Abstract
BackgroundChromocenters are defined as a punctate condensed blocks of chromatin in the interphase cell nuclei of certain cell types with unknown biological significance. In recent years a progress in revealing of chromocenters protein content has been made although the details of DNA content within constitutive heterochromatin still remain unclear. It is known that these regions are enriched in tandem repeats (TR) and transposable elements. Quick improvement of genome sequencing does not help to assemble the heterochromatic regions due to lack of appropriate bioinformatics techniques.ResultsChromocenters DNA have been isolated by a biochemical approach from mouse liver cells nuclei and sequenced on the Illumina MiSeq resulting in ChrmC dataset. Analysis of ChrmC dataset by the bioinformatics tools available revealed that the major component of chromocenter DNA are TRs: ~ 66% MaSat and ~ 4% MiSat. Other previously classified TR families constitute ~ 1% of ChrmC dataset. About 6% of chromocenters DNA are mostly unannotated sequences. In the contigs assembled with IDBA_UD there are many fragments of heterochromatic Y-chromosome, rDNA and other pseudo-genes and non-coding DNA. A protein coding sfi1 homolog gene fragment was also found in contigs. The Sfi1 homolog gene is located on the chromosome 11 in the reference genome very close to the Golden Pass Gap (a ~ 3 Mb empty region reserved to the pericentromeric region) and proves the purity of chromocenters isolation. The second major fraction are non-LTR retroposons (SINE and LINE) with overwhelming majority of LINE - ~ 11% of ChrmC. Most of the LINE fragments are from the ~ 2 kb region at the end of the 2nd ORF and its’ flanking region. The precise LINEs’ segment of ~ 2 kb is the necessary mouse constitutive heterohromatin component together with TR. The third most abundant fraction are ERVs. The ERV distribution in chromocenters differs from the whole genome: IAP (ERV2 class) is the most numerous in ChrmC while MaLR (ERV3 class) prevails in the reference genome. IAP and its LTR also prevail in TR containing contigs extracted from the WGS dataset. In silico prediction of IAP and LINE fragments in chromocenters was confirmed by direct fluorescent in situ hybridization (FISH).ConclusionOur data of chromocenters’ DNA (ChrmC) sequencing demonstrate that IAP with LTR and a precise ~ 2 kb fragment of LINE represent a substantial fraction of mouse chromocenters (constitutive heteroсhromatin) along with TRs.
Highlights
Chromocenters are defined as a punctate condensed blocks of chromatin in the interphase cell nuclei of certain cell types with unknown biological significance
Investigation of the transcription burst may prove to be vital in resolving at least some of the mysteries surrounding the role of constitutive heterochromatin in development, cell differentiation and responses to stress, RNA only could be recognized if corresponding DNA sequences are known
The underrepresentation of Short interspersed nuclear element (SINE) in chromocenters’ DNA (ChrmC) is visible in comparison with whole genome High-throughput sequencing (wgHTS) and especially with the reference genome
Summary
Chromocenters are defined as a punctate condensed blocks of chromatin in the interphase cell nuclei of certain cell types with unknown biological significance. The nuclear morphology of the neuron changes from a small, heterochromatic nucleus with many randomly-located chromocenters and nucleoli, to a large, mostly euchromatic nucleus with fewer, larger chromocenters associated with a large, centrally located nucleolus [8,9,10,11,12] This non-random reorganization suggests that these changes occur via clustering and relocation of these structures during terminal differentiation and these global chromatin changes have been observed in terminally differentiating neurons in a variety of species, strongly indicating functional significance [9, 13]. Murine periCEN major satellite (MaSat) is highly transcribed during embryogenesis, and transcripts are responsible for reorganization of periCEN DNA into chromocenters Destruction of these transcripts led to developmental arrest indicating their role in de novo heterochromatin formation and proper developmental progression [16]. Investigation of the transcription burst may prove to be vital in resolving at least some of the mysteries surrounding the role of constitutive heterochromatin in development, cell differentiation and responses to stress, RNA only could be recognized if corresponding DNA sequences are known
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