Project description:Species often produce sterile hybrids early in their evolutionary divergence, and some evidence suggests that hybrid sterility may be associated with deviations or disruptions in gene expression. In support of this idea, many studies have shown that a high proportion of male-biased genes are underexpressed compared to non-sex-biased genes in sterile F1 male hybrids of Drosophila species. In this study, we examined and compared patterns of misexpression in F1 hybrid male third instar larvae of Drosophila simulans and its sibling species, D. sechellia. We analyzed hybrids using custom cDNA arrays we developed from RT-PCRs of spermatogenesis-related transcripts from these species and another sibling species (D. mauritiana). The results from a commercial genome-wide array and custom chip for adults of this species pair, from the custom chip and the genome-wide chip for adults of the D. simulans-D. mauritiana species pair, and from the larvae of the D. simulans-D. mauritiana species pair, are presented separately. Keywords: Comparison of pure-species Drosophila expression to hybrid expression in larvae
Project description:CP190 is a Drosophila protein of unknown function that exhibits cell cycle-specific localisation to the centrosome during mitosis and the nucleus during the interphase. CP190 associates with multiple sites on polytene chromosomes and its predicted amino acid sequence identifies four C2h2 zinc finger motifs along with an N-terminal BTB/POZ domain (similar to GAGA factor). Experimental studies have identified a 128 amino acid domain that direct centrosome association and a bipartite NLS. Interest in the function of CP190 was initially driven by an interest in centrosome function. Is is also fascinating to know why a protein that forms a core component of the centrosome also shares the attributes and distribution expected of a regulator of gene expression. CP190 mutants (EMS generated) were recently isolated whose recessive lethal phenotype indicates that CP190 is essential. The two best-characterised isolates, CP190[1] and CP190[2], are either nulls or severe hypomorphs (CP190 protein is undetectable on Western blots of total protein from homozygous mutatant third instar larvae). Both mutants can be rescued as homozygotes by expression of wild type CP190 from a transgenic second chromosome carrying a CP190 cDNA under the control of the polyubiquitin promoter. Stocks homozygous or hemizygous for either of these mutant alleles die as pharate adults that show no obvious externally visible defects. Mutant larvae show a slight developmental delay compared to their heterozygous siblings, but their brains and discs are of normal size and brain squash preparation show no obvious mitotic phenotype. Since CP190 is a chromatin-associated protein and has motifs that are typical of proteins involved in regulating gene expression, it seems likely that the gene expression profile of CP190 mutants will be aberrant. This experiment hopes to identify genes that exhibit aberrant expression in the mutant compared to the wild type. Target RNA were isolated from: homozygous CP190[1] 0 hour white prepupae (non tubby, red malpighian tubules) homozygous CP190[2] 0 hour white prepupae (non tubby, red malpighian tubules) homozygous red[1] e[1] 0 h white prepupae (source from which the mutated chromosome was obtained) For each CP190 mutant, total RNA preparations were made from three separate groups of 100 prepupae selected from independent cultures, similarly, six independent RNA samples were prepared from red[1] e[1] stock as 'wild type' controls, giving a total of 12 target RNA samples. Prepupae were collected from bottles every 30 min and frozen directly in liquid N2. Frozen samples were pooled and ground to a powder in liquid N2 before resuspension in TRIZOL and shipment to Flychip facility at the University of Cambridge, UK.
Project description:Species often produce sterile hybrids early in their evolutionary divergence, and some evidence suggests that hybrid sterility may be associated with deviations or disruptions in gene expression. In support of this idea, many studies have shown that a high proportion of male-biased genes are underexpressed compared to non-sex-biased genes in sterile F1 male hybrids of Drosophila species. In this study, we examined and compared patterns of misexpression in F1 hybrid male third instar larvae of Drosophila simulans and its sibling species, D. mauritiana. We analyzed hybrids using custom cDNA arrays we developed from RT-PCRs of spermatogenesis-related transcripts from these species and another sibling species (D. sechellia). The results from a commercial genome-wide array and custom chip for adults of this species pair, from the custom chip and the genome-wide chip for adults of the D. simulans-D. sechellia species pair, and from the larvae of the D. simulans-D. sechellia species pair, are presented separately. Keywords: Comparison of pure-species Drosophila expression to hybrid expression
Project description:We used whole larvae of both wildtype and Nxt1 trans-heterozygotes for RNA paired-end sequencing and obtained a read depth between 6.4M to 11.1M reads across all samples. In wandering larvae, stationary larvae and white prepupae, a total of 435, 855, and 611 genes were differentially expressed. A high percentage of down-regulated genes were highly expressed/testis-specific in the Drosophila testis.