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Dissection of cell cycle-dependent dynamics of Dnmt1 by FRAP and diffusion-coupled modeling

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Dissection of cell cycle-dependent dynamics of Dnmt1 by FRAP and diffusion-coupled modeling. / Schneider, Katrin; Fuchs, Christiane; Dobay, Akos; Rottach, Andrea; Qin, Weihua; Wolf, Patricia; Álvarez-Castro, José M; Nalaskowski, Marcus M; Kremmer, Elisabeth; Schmid, Volker; Leonhardt, Heinrich; Schermelleh, Lothar.

In: NUCLEIC ACIDS RES, Vol. 41, No. 9, 01.05.2013, p. 4860-76.

Research output: SCORING: Contribution to journal › SCORING: Journal article › Research › peer-review

Harvard

Schneider, K, Fuchs, C, Dobay, A, Rottach, A, Qin, W, Wolf, P, Álvarez-Castro, JM, Nalaskowski, MM, Kremmer, E, Schmid, V, Leonhardt, H & Schermelleh, L 2013, 'Dissection of cell cycle-dependent dynamics of Dnmt1 by FRAP and diffusion-coupled modeling', NUCLEIC ACIDS RES, vol. 41, no. 9, pp. 4860-76. https://doi.org/10.1093/nar/gkt191

APA

Schneider, K., Fuchs, C., Dobay, A., Rottach, A., Qin, W., Wolf, P., Álvarez-Castro, J. M., Nalaskowski, M. M., Kremmer, E., Schmid, V., Leonhardt, H., & Schermelleh, L. (2013). Dissection of cell cycle-dependent dynamics of Dnmt1 by FRAP and diffusion-coupled modeling. NUCLEIC ACIDS RES, 41(9), 4860-76. https://doi.org/10.1093/nar/gkt191

Vancouver

Schneider K, Fuchs C, Dobay A, Rottach A, Qin W, Wolf P et al. Dissection of cell cycle-dependent dynamics of Dnmt1 by FRAP and diffusion-coupled modeling. NUCLEIC ACIDS RES. 2013 May 1;41(9):4860-76. https://doi.org/10.1093/nar/gkt191

Bibtex

@article{8573cdb228ba4712a25128448743e10e,
title = "Dissection of cell cycle-dependent dynamics of Dnmt1 by FRAP and diffusion-coupled modeling",
abstract = "DNA methyltransferase 1 (Dnmt1) reestablishes methylation of hemimethylated CpG sites generated during DNA replication in mammalian cells. Two subdomains, the proliferating cell nuclear antigen (PCNA)-binding domain (PBD) and the targeting sequence (TS) domain, target Dnmt1 to the replication sites in S phase. We aimed to dissect the details of the cell cycle-dependent coordinated activity of both domains. To that end, we combined super-resolution 3D-structured illumination microscopy and fluorescence recovery after photobleaching (FRAP) experiments of GFP-Dnmt1 wild type and mutant constructs in somatic mouse cells. To interpret the differences in FRAP kinetics, we refined existing data analysis and modeling approaches to (i) account for the heterogeneous and variable distribution of Dnmt1-binding sites in different cell cycle stages; (ii) allow diffusion-coupled dynamics; (iii) accommodate multiple binding classes. We find that transient PBD-dependent interaction directly at replication sites is the predominant specific interaction in early S phase (residence time Tres ≤ 10 s). In late S phase, this binding class is taken over by a substantially stronger (Tres ∼22 s) TS domain-dependent interaction at PCNA-enriched replication sites and at nearby pericentromeric heterochromatin subregions. We propose a two-loading-platform-model of additional PCNA-independent loading at postreplicative, heterochromatic Dnmt1 target sites to ensure faithful maintenance of densely methylated genomic regions.",
keywords = "Animals, Cell Cycle, Cell Line, Cell Nucleus, DNA (Cytosine-5-)-Methyltransferase, Diffusion, Fluorescence Recovery After Photobleaching, Heterochromatin, Kinetics, Mice, Models, Biological, Protein Structure, Tertiary, S Phase",
author = "Katrin Schneider and Christiane Fuchs and Akos Dobay and Andrea Rottach and Weihua Qin and Patricia Wolf and {\'A}lvarez-Castro, {Jos{\'e} M} and Nalaskowski, {Marcus M} and Elisabeth Kremmer and Volker Schmid and Heinrich Leonhardt and Lothar Schermelleh",
year = "2013",
month = may,
day = "1",
doi = "10.1093/nar/gkt191",
language = "English",
volume = "41",
pages = "4860--76",
journal = "NUCLEIC ACIDS RES",
issn = "0305-1048",
publisher = "Oxford University Press",
number = "9",

