Chromosome Engineering Allows the Efficient Isolation of Vertebrate Neocentromeres

Autor: Shang, W. H., Hori, T., Martins, N. M., Toyoda, A., Misu, S., Monma, N., Hiratani, I., Maeshima, K., Ikeo, K., Fujiyama, A., Kimura, Hiroshi, Earnshaw, W. C., Fukagawa, T.
Jazyk: angličtina
Rok vydání: 2013
Předmět:
DNA Replication
Chromosome engineering
Chromatin Immunoprecipitation
Neocentromere
Transcription
Genetic
Chromosomal Proteins
Non-Histone
Centromere
Autoantigens/genetics
Autoantigens
General Biochemistry
Genetics and Molecular Biology
Article
Chromosomes
Cell Line
Epigenesis
Genetic
Chromosomal Proteins
Non-Histone/genetics
Chromosomes/*genetics
03 medical and health sciences
Histone H3
0302 clinical medicine
Animals
Molecular Biology
030304 developmental biology
Genetics
0303 health sciences
Replication timing
biology
Base Sequence
DNA replication
Cell Biology
Sequence Analysis
DNA
DNA Methylation
Cell biology
Chickens/*genetics
Histone
Centromere/*genetics/metabolism
biology.protein
Genetic Engineering
Chromatin immunoprecipitation
Chickens
030217 neurology & neurosurgery
Centromere Protein A
Developmental Biology
Zdroj: Developmental Cell
ISSN: 1878-1551
1534-5807
Popis: Summary Centromeres are specified by sequence-independent epigenetic mechanisms in most organisms. Rarely, centromere repositioning results in neocentromere formation at ectopic sites. However, the mechanisms governing how and where neocentromeres form are unknown. Here, we established a chromosome-engineering system in chicken DT40 cells that allowed us to efficiently isolate neocentromere-containing chromosomes. Neocentromeres appear to be structurally and functionally equivalent to native centromeres. Chromatin immunoprecipitation sequencing (ChIP-seq) analysis with 18 neocentromeres revealed that the centromere-specific histone H3 variant CENP-A occupies an ∼40 kb region at each neocentromere, which has no preference for specific DNA sequence motifs. Furthermore, we found that neocentromeres were not associated with histone modifications H3K9me3, H3K4me2, and H3K36me3 or with early replication timing. Importantly, low but significant levels of CENP-A are detected around endogenous centromeres, which are capable of seeding neocentromere assembly if the centromere core is removed. In summary, our experimental system provides valuable insights for understanding how neocentromeres form.
Graphical Abstract Highlights ► Chromosome engineering efficiently generates neocentromeres in chicken DT40 cells ► CENP-A reproducibly occupies an ∼40 kb genomic region at each neocentromere ► Nonkinetochore CENP-A appears to function as a seed for neocentromere assembly
Centromeres are specified by sequence-independent epigenetic mechanisms. Shang et al. generated a collection of chicken neocentromeres in DT40 cells. Their analysis indicates that neocentromere formation does not correlate with the expected histone modifications or with replication timing, but rather depends on the histone H3 variant CENP-A to seed assembly.
Databáze: OpenAIRE
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