Project description:In Drosophila, two chromosome-wide compensatory systems have been characterized; the dosage compensation system acting on the male X-chromosome and the chromosome specific regulation of genes located on the heterochromatic 4th chromosome. Dosage compensation in Drosophila is accomplished by hypertranscription of the single male X-chromosome mediated by the MSL-complex. The mechanism for this compensation is suggested to be an MSL-complex mediated enhanced transcriptional elongation while the mechanism for the compensation mediated by Painting of fourth (POF) on the 4th chromosome has remained elusive. Here we show that POF binds to nascent RNA and this binding is associated with an increase in amount of chromosome 4 transcripts. Furthermore, genes located on the 4th chromosome are enriched in binding of the nucleoplasmic nucleporin component NUP98 and this enrichment correlates to increased POF binding. We also show that genes located in heterochromatic regions have a shorter transition time from site of transcription and to the nuclear envelope. Our current work broadens the understanding about how genes in heterochromatic regions can overcome the repressive influence of their hostile environment.
Project description:Drosophila X chromosomes are subject to dosage compensation in males and are known to have a specialized chromatin structure in the male soma. We are interested in how specific chromatin structure change contributes to X chromosome hyperactivity and dosage compensation. We have conducted a global analysis of localize two dosage compensation complex dependent histone marks H4AcK16 and H3PS10 and one dosage compensation complex independent histone mark H3diMeK4 in the genome, especially on X chromosome by ChIP-chip approach in both male and female adult flies. We also probed general genomewide chromatin structure by deep DNA sequencing of sheared ChIP input DNA from male and female adult flies.
Project description:In Drosophila, two chromosome-wide compensatory systems have been characterized; the dosage compensation system acting on the male X-chromosome and the chromosome specific regulation of genes located on the heterochromatic 4th chromosome. Dosage compensation in Drosophila is accomplished by hypertranscription of the single male X-chromosome mediated by the MSL-complex. The mechanism for this compensation is suggested to be an MSL-complex mediated enhanced transcriptional elongation while the mechanism for the compensation mediated by Painting of fourth (POF) on the 4th chromosome has remained elusive. Here we show that POF binds to nascent RNA and this binding is associated with an increase in amount of chromosome 4 transcripts. Furthermore, genes located on the 4th chromosome are enriched in binding of the nucleoplasmic nucleporin component NUP98 and this enrichment correlates to increased POF binding. We also show that genes located in heterochromatic regions have a shorter transition time from site of transcription and to the nuclear envelope. Our current work broadens the understanding about how genes in heterochromatic regions can overcome the repressive influence of their hostile environment. Pof mutant vs. wild type, 3 replicates
Project description:In Drosophila, two chromosome-wide compensatory systems have been characterized; the dosage compensation system acting on the male X-chromosome and the chromosome specific regulation of genes located on the heterochromatic 4th chromosome. Dosage compensation in Drosophila is accomplished by hypertranscription of the single male X-chromosome mediated by the MSL-complex. The mechanism for this compensation is suggested to be an MSL-complex mediated enhanced transcriptional elongation while the mechanism for the compensation mediated by Painting of fourth (POF) on the 4th chromosome has remained elusive. Here we show that POF binds to nascent RNA and this binding is associated with an increase in amount of chromosome 4 transcripts. Furthermore, genes located on the 4th chromosome are enriched in binding of the nucleoplasmic nucleporin component NUP98 and this enrichment correlates to increased POF binding. We also show that genes located in heterochromatic regions have a shorter transition time from site of transcription and to the nuclear envelope. Our current work broadens the understanding about how genes in heterochromatic regions can overcome the repressive influence of their hostile environment.
Project description:Drosophila X chromosomes are subject to dosage compensation in males and are known to have a specialized chromatin structure in the male soma. We are interested in how specific chromatin structure change contributes to X chromosome hyperactivity and dosage compensation. We have conducted a global analysis of localize two dosage compensation complex dependent histone marks H4AcK16 and H3PS10 and one dosage compensation complex independent histone mark H3diMeK4 in the genome, especially on X chromosome by ChIP-chip approach in both male and female adult flies. We also probed general genomewide chromatin structure by deep DNA sequencing of sheared ChIP input DNA from male and female adult flies. Chromatin immunoprecipitations were performed in 5-7 day aged adult male and female flies with three histone modification antibodies. ChIP enriched DNA and input DNA was labeled by Cy3 or Cy5 dye separately and hybridized simultaneously to the Drosophila FlyGEM arrays. At least two biological replicates were performed for each antibody and sex. DNA-seq (NIDDK-Drosophila-Illumina-DNASeq) were performed on ChIP-input sheared DNA to check the general chromatin structure of different chromosome.
Project description:Chromosomal and segmental aneuploidies are usually lethal or common features of cancer cells and variety of disease, but little is known about how copy number relates to gene expression. Drosophila males have a single X chromosome and two sets of autosomes, but X chromosome and autosome transcripts are equally abundant. This 2-fold increase in X chromosome gene expression requires a dosage compensation complex (MSL) that acetylates histone 4 at lysine 16 (H4AcK16). To determine the contribution of general buffering and MSL to X chromosome dosage compensation, we analyzed genome-wide copy number, expression and histone modification pattern in male Drosophila S2 cells with and without MSL. Keywords: Gene Regulation Study, genomic analyses
Project description:The essential process of dosage compensation, which corrects for the imbalance in X-linked gene expression between XX females and XY males, represents a key model for how genes are targeted for coordinated regulation. However, the mechanism by which dosage compensation complexes identify the X-chromosome during early development remained unknown because of the difficulty of sexing embryos prior to zygotic transcription. We used meiotic drive to sex Drosophila embryos prior to zygotic transcription and ChIP-seq to measure dynamics of dosage compensation factor targeting. The Drosophila Male-Specific Lethal dosage compensation complex (MSLc) requires the ubiquitous zinc-finger protein Chromatin-Linked Adaptor for MSL Proteins (CLAMP) to identify the X-chromosome. We observe a multi-stage process in which MSLc first identifies CLAMP binding sites throughout the genome followed by concentration at the strongest X-linked MSLc sites. We provide insight into the dynamic mechanism by which a large transcription complex identifies its binding sites during early development.