Simple and Complex Centromeric Satellites in Drosophila Sibling Species
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ABSTRACT:
PROVIDER: PRJNA414639 | ENA |
REPOSITORIES: ENA
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Project description:Centromeres are the chromosomal sites of assembly for kinetochores, the protein complexes that attach to spindle fibers and mediate separation of chromosomes to daughter cells in mitosis and meiosis. In most multicellular organisms, centromeres comprise a single specific family of tandem repeats, often 100-400 bp in length, found on every chromosome, typically in one location within heterochromatin. Drosophila melanogaster is unusual in that the heterochromatin contains many families of mostly short (5-12 bp) tandem repeats, none of which appears to be present at all centromeres, and none of which is found only at centromeres. Although centromere sequences from a minichromosome have been identified and candidate centromere sequences have been proposed, the DNA sequences at native Drosophila centromeres remain unknown. Here we use native chromatin immunoprecipitation to identify the centromeric sequences bound by the foundational kinetochore protein cenH3, known in vertebrates as CENP-A. In D. melanogaster, these sequences include a few families of 5-bp and 10-bp repeats, but in closely related D. simulans, a partially overlapping set of short repeats and more complex repeats comprise the centromeres. The results suggest that a recent expansion of short repeats is replacing more complex centromeric repeats in the melanogaster subgroup of Drosophila.
2018-01-08 | GSE105100 | GEO
Project description:Centromeres are the chromosomal sites of assembly for kinetochores, the protein complexes that attach to spindle fibers and mediate separation of chromosomes to daughter cells in mitosis and meiosis. In most multicellular organisms, centromeres comprise a single specific family of tandem repeats-often 100-400 bp in length-found on every chromosome, typically in one location within heterochromatin. Drosophila melanogaster is unusual in that the heterochromatin contains many families of mostly short (5-12 bp) tandem repeats, none of which appear to be present at all centromeres, and none of which are found only at centromeres. Although centromere sequences from a minichromosome have been identified and candidate centromere sequences have been proposed, the DNA sequences at native Drosophila centromeres remain unknown. Here we use native chromatin immunoprecipitation to identify the centromeric sequences bound by the foundational kinetochore protein cenH3, known in vertebrates as CENP-A. In D. melanogaster, these sequences include a few families of 5- and 10-bp repeats; but in closely related D. simulans, the centromeres comprise more complex repeats. The results suggest that a recent expansion of short repeats has replaced more complex centromeric repeats in D. melanogaster.
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Project description:Centromeres of most eukaryotes are multi-megabase arrays of satellite DNA that assemble proteinaceous kinetochores to facilitate faithful chromosome segregation. However, the nature of the chromatin landscape at centromeres remains unclear, in part due to the difficulty of isolating intact centromeric chromatin rendered insoluble by megadalton-size kinetochore protein complexes. To address this challenge we combined classical salt fractionation with chromatin immunoprecipitation to recover human centromeric chromatin under native conditions. We found that >85% of the total centromeric chromatin is insoluble under conditions typically used for native chromatin extraction. To map both soluble and insoluble chromatin in situ, we combined CUT&RUN, a targeted nuclease method, with salt fractionation. Using this approach, we found that the strength of Centromere Protein B (CENP-B) binding and the density of CENP-B motifs corresponds to the occupancy of Constitutive Centromere-Assocated Network (CCAN) complexes bound to α-satellite arrays. We also observed unexpected structural variations of CENP-A-containing complexes on different α-satellite dimeric units within highly homogenous arrays. Our results suggest that slight α-satellite sequence differences controls the structure and occupancy of the associated centromeric chromatin complex.
2018-01-31 | GSE104805 | GEO
Project description:Unexpected conformational variations of the human centromeric chromatin complex
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Project description:Speciation involves the reproductive isolation of natural populations due to the sterility or lethality of their hybrids. However, the molecular basis of hybrid lethality and the evolutionary driving forces that provoke it, remain largely elusive. The hybrid male rescue (Hmr) and the lethal hybrid rescue (Lhr) genes serve as a model to study speciation in Drosophilids as their interaction causes lethality in male hybrid offspring. Here we show that HMR and LHR form a centromeric complex necessary for proper chromosome segregation. We find that the Hmr expression level is substantially higher in D. melanogaster whereas Lhr expression levels are increased in D. simulans. The resulting elevated amount of HMR/LHR complex in hybrids results in an extensive mislocalisation of the complex, an interference with the regulation of transposable elements and an impairment of cell proliferation. Our findings provide evidence for a major role of centromere divergence in the generation of biodiversity. Raw data were analysed using the MaxQuant 1.2.2.5 software package. Identified proteins were considered as interation partners if their MaxQuant iBAQ values displayed a greater than 16fold enrichment compared to control anti-FLAG purifications from Schneider cell nuclear extracts not expressing any FLAG-tagged protein.
