The chicken lysozyme gene domain is distinguished by a broad knowledge of how its expression is regulated. Here, we examined the in vivo replication of the lysozyme gene locus using polymerase chain reaction amplification and competitive polymerase chain reaction of size-fractionated, nascent DNA strands. We found that DNA replication initiates at multiple sites within a broad initiation zone spanning at least 20 kilobases, which includes most of the lysozyme gene domain. The 5 border of this zone is probably located downstream of the lysozyme 5 nuclear matrix attachment region. Preferred initiation occurs in a 3-located subzone. The initiation zone at the lysozyme gene locus is also active in nonexpressing liver DU249 cells. Furthermore, examining the timing of DNA replication at the lysozyme gene locus revealed that the gene locus replicates early during S phase in both HD11 and DU249 cells, irrespective of its transcriptional activity.
Summary Chromatin in eukaryotic cells is partitioned into loop‐domains by binding of MARs to the nuclear matrix. It is increasingly accepted that the replication event is attached to the nuclear matrix. In this study, chicken myelomonocytic HD 11 cells were synchronized at the G1/S phase boundary, and after release into the S phase, were immediately pulse‐labelled with 5‐bromodeoxyuridine (BrdU). The chromatin was then fractionated into the nuclear matrix and a solubilized fraction to determine the partitioning of BrdU in newly replicated DNA. We found that in early S phase‐synchronized cells newly replicated DNA was highly enriched in the nuclear matrix. This result indicates that origins of DNA replication in eukaryotic cells are attached at the nuclear matrix. Zusammenfassung DNA‐Replikation ereignet sich an der Kernmatrix Das eukaryotische Chromatin ist durch die Bindung von MARs an die Kernmatrix in schleifenartige Domainen organisiert. Es wird für wahrscheinlich gehalten, daß der Replikationsvorgang an die Kernmatrix gebunden ist. In dieser Untersuchung wurden myelomonozytische HD 11‐Zellen synchronisiert und am Beginn der S‐Phase mit 5‐Bromdesoxyuridin (BrdU) pulsmarkiert. Nach Fraktionierung des Chromatins in die Kernmatrix und eine lösliche Fraktion wurde die Verteilung von BrdU bestimmt. Zu Beginn der S‐Phase replizierte DNA in synchronisierten Zellen war in der Kernmatrix hoch angereichert. Das Ergebnis liefert somit einen weiteren Hinweis für eine Anheftung der Origins der DNA‐Replikation an die Kernmatrix.
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