery of bb+ phenotype is strongly disturbed and the rDNA redundancy value fails to reach the wild-type level. The strong effect of this meiotic mutation on rDNA magnification suggests a close relationship between this phenomenon and the repair-recombination processes. HE bobbed (bb) locus of D. melanogaster maps proximal to the centromeres T on the X chromosome and on the Y short arm. Wild-type (bb+) loci consist of a cluster of 150-250 tandemly repeated genes for ribosomal RNA (for a review see RITOSSA 1976). The rRNA genes have an additive effect, that is the phenotype depends on the total number of the genes carried by both the X and Y chromosomes. The bb phenotype reduction of thickness and length of bristles, etched abdominal cuticle, delayed hatching time (LINDSLEY and GRELL 1968) arises when the diploid (i.e., X plus X, or X plus Y) number of rDNA repeats are below a certain value, usually about half of the stock rDNA content. Bobbed loci show a remarkable random instability of redundancy, and several mechanisms capable of changing rRNA gene multiplicity have been demonstrated. In addition to meiotic unequal crossing-over within the bobbed loci (SCHALET 1969), somatic gene compensation (TARTOF 1971; SPEAR and GALL 1973) and rDNA magnification (RITOSSA 1968; BONCINELLI et al. 1972; MALVA et al. 1972) have been described. Magnification occurs in males of bobbed phenotype with several possible genotypes: Xbb/Ybb-; Xbb/O; XNo-/Ybb; =bb/O; Xbb/Ybb. When an X chromosome carrying a partial or a total deletion of the bb locus is kept in combination with a Y chromosome also deleted in rDNA, a sudden and heritable increase of the rRNA gene number is observed. Males of this first generation, called premagnified (pre), still exhibit a strong bobbed phenotype with respect to their level of rDNA redundancy. This observation and other data (RITOSSA et al. 1971; GRAZIANI and GARGANO 1976) suggest that the new rDNA copies synthesized at this stage are not fully functional. If premagnified males are backcrossed with suitable females in order to maintain the same
ABSTRACT The rDNA magnification process consists of a rapid and inheritable rDNA increase occurring in bobbed males: in a few generations the bb loci acquire the wild-type rDNA value and reach a bb+ phenotype.—We have analyzed the rDNA magnification process in the repair-recombination-deficient mutant mei9a, both at the phenotypical and rDNA content levels. In mei9a bb double mutants the recovery of bb+ phenotype is strongly disturbed and the rDNA redundancy value fails to reach the wild-type level. The strong effect of this meiotic mutation on rDNA magnification suggests a close relationship between this phenomenon and the repair-recombination processes.
Abstract We report the characterization of two novel genes of Drosophila melanogaster , named mst36Fa and mst36Fb . They define a novel gene family, showing identical time and tissue‐specificity limited to male germ cells where their transcription starts during meiotic prophase. These two genes encode for two slightly basic proteins highly homologous to each other and fairly rich in leucine and glutamic acid. Although strictly clustered, these genes utilize different promoter regions as revealed by examination of transgenic flies bearing mst36F‐promoter‐lacZ reporter constructs and by reverse transcription–polymerase chain reaction assays. Our data suggest that at least one gene ( mst36Fa ) of the cluster is under translational repression until spermiogenesis suggesting a putative role in the spermatides differentiation. The present study is aimed at the structural analysis of these genes.
In recent years there has been growing evidence that all organisms and the environment are exposed to hormone-like chemicals, known as endocrine disruptor chemicals (EDCs). These chemicals may alter the normal balance of endocrine systems and lead to adverse effects, as well as an increasing number of hormonal disorders in the human population or disturbed growth and reduced reproduction in the wildlife species. For some EDCs, there are documented health effects and restrictions on their use. However, for most of them, there is still no scientific evidence in this sense. In order to verify potential endocrine effects of a chemical in the full organism, we need to test it in appropriate model systems, as well as in the fruit fly, Drosophila melanogaster. Here we report detailed in vivo protocols to study endocrine disruption in Drosophila, addressing EDC effects on the fecundity/fertility, developmental timing, and lifespan of the fly. In the last few years, we used these Drosophila life traits to investigate the effects of exposure to 17-α-ethinylestradiol (EE2), bisphenol A (BPA), and bisphenol AF (BPA F). Altogether, these assays covered all Drosophila life stages and made it possible to evaluate endocrine disruption in all hormone-mediated processes. Fecundity/fertility and developmental timing assays were useful to measure the EDC impact on the fly reproductive performance and on developmental stages, respectively. Finally, the lifespan assay involved chronic EDC exposures to adults and measured their survivorship. However, these life traits can also be influenced by several experimental factors that had to be carefully controlled. So, in this work, we suggest a series of procedures we have optimized for the right outcome of these assays. These methods allow scientists to establish endocrine disruption for any EDC or for a mixture of different EDCs in Drosophila, although to identify the endocrine mechanism responsible for the effect, further essays could be needed.
Abstract mst36Fa and mst36Fb are two male‐specific genes that are part of a novel gene family recently characterized in Drosophila melanogaster . The genes are strictly clustered and show an identical tissue and temporal expression pattern limited to the male germline. Here we demonstrate that the transcription of these two genes, which is triggered by different cis regulatory elements, responds to the same testis‐specific factors encoded by the aly and can class meiotic arrest genes. RNA interference was used to decrease expression of these two genes. We obtained a reduction of fertility in the transgenic adult males compared to the wild type. These data suggest that the Mst36Fa and Mst36Fb proteins may have an important role in the production of functional sperm.
The intersex (ix) gene works in concert with doublesex (dsx) at the bottom of the sex-determination hierarchy to control somatic sexual differentiation in Drosophila melanogaster females. Here we report the isolation and characterization of the Drosophila intersex (ix) homologue in the pest lepidopteron Maruca vitrata (Mvix). The Mvix gene exhibits major complexity with respect to the Drosophila homolog. It is expressed in males and females and its pre-mRNA is subject to differential splicing events which affect both the protein coding and the non-coding regions. Moreover, Northern blot experiments revealed the presence of a female-specific transcript in pupae RNA, which appears to be the first described sex specific transcript of ix homologs characterized to date. The expression of Mvix cDNA in D.melanogaster transgenic flies indicates that the MvIX product, which shares a relatively high degree of homology with the D.melanogaster IX protein, is able to partially rescues the Drosophila mutant phenotype.
