title: Dynamics of plant DNA replication based on PCNA visualization.
abstract: DNA replication is an essential process for the copying of genomic information in living organisms. Imaging of DNA replication in tissues and organs is mainly performed using fixed cells after incorporation of thymidine analogs. To establish a useful marker line to measure the duration of DNA replication and analyze the dynamics of DNA replication, we focused on the proliferating cell nuclear antigen (PCNA), which functions as a DNA sliding clamp for replicative DNA polymerases and is an essential component of replisomes. In this study we produced an Arabidopsis thaliana line expressing PCNA1 fused with the green fluorescent protein under the control of its own promoter (pAtPCNA1::AtPCNA1-sGFP). The duration of the S phase measured using the expression line was consistent with that measured after incorporation of a thymidine analog. Live cell imaging revealed that three distinct nuclear localization patterns (whole, dotted, and speckled) were sequentially observable. These whole, dotted, and speckled patterns of subnuclear AtPCNA1 signals were indicative of the G1 or G2 phase, early S phase and late S phase, respectively. The results indicate that the pAtPCNA1::AtPCNA1-sGFP line is a useful marker line for visualization of S-phase progression in live plant organs.
authors.affiliation: Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan.
authors.forename: Ryohei
authors.initials: R
authors.lastname: Yokoyama
authors.affiliation: Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Hirakawa
authors.affiliation: Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan.
authors.forename: Seri
authors.initials: S
authors.lastname: Hayashi
authors.affiliation: Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Sakamoto
authors.affiliation: Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan.
authors.forename: Sachihiro
authors.initials: S
authors.lastname: Matsunaga
mesh_terms: D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D004261:DNA Replication; D018744:DNA, Plant / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000378:metabolism; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016196:S Phase
chemical_list: D029681:Arabidopsis Proteins; D018744:DNA, Plant; C000604257:PCNA1 antigen, Arabidopsis; D018809:Proliferating Cell Nuclear Antigen; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins