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Late replication of the inactive x chromosome is independent of the compactness of chromosome territory in human pluripotent stem cells

2013-04-01 13:45:39

ActaNaturae; 2013 Apr; 5(2):54-61


Panova AV, Nekrasov ED, Lagarkova MA, Kiselev SL, Bogomazova AN



Introduction



The chromatin structure and architecture of the nucleus are the crucial elements in the regulation of transcription and replication, the key genetic processes that occur in nuclei. Chromosomes occupy certain non-overlapping regions in the interphase nucleus, forming the so-called chromosome territories. The densely packed chromatin mostly localizes in the peripheral and perinucleolar regions of the nucleus. The replication of dispersed euchromatin and densely packed heterochromatin is separated both in space and in time. Dispersed euchromatin replicates during the early S-phase, while the condensed heterochromatin replicates in the late S-phase. T. Ryba et al. have put forward a hypothesis that late replication of densely packed chromatin domains can be attributed to the fact that access for the replication initiation factors to these regions is hindered.



An inactivated X chromosome (Xi), which becomes transcriptionally silent as a result of the dosage compensation, forms the compact structure known as the Barr body on the nuclear periphery and replicates in the late S-phase, is an example of a bulk heterochromatin domain inside the nucleus in female mammalian somatic cells.



The variability of the status of X chromosome inactivation in female human pluripotent stem cells (PSCs) provides an interesting opportunity for studying the relationship between the different epigenetic states of chromatin, the architecture of the chromosome territories in the interphase nucleus, and the regulation of replication.



Up to now, female human embryonic stem cell (hESCs) lines with two active X chromosomes (Xa), one inactivated X chromosome, and hESC lines without any conventional cytological indicators of X inactivation have been described. As for human induced pluripotent stem cells (iPSCs), there is no clear opinion about the possibility of complete reactivation of the X chromosome and the possibility of long-term maintenance of the active status in vitro. However, the X chromosome during reprogramming undoubtedly undergoes a number of significant epigenetic changes associated at least with partial reactivation. Our study was aimed at searching for a relationship between the replication timing of the X chromosomes in human PSCs with different statuses of X chromosome inactivation and the degree of compactness of their chromosome territories.



Our results demonstrate that replication of the inactive X chromosome in the late S-phase of the cell cycle can be unrelated to the compactness of the chromosome area and that the late-replicating and transcriptionally silent Xi can be present in female PSCs in the relaxed state. Nevertheless, the X chromosome territory relaxes as the X chromosome becomes active, which is accompanied by synchronization of the replication of homologous X chromosomes.



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