Chromosomal replication initiates and terminates at random sequences but at regular intervals in the ribosomal DNA of Xenopus early embryos.

Abstract

We have analysed the replication of the chromosomal ribosomal DNA (rDNA) cluster in Xenopus embryos before the midblastula transition. Two‐dimensional gel analysis showed that replication forks are associated with the nuclear matrix, as in differentiated cells, and gave no evidence for single‐stranded replication intermediates (RIs). Bubbles, simple forks and double Ys were found in each restriction fragment analysed, showing that replication initiates and terminates without detectable sequence specificity. Quantification of the results and mathematical analysis showed that the average rDNA replicon replicates in 7.5 min and is 9‐12 kbp in length. This time is close to the total S phase duration, and this replicon size is close to the maximum length of DNA which can be replicated from a single origin within this short S phase. We therefore infer that (i) most rDNA origins must be synchronously activated soon in S phase and (ii) origins must be evenly spaced, in order that no stretch of chromosomal DNA is left unreplicated at the end of S phase. Since origins are not specific sequences, it is suggested that this spatially and temporally concerted pattern of initiation matches some periodic chromatin folding, which itself need not rely on DNA sequence.

DNA replication timing during development anticipates transcriptional programs and parallels enhancer activation

“…Cold Spring Harbor Laboratory Press on May 11, 2018 -Published by genome.cshlp.org Downloaded from domains are replicated in a reproducible manner but origins and origin clusters fire stochastically (Mills et al 1989;Hyrien and Méchali 1993;Dimitrova and Gilbert 1999;Labit et al 2008). This study established that at low temporal resolution the program appears random; however, at high temporal and spatial resolution allowed by the sequencing-based, genome-wide approach, a fine yet significantly structured program emerges and is visually evident with normalized replication timing profiles.…”

Perturbation of the Activity of Replication Origin by Meiosis-specific Transcription

“…Bars in the bottom graph are standard errors of two experiments. observed at the time of the mid-blastula transition (32,33). The mechanisms behind the origin specification remain poorly understood.…”

Persistence Length of Chromatin Determines Origin Spacing in Xenopus Early-Embryo DNA Replication: Quantitative Comparisons between Theory and Experiment

“…Note again that Equation 1 does not accurately describe this large-l limit, which has been modeled more accurately as a Gaussian chain. 23 If the dynamics are diffusive, i.e., if the reactive groups bind the first time they encounter each other within some small reaction range α (< 1), we can show that the SY approximation continues to hold in the regime where the loop-size is less than a few times the persistence length, and the loop-formation time τ c is given by (2) where C is a dimensionless prefactor that is practically a constant (~10 -1 ) for all l, and D is the diffusion constant. 24 This "first return time" τ c predicted by Equation 2 is very short (10 -3 to 10 -2 seconds for chromatin, comparable to that of linear dsDNA 25 ), implying that loop-formation dynamics are much faster than replication time scales (~20 minutes).…”

“…Because the length of S phase is determined by the replication of the entire genome, even relatively rare long gaps could prolong S phase beyond its observed duration of 10-20 minutes for complete duplication of the whole genome (3 billion bases). 2,3 The problem is all the more acute in that early embryo cells lack an efficient S/M checkpoint, 4 which is used by many eukaryotic cells to delay entry into mitosis in the presence of unreplicated DNA. This problem is formally stated as the "random-completion problem," 5 and, because of the reasons explained above, its solution requires a mechanism that regulates replication other than sequence.…”