2013-11-20 | PXD000489 | Pride
Project description:To investigate the exact locations of CCTT-chromatin interaction on a genome-wide scale, we modified previously reported ChIRP-seq (chromatin isolation by RNA purification followed by deep-sequencing) with a crosslinker 4’- aminomethyltrioxalen (AMT, a psoralen derivative), which allows fixation of nucleic acid interaction by ultraviolet light without crosslinking proteins. We designed 8 complementary DNA oligonucleotides that tiled the Alu-depleted part of CCTT. Affinity-purified CCTT-binding DNAs were sequenced and mapped to the HuRef genome hg38, which contains human α-satellite sequence models in each centromeric region. In contrast with 5.38% of input reads mapping to α-satellite sequence, 16.89% of CCTT-binding reads were enriched at α-satellites. Peaks bound by lnc-CCTT were identified by MACS2 algorithm, which compared the reads in CCTT-captured samples with that in input ones. CCTT-binding peaks mapped across the centromeric regions of all 23 reference centromeres. Next, we validated the enrichment of centromeric peaks located at all chromosomes via ChIRP-qPCR. To validate our ChIRP-seq results, we selected representatives of three major subpopulations: multimapping peaks in one specific chromosome and several chromosomes, as well as single-copy peaks. We performed ChIRP-qPCR and found abundant CCTT-binding centromeric peaks throughout all 23 chromosomes. Importantly, CCTT ChIRP-seq profile was highly correlated with the previously reported ChIP-seq profiles of CENP-C (Pearson correlation R = 0.82), both of which showed very strong, extensive signals across entire centromeric regions in HeLa cells.
2022-04-20 | GSE149532 | GEO
Project description:In human cells chromosome specific aneuploidy was measured. Then, using NGS methods, centromeric features were measured for each chromosome (including centromere length and amount of centromeric proteins). The study aims at testing an association between centromeric features and missegregation rate.
2019-11-26 | GSE132193 | GEO
Project description:Drosophila simulans relies exclusively on the fruits of Morinda citrifolia, which are toxic to most insects, including its sibling species D. melanogaster and D. simulans. Although several odorant binding protein (Obp) genes and olfactory receptor (Or) genes were suggested to be associated with the D. simulans host shift, a broad view of how chemosensory genes have contributed to this shift is still lacking. We therefore studied the antennal transcriptomes, the main organ responsible for detecting food resource and oviposition, of D. simulans and its two sibling species. We wanted to know whether gene expression, particularly chemosensory genes, has diverged between D. simulans and its two sibling species. Using a very stringent definition of differential gene expression, we found 147 genes (including 11 chemosensory genes) were up-regulated while only 81 genes (including 5 chemosensory genes) were down-regulated in D. simulans. Interestingly, Obp50a exhibited the highest up-regulation, a ~100 fold increase, and Or85c – previously reported to be a larva-specific gene– showed ~20 fold up-regulation in D. simulans. Furthermore, Ir84a, proposed to be associated with male courtship behavior, is significantly up-regulated in D. simulans. We also found expression divergence in most of the receptor gene families between D. simulans and the two sibling species. Our observations suggest that the host shift of D. simulans is associated with expression profile divergence in all chemosensory gene families and is achieved mostly by up-regulation of chemosensory genes.
2015-10-01 | GSE67862 | GEO
Project description:Drosophila sechellia relies exclusively on the fruits of Morinda citrifolia, which are toxic to most insects, including its sibling species D. melanogaster and D. simulans. Although several odorant binding protein (Obp) genes and olfactory receptor (Or) genes were suggested to be associated with the D. sechellia host shift, a broad view of how chemosensory genes have contributed to this shift is still lacking. We therefore studied the antennal transcriptomes, the main organ responsible for detecting food resource and oviposition, of D. sechellia and its two sibling species. We wanted to know whether gene expression, particularly chemosensory genes, has diverged between D. sechellia and its two sibling species. Using a very stringent definition of differential gene expression, we found 147 genes (including 11 chemosensory genes) were up-regulated while only 81 genes (including 5 chemosensory genes) were down-regulated in D. sechellia. Interestingly, Obp50a exhibited the highest up-regulation, a ~100 fold increase, and Or85c – previously reported to be a larva-specific gene– showed ~20 fold up-regulation in D. sechellia. Furthermore, Ir84a, proposed to be associated with male courtship behavior, is significantly up-regulated in D. sechellia. We also found expression divergence in most of the receptor gene families between D. sechellia and the two sibling species. Our observations suggest that the host shift of D. sechellia is associated with expression profile divergence in all chemosensory gene families and is achieved mostly by up-regulation of chemosensory genes.
2015-10-01 | GSE67587 | GEO