Monitoring and quantifying replication fork dynamics with high-throughput methods.

Before each cell division, eukaryotic cells must replicate their chromosomes to ensure the accurate transmission of genetic information. Chromosome replication involves more than just DNA duplication; it also includes chromatin assembly, inheritance of epigenetic marks, and faithful resumption of all genomic functions after replication. Recent progress in quantitative technologies has revolutionized our understanding of the complexity and dynamics of DNA replication forks at both molecular and genomic scales.

Prospectively defined patterns of APOBEC3A mutagenesis are prevalent in human cancers.

Mutational signatures defined by single base substitution (SBS) patterns in cancer have elucidated potential mutagenic processes that contribute to malignancy. Two prevalent mutational patterns in human cancers are attributed to the APOBEC3 cytidine deaminase enzymes. Among the seven human APOBEC3 proteins, APOBEC3A is a potent deaminase and proposed driver of cancer mutagenesis.

Genome-wide and sister chromatid-resolved profiling of protein occupancy in replicated chromatin with ChOR-seq and SCAR-seq.

Elucidating the mechanisms underlying chromatin maintenance upon genome replication is critical for the understanding of how gene expression programs and cell identity are preserved across cell divisions. Here, we describe two recently developed techniques, chromatin occupancy after replication (ChOR)-seq and sister chromatids after replication (SCAR)-seq, that profile chromatin occupancy on newly replicated DNA in mammalian cells in 5 d of bench work. Both techniques share a common strategy that includes pulse labeling of newly synthesized DNA and chromatin immunoprecipitation (ChIP), followed by purification and high-throughput sequencing.

DNA molecular combing-based replication fork directionality profiling.

The replication strategy of metazoan genomes is still unclear, mainly because definitive maps of replication origins are missing. High-throughput methods are based on population average and thus may exclusively identify efficient initiation sites, whereas inefficient origins go undetected. Single-molecule analyses of specific loci can detect both common and rare initiation events along the targeted regions.

Staying true to yourself: mechanisms of DNA methylation maintenance in mammals.

DNA methylation is essential to development and cellular physiology in mammals. Faulty DNA methylation is frequently observed in human diseases like cancer and neurological disorders. Molecularly, this epigenetic mark is linked to other chromatin modifications and it regulates key genomic processes, including transcription and splicing.

Chromatin replication and epigenetic cell memory.

Propagation of the chromatin landscape across cell divisions is central to epigenetic cell memory. Mechanistic analysis of the interplay between DNA replication, the cell cycle, and the epigenome has provided insights into replication-coupled chromatin assembly and post-replicative chromatin maintenance. These breakthroughs are critical for defining how proliferation impacts the epigenome during cell identity changes in development and disease.

Accurate Recycling of Parental Histones Reproduces the Histone Modification Landscape during DNA Replication.

Chromatin organization is disrupted genome-wide during DNA replication. On newly synthesized DNA, nucleosomes are assembled from new naive histones and old modified histones. It remains unknown whether the landscape of histone post-translational modifications (PTMs) is faithfully copied during DNA replication or the epigenome is perturbed.

MCM2 promotes symmetric inheritance of modified histones during DNA replication.

During genome replication, parental histones are recycled to newly replicated DNA with their posttranslational modifications (PTMs). Whether sister chromatids inherit modified histones evenly remains unknown. We measured histone PTM partition to sister chromatids in embryonic stem cells.

Replication landscape of the human genome.

Despite intense investigation, human replication origins and termini remain elusive. Existing data have shown strong discrepancies. Here we sequenced highly purified Okazaki fragments from two cell types and, for the first time, quantitated replication fork directionality and delineated initiation and termination zones genome-wide.

Functional study of the Hap4-like genes suggests that the key regulators of carbon metabolism HAP4 and oxidative stress response YAP1 in yeast diverged from a common ancestor.

The transcriptional regulator HAP4, induced by respiratory substrates, is involved in the balance between fermentation and respiration in S. cerevisiae. We identified putative orthologues of the Hap4 protein in all ascomycetes, based only on a conserved sixteen amino acid-long motif.

From simple bacterial and archaeal replicons to replication N/U-domains.

The Replicon Theory proposed 50 years ago has proven to apply for replicons of the three domains of life. Here, we review our knowledge of genome organization into single and multiple replicons in bacteria, archaea and eukarya. Bacterial and archaeal replicator/initiator systems are quite specific and efficient, whereas eukaryotic replicons show degenerate specificity and efficiency, allowing for complex regulation of origin firing time.