title: Tracking replication enzymology in vivo by genome-wide mapping of ribonucleotide incorporation.
abstract: Ribonucleotides are frequently incorporated into DNA during replication in eukaryotes. Here we map genome-wide distribution of these ribonucleotides as markers of replication enzymology in budding yeast, using a new 5' DNA end-mapping method, hydrolytic end sequencing (HydEn-seq). HydEn-seq of DNA from ribonucleotide excision repair-deficient strains reveals replicase- and strand-specific patterns of ribonucleotides in the nuclear genome. These patterns support the roles of DNA polymerases α and δ in lagging-strand replication and of DNA polymerase ɛ in leading-strand replication. They identify replication origins, termination zones and variations in ribonucleotide incorporation frequency across the genome that exceed three orders of magnitude. HydEn-seq also reveals strand-specific 5' DNA ends at mitochondrial replication origins, thus suggesting unidirectional replication of a circular genome. Given the conservation of enzymes that incorporate and process ribonucleotides in DNA, HydEn-seq can be used to track replication enzymology in other organisms.
authors.affiliation: Genome Integrity &Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institute of Health (NIH), Research Triangle Park, North Carolina, USA.
authors.forename: Anders R
authors.initials: AR
authors.lastname: Clausen
authors.affiliation: Genome Integrity &Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institute of Health (NIH), Research Triangle Park, North Carolina, USA.
authors.forename: Scott A
authors.identifier: 0000000324008325
authors.initials: SA
authors.lastname: Lujan
authors.affiliation: Integrative Bioinformatics, National Institute of Environmental Health Sciences, NIH, Research Triangle Park, North Carolina, USA.
authors.forename: Adam B
authors.initials: AB
authors.lastname: Burkholder
authors.affiliation: Genome Integrity &Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institute of Health (NIH), Research Triangle Park, North Carolina, USA.
authors.forename: Clinton D
authors.initials: CD
authors.lastname: Orebaugh
authors.affiliation: Genome Integrity &Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institute of Health (NIH), Research Triangle Park, North Carolina, USA.
authors.forename: Jessica S
authors.initials: JS
authors.lastname: Williams
authors.affiliation: Department of Genetics, High Throughput Sequencing Facility, University of North Carolina, Chapel Hill, North Carolina, USA.
authors.forename: Maryam F
authors.initials: MF
authors.lastname: Clausen
authors.affiliation: Department of Genetics, High Throughput Sequencing Facility, University of North Carolina, Chapel Hill, North Carolina, USA.
authors.forename: Ewa P
authors.initials: EP
authors.lastname: Malc
authors.affiliation: Department of Genetics, High Throughput Sequencing Facility, University of North Carolina, Chapel Hill, North Carolina, USA.
authors.forename: Piotr A
authors.initials: PA
authors.lastname: Mieczkowski
authors.affiliation: Integrative Bioinformatics, National Institute of Environmental Health Sciences, NIH, Research Triangle Park, North Carolina, USA.
authors.forename: David C
authors.initials: DC
authors.lastname: Fargo
authors.affiliation: Center for Genomics and Systems Biology, Department of Biology, New York University, New York, New York, USA.
authors.forename: Duncan J
authors.initials: DJ
authors.lastname: Smith
authors.affiliation: Genome Integrity &Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institute of Health (NIH), Research Triangle Park, North Carolina, USA.
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Kunkel
mesh_terms: D002874:Chromosome Mapping; D004261:DNA Replication / Q000502:physiology*; D004272:DNA, Mitochondrial / Q000737:chemistry; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism / Q000502:physiology; D005656:Fungal Proteins / Q000378:metabolism / Q000502:physiology; D059014:High-Throughput Nucleotide Sequencing; D018741:Replication Origin; D012265:Ribonucleotides / Q000737:chemistry / Q000502:physiology*; D012441:Saccharomyces cerevisiae / Q000235:genetics; D017421:Sequence Analysis
chemical_list: D004272:DNA, Mitochondrial; D005656:Fungal Proteins; D012265:Ribonucleotides; D004259:DNA-Directed DNA Polymerase