Maturation of newly replicated chromatin

We undertook a study to obtain a clearer understanding of the maturation process in rat hepatoma tissue culture (HTC) cells. HTC cells were labeled with short pulses of [3H] thymidine and chased with cold thymidine for varying lengths of time in order to identify newly synthesized DNA at different stages of the maturation process. Nuclei were isolated directly or after an extensive fixation with formaldehyde to prevent the dissociation of histones from the DNA. An analysis of the repeat lengths obtained from nascent and bulk chromatin revealed that there are two stages in the maturation of newly replicated chromatin in HTC cells. The first stage is relatively brief (< 5 min), involves the chromatin which is immediately adjacent to the replication fork, and is characterized by a diffuse organization of nucleosomes on nascent DNA. The second stage, which lasts from 5-30 minutes post-replication, is characterized by a normal repeat length for the nascent nucleosomes, which are more easily able to slide along the DNA than bulk nucleosomes, resulting in an apparently shorter nucleosomal repeat length