Studies Of DNA Composition And Structure Of The Centromere And Chromosomes Spatial Distribution In Cotton

As an essential functional element in eukaryotic chromosomes,centromere is involved in kinetochore formation,chromosomes separation,sister chromatid bonding,homologous chromosome pairing and other important biological processes.Centromeric sequences are mainly composed of retrotransposons and satellites,but are highly variable between different species and even between close relatives.Therefore,it is difficult to be cloned and further analyzed.Here,we conducted an isolation and analysis of centromere DNA by chromatin immunoprecipitation and high-throughput sequencing(ChIP-seq)against centromere-specific histone CENH3(centromeric histone H3;also known as CENP-A)in Gossypium raimondii(D-genome).Gossypium raimondii is a putative diploid progenitor of the allopolyploid(AD-genome)cottons.The results showed that:(1)Centromeric repeat of the cotton were LTR-like retrotransposons,belonging to plant CRM retrotransposons family.(2)By comparative analyses,we found that these sequences did not exist or only had a very low copy number in Gossypium arboreum(AA),which is another putative progenitor of the allopolyploid(AD-genome)cottons.However,there was abundant amplification in the tetraploid upland cotton(AADD),suggesting that centromeric repeat of D-genome cotton had a burst growth after polyploidization in cotton.(3)Meanwhile,a joint analysis of the transcriptomic data were conducted.We found that there is still transcriptionally active genes in centromere regions.However,the gene density and expression level were significantly lower than other regions of chromosome.(4)In addition,identification of centromere regions in the existing genome revealed that some chromosomes might assembly errors in centromere regions.The development of this work provides important clues for resolving centromere structure,function and its evolutionary mechanisms.It also provides basic knowledge for the future development of artificial chromosomes in cotton.Chromosomes are not randomly distributed within the nucleus but occupy certain nucleus space and form chromosome territory(CT).Studies have shown that this ordered organization of genome is critical for the gene expression regulation and other genomic functions.Therefore,the analysis of genomic three-dimensional structure has become an important component of functional genomics research in recent years.We conducted a study on the distribution of metaphase chromosomes in Gossypium hirsutum(AADD).We found that:(1)The two subgenome,A and D,were spatially separated and showed a typical radial arrangement pattern,in which the small D subgenome chromosomes tended to concentrate at the center and the large A subgenome chromosomes were scattered periphery.(2)The observation of chromosomes distribution in an artificial hexaploid(AADDGG)revealed that ordered chromosomes arrangement of A subgenome chromosomes and D subgenome chromosomes were disturbed due to the entry of G subgenome chromosomes.However,the radial separation pattern of A subgenome chromosomes and D subgenome chromosomes could be recovered with the continuous loss of G subgenome chromosomes after backcrossing the artificial hexaploid with the Gossypium hirsutum.(3)Additionally,a similar genome separation phenomenon was also found in synthesized tetraploid cotton(AAGG).These results indicate that the genome separation pattern could be established immediately after tetraploid cotton formation and could be stably inherited in tetraploid cotton.Given the research in other plants and animals,we speculated that genome separation might be a normal phenomenon between different genomes.This finding provides evidence and reference for elucidating the genomic conformation regular and function in plant cells.