The dynamics of replication licensing in live Caenorhabditis elegans embryos

TY - JOUR

T1 - The dynamics of replication licensing in live Caenorhabditis elegans embryos

A1 - Sonneville,Remi

A1 - Querenet,Matthieu

A1 - Craig,Ashley

A1 - Gartner,Anton

A1 - Blow,J. Julian

AU - Sonneville,Remi

AU - Querenet,Matthieu

AU - Craig,Ashley

AU - Gartner,Anton

AU - Blow,J. Julian

PY - 2012/1/23

Y1 - 2012/1/23

N2 - <p>Accurate DNA replication requires proper regulation of replication licensing, which entails loading MCM-2-7 onto replication origins. In this paper, we provide the first comprehensive view of replication licensing in vivo, using video microscopy of Caenorhabditis elegans embryos. As expected, MCM-2-7 loading in late M phase depended on the prereplicative complex (pre-RC) proteins: origin recognition complex (ORC), CDC-6, and CDT-1. However, many features we observed have not been described before: GFP-ORC-1 bound chromatin independently of ORC-2-5, and CDC-6 bound chromatin independently of ORC, whereas CDT-1 and MCM-2-7 DNA binding was interdependent. MCM-3 chromatin loading was irreversible, but CDC-6 and ORC turned over rapidly, consistent with ORC/CDC-6 loading multiple MCM-2-7 complexes. MCM-2-7 chromatin loading further reduced ORC and CDC-6 DNA binding. This dynamic behavior creates a feedback loop allowing ORC/CDC-6 to repeatedly load MCM-2-7 and distribute licensed origins along chromosomal DNA. During S phase, ORC and CDC-6 were excluded from nuclei, and DNA was overreplicated in export-defective cells. Thus, nucleocytoplasmic compartmentalization of licensing factors ensures that DNA replication occurs only once.</p>

AB - <p>Accurate DNA replication requires proper regulation of replication licensing, which entails loading MCM-2-7 onto replication origins. In this paper, we provide the first comprehensive view of replication licensing in vivo, using video microscopy of Caenorhabditis elegans embryos. As expected, MCM-2-7 loading in late M phase depended on the prereplicative complex (pre-RC) proteins: origin recognition complex (ORC), CDC-6, and CDT-1. However, many features we observed have not been described before: GFP-ORC-1 bound chromatin independently of ORC-2-5, and CDC-6 bound chromatin independently of ORC, whereas CDT-1 and MCM-2-7 DNA binding was interdependent. MCM-3 chromatin loading was irreversible, but CDC-6 and ORC turned over rapidly, consistent with ORC/CDC-6 loading multiple MCM-2-7 complexes. MCM-2-7 chromatin loading further reduced ORC and CDC-6 DNA binding. This dynamic behavior creates a feedback loop allowing ORC/CDC-6 to repeatedly load MCM-2-7 and distribute licensed origins along chromosomal DNA. During S phase, ORC and CDC-6 were excluded from nuclei, and DNA was overreplicated in export-defective cells. Thus, nucleocytoplasmic compartmentalization of licensing factors ensures that DNA replication occurs only once.</p>

KW - ORIGIN RECOGNITION COMPLEX

KW - EUKARYOTIC DNA-REPLICATION

KW - C-ELEGANS

KW - CELL-CYCLE

KW - MCM2-7 HELICASE

KW - GEMININ HOMOLOG

KW - ATP-HYDROLYSIS

KW - RE-REPLICATION

KW - S-PHASE

KW - CHROMATIN

U2 - 10.1083/jcb.201110080

DO - 10.1083/jcb.201110080

M1 - Article

JO - Journal of Cell Biology

JF - Journal of Cell Biology

SN - 0021-9525

IS - 2

VL - 196

SP - 233

EP - 246

ER -