}

RIS

TY - JOUR

T1 - Dissection of cell cycle-dependent dynamics of Dnmt1 by FRAP and diffusion-coupled modeling

AU - Schneider, Katrin

AU - Fuchs, Christiane

AU - Dobay, Akos

AU - Rottach, Andrea

AU - Qin, Weihua

AU - Wolf, Patricia

AU - Álvarez-Castro, José M

AU - Nalaskowski, Marcus M

AU - Kremmer, Elisabeth

AU - Schmid, Volker

AU - Leonhardt, Heinrich

AU - Schermelleh, Lothar

PY - 2013/5/1

Y1 - 2013/5/1

N2 - DNA methyltransferase 1 (Dnmt1) reestablishes methylation of hemimethylated CpG sites generated during DNA replication in mammalian cells. Two subdomains, the proliferating cell nuclear antigen (PCNA)-binding domain (PBD) and the targeting sequence (TS) domain, target Dnmt1 to the replication sites in S phase. We aimed to dissect the details of the cell cycle-dependent coordinated activity of both domains. To that end, we combined super-resolution 3D-structured illumination microscopy and fluorescence recovery after photobleaching (FRAP) experiments of GFP-Dnmt1 wild type and mutant constructs in somatic mouse cells. To interpret the differences in FRAP kinetics, we refined existing data analysis and modeling approaches to (i) account for the heterogeneous and variable distribution of Dnmt1-binding sites in different cell cycle stages; (ii) allow diffusion-coupled dynamics; (iii) accommodate multiple binding classes. We find that transient PBD-dependent interaction directly at replication sites is the predominant specific interaction in early S phase (residence time Tres ≤ 10 s). In late S phase, this binding class is taken over by a substantially stronger (Tres ∼22 s) TS domain-dependent interaction at PCNA-enriched replication sites and at nearby pericentromeric heterochromatin subregions. We propose a two-loading-platform-model of additional PCNA-independent loading at postreplicative, heterochromatic Dnmt1 target sites to ensure faithful maintenance of densely methylated genomic regions.

AB - DNA methyltransferase 1 (Dnmt1) reestablishes methylation of hemimethylated CpG sites generated during DNA replication in mammalian cells. Two subdomains, the proliferating cell nuclear antigen (PCNA)-binding domain (PBD) and the targeting sequence (TS) domain, target Dnmt1 to the replication sites in S phase. We aimed to dissect the details of the cell cycle-dependent coordinated activity of both domains. To that end, we combined super-resolution 3D-structured illumination microscopy and fluorescence recovery after photobleaching (FRAP) experiments of GFP-Dnmt1 wild type and mutant constructs in somatic mouse cells. To interpret the differences in FRAP kinetics, we refined existing data analysis and modeling approaches to (i) account for the heterogeneous and variable distribution of Dnmt1-binding sites in different cell cycle stages; (ii) allow diffusion-coupled dynamics; (iii) accommodate multiple binding classes. We find that transient PBD-dependent interaction directly at replication sites is the predominant specific interaction in early S phase (residence time Tres ≤ 10 s). In late S phase, this binding class is taken over by a substantially stronger (Tres ∼22 s) TS domain-dependent interaction at PCNA-enriched replication sites and at nearby pericentromeric heterochromatin subregions. We propose a two-loading-platform-model of additional PCNA-independent loading at postreplicative, heterochromatic Dnmt1 target sites to ensure faithful maintenance of densely methylated genomic regions.

KW - Animals

KW - Cell Cycle

KW - Cell Line

KW - Cell Nucleus

KW - DNA (Cytosine-5-)-Methyltransferase

KW - Diffusion

KW - Fluorescence Recovery After Photobleaching

KW - Heterochromatin

KW - Kinetics

KW - Mice

KW - Models, Biological

KW - Protein Structure, Tertiary

KW - S Phase

U2 - 10.1093/nar/gkt191

DO - 10.1093/nar/gkt191

M3 - SCORING: Journal article

C2 - 23535145

VL - 41

SP - 4860

EP - 4876

JO - NUCLEIC ACIDS RES

JF - NUCLEIC ACIDS RES

SN - 0305-1048

IS - 9

ER -