Project description:CGH analysis of translocations with breakpoints at the euchromatin/heterochromatin boundary. Three translocations with breakpoint at the euchromatin/heterochromatin boundary of 2L, 3L and X, respectively, were analyzed by CGH to distinguish heterochromatic sequences from euchromatic sequences. X: 101042(T(1;Y)B91); 2L: 130186 (T(Y;2)R146); and 3L: 102004(T(2;3)H31). To obtain embryos lacking the euchromatin portion of the chromosome arms, translocation males bearing breakpoint at the euchromatin/heterochromatin boundary of 2L, 3L and X were crossed to C(2)EN, C(3)EN or attached X females, respectively. All embryos were collected at room temperature.
Project description:We performed mRNA transcriptional profiling on 99 hemizygotic lines (Df/+) from the DrosDel project covering 68% of chromosome 2L, in order to understand how changes in gene copy number affect overall transcriptome.
Project description:Chromosomal instability which involves deletion and duplication of chromosomes or chromosome parts is a common feature of cancers, and deficiency screens are commonly used as a method to find genes involved in different biological pathways. Still, how gene expression from whole chromosomes or large chromosomal domains is affected by deficiencies, duplications or chromosome loss is largely unknown. Using expression microarrays of deficiency hemizygotes and a duplication hemizygote we show that expressed genes are significantly buffered when present in a deficiency hemizygote and that the buffering effect is general and not mainly caused by feedback regulation of individual genes. Differentially expressed genes are in general better buffered than ubiquitously expressed genes when present in one copy. When present in three copies, differentially expressed genes are in general less buffered than ubiquitously expressed genes. Furthermore, we show that the 4th chromosome is compensated in response to dose differences. Our results suggest that general mechanisms exist to stimulate and to repress gene expression of aneuploidy regions and on the 4th chromosome this compensation is mediated by POF (Painting of Fourth). Experiment Overall Design: We prepared total RNA from flies heterozygous for three different deletions, Df(2L)J-H, Df (2L)ED4470, Df(2L)ED4651, flies heterozygous for the duplication Dp(2;2)Cam3 and flies with only one chromosome 4, or three copies of the 4th chromosome, as well as from wild type control flies. Three biological replicates of all genotypes were prepared.
Project description:Heterochromatin protein 1 (HP1) proteins are important regulators of heterochromatin mediated gene silencing and chromosome structure and it is well known as the reader of the heterochromatin mark methylation of histone H3 lysine 9 (H3K9me). In Drosophila three different histone lysine methyl transferases (HKMTs) are associated with the methylation of H3K9; Su(var)3-9, Setdb1 and G9a. To gain insights on the dependence of HP1a on the three different HKMTs, the division of labor between these methyl transferases and the dependence of HP1a on H3K9me we have studied HP1a binding in relation to H3K9me in mutants of these HKMTs. We show that Su(var)3-9 is responsible for the HP1a H3K9me-dependent binding in pericentromeric regions while Setdb1 controls the HP1a H3K9me-dependent binding to cytological region 2L:31 and together with POF chromosome 4. HP1a binds to the promoters and within gene bodies of active genes in these three regions. More importantly, HP1a bound at promoters of active genes are independent of H3K9me and POF and is associated to heterochromatin protein 2 (HP2) and open chromatin. Our results supports a model where HP1a nucleates with high affinity independent of H3K9me in promoters of active genes and then spreads via H3K9 methylation and transient looping contacts with those H3K9me target sites.
Project description:Mapping the Drosophila melanogaster centromeric heterochromatin by CGH analysis of embryos lacking specific chromosomes or chromosome arms. Nine chromosome or chromosome arm deletions were tested: embryos lacking the entire second chromosome (2En-), 2L (2L-), 2R (2R-), the entire third chromosome (3En-), 3L (3L-), 3R (3R-), the entire fourth chromosome (4En-), the X chromosome (X-), or both X and Y chromosomes (XY-). Control: Blastoderm stage wild type Oregon R embryos. For each experiment 100-150 embryos of the appropriate genotype were collected. DNA from randomly staged 0-8 hr wild type Oregon R embryos was used as reference for all experiments. Embryos with no X chromosome were obtained by crossing attached-X/Y females (C(1)DX, y f) to X/Y males. Embryos with no X and Y chromosomes were obtained by crossing attached-X/Y females (C(1)RM, y2suwawa) to attached-XY males (YSX YL, In(1)EN, y B). The compound II chromosomes RM(2L); RM(2R)=C(2)v and the compound III chromosomes RM(3L); RM(3R)=C(3)se were used to generate 2L- and 2R-, and 3L- and 3R- embryos, respectively. The compound II C(2)EN and compound III C(3)EN st1 cu1es stocks were used to generate embryos deficient for the entire second and third chromosome, respectively. The compound IV C(4)RM, ci1eyR/0 were used to generate embryos deficient for the fourth chromosome. Embryos deficient for chromosome 4 were identified by their defects in denticle belt patterning during late embryogenesis, whereas embryos deficient for other chromosome/chromosome arm were recognized based on their specific phenotypic defects during early embryonic development. All embryos were collected at room temperature.
Project description:Drosophila Haspin kinase phosphorylates Histone H3 at threonine 3 at centromeric heterochromatin and either lamin- or polycomb-enriched euchromatic regions, being required for nuclear organization of interphase cells and polycomb-dependent gene silencing.