Concerted Loading of Mcm2–7 Double Hexamers around DNA during DNA Replication Origin Licensing

Remus, Dirk; Beuron, Fabienne; Tolun, Gökhan; Griffith, Jack D.; Morris, Edward P.; Diffley, John F.X.

doi:10.1016/j.cell.2009.10.015

Cited by 608 publications

(899 citation statements)

References 74 publications

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“…The cooperative loading of 2 MCM2-7 complexes results after ATP-dependent dissociation of ORC, Cdc6 and Cdt1 3 in the stable association of MCM2-7 double hexamers. 4 In S phase, the MCM complex provides the helicase activity required for DNA strand separation during DNA replication.…”

Section: Introductionmentioning

confidence: 99%

Spatio-temporal regulation of the human licensing factor Cdc6 in replication and mitosis

et al. 2015

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To maintain genome stability, the thousands of replication origins of mammalian genomes must only initiate replication once per cell cycle. This is achieved by a strict temporal separation of ongoing replication in S phase, and the formation of pre-replicative complexes in the preceding G1 phase, which "licenses" each origin competent for replication. The contribution of the loading factor Cdc6 to the timing of the licensing process remained however elusive due to seemingly contradictory findings concerning stabilization, degradation and nuclear export of Cdc6. Using fluorescently tagged Cdc6 (Cdc6-YFP) expressed in living cycling cells, we demonstrate here that Cdc6-YFP is stable and chromatin-associated during mitosis and G1 phase. It undergoes rapid proteasomal degradation during S phase initiation followed by active export to the cytosol during S and G2 phases. Biochemical fractionation abolishes this nuclear exclusion, causing aberrant chromatin association of Cdc6-YFP and, likely, endogenous Cdc6, too. In addition, we demonstrate association of Cdc6 with centrosomes in late G2 and during mitosis. These results show that multiple Cdc6-regulatory mechanisms coexist but are tightly controlled in a cell cycle-specific manner.

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Section: Introductionmentioning

confidence: 99%

Spatio-temporal regulation of the human licensing factor Cdc6 in replication and mitosis

et al. 2015

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“…A six subunit origin recognition complex (ORC) binds to a limited number of ACSs throughout the cell cycle (Bell and Stillman 1992;Diffley and Cocker 1992). Then in G1, double hexamers of the minichromosome maintenance (MCM) complex are subsequently recruited to origins to form the pre-replicative complex (pre-RC) (Remus et al 2009). Once the pre-RC is formed, origins are licensed to fire.…”

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confidence: 99%

Nucleosome occupancy as a novel chromatin parameter for replication origin functions

et al. 2016

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Eukaryotic DNA replication initiates from multiple discrete sites in the genome, termed origins of replication (origins). Prior to S phase, multiple origins are poised to initiate replication by recruitment of the pre-replicative complex (pre-RC). For proper replication to occur, origin activation must be tightly regulated. At the population level, each origin has a distinct firing time and frequency of activation within S phase. Many studies have shown that chromatin can strongly influence initiation of DNA replication. However, the chromatin parameters that affect properties of origins have not been thoroughly established. We found that nucleosome occupancy in G1 varies greatly around origins across the S. cerevisiae genome, and nucleosome occupancy around origins significantly correlates with the activation time and efficiency of origins, as well as pre-RC formation. We further demonstrate that nucleosome occupancy around origins in G1 is established during transition from G2/M to G1 in a pre-RC-dependent manner. Importantly, the diminished cell-cycle changes in nucleosome occupancy around origins in the orc1-161 mutant are associated with an abnormal global origin usage profile, suggesting that proper establishment of nucleosome occupancy around origins is a critical step for regulation of global origin activities. Our work thus establishes nucleosome occupancy as a novel and key chromatin parameter for proper origin regulation.

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“…The association of CDC6 and CDT1 to the ORC is a prerequisite for the loading of a hexameric complex containing six different minichromosome maintenance proteins (MCM 2-7) to chromatin, licensing the origin for a new replication round. At this stage, MCMs that are members of the AAA+ class of ATPases (22) acquire their helicase activity to unwind DNA. The molecular components and the basic interaction of pre-RC components are strikingly conserved from yeast to metazoa.…”

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confidence: 99%

KLF15 negatively regulates estrogen-induced epithelial cell proliferation by inhibition of DNA replication licensing

Ray

Pollard²

2012

Proc. Natl. Acad. Sci. U.S.A.

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In the epithelial compartment of the uterus, estradiol-17β (E 2 ) induces cell proliferation while progesterone (P 4 ) inhibits this response and causes differentiation of the cells. In this study, we identified the mechanism whereby E 2 and P 4 reciprocally regulate the expression of minichromosome maintenance (MCM)-2, a protein that is an essential component of the hexameric MCM-2 to 7 complex required for DNA synthesis initiation. We show in the uterine epithelium that Kruppel-like transcription (KLF) factors, KLF 4 and 15, are inversely expressed; most importantly, they bind to the Mcm2 promoter under the regulation of E 2 and P 4 E 2 , respectively. After P 4 E 2 exposure and in contrast to E 2 treated mice, the Mcm2 promoter displays increased histone 3 (H3) methylation and the recruitment of histone deacetylase 1 and 3 with the concomitant deacetylation of H3. This increased methylation and decreased acetylation is associated with an inhibition of RNA polymerase II binding, indicating an inactive Mcm2 promoter following P 4 E 2 treatment. Using transient transfection assays in the Ishikawa endometrial cell line, we demonstrate that Mcm2 promoter activity is hormonally stimulated by E 2 and that KLF15 inhibits this E 2 enhanced transcription. KLF15 expression also blocks Ishikawa cell proliferation through inhibition of MCM2 protein level. Importantly, in vivo expression of KLF15 in an estrogenized uterus mimics P 4 's action by inhibiting E 2 -induced uterine epithelial MCM-2 expression and DNA synthesis. KLF15 is therefore a downstream physiological mediator of progesterone's cell cycle inhibitory action in the uterine epithelium.endometrium | implantation | menstrual cycle | endometriosis E stradiol-17β (E 2 ) and progesterone (P 4 ) directs uterine preparation for pregnancy. These hormones synthesized cyclically in humans cause a sequential series of proliferative and differentiation events in the uterine stroma and epithelium that result in the epithelium being receptive to the blastocyst for attachment and subsequent implantation (1). Despite this close control of uterine cell proliferation, aberrant proliferative conditions of the human endometrium are common. For example, endometrial polyps and endometriosis are caused by inappropriate proliferation of the uterus, while unopposed estrogen stimulation is associated with menstrual irregularities and endometrial hyperplasia/adenocarcinoma (2). Endometrial cancer is the most common female genital tract malignancy and is responsible for 6% of cancer deaths of women in the United States and more worldwide (3). However, the molecular mechanisms underlying these pathologies are still obscure, as are the molecular mechanisms involved in normal hormonal regulation of cell proliferation in the endometrium, which is essential for successful pregnancy (4).The mouse uterus provides a unique in vivo model to study the regulation of epithelial cell proliferation as the physiological actions of E 2 and P 4 E 2 can be recapitulated in ovariectomized animals by treat...

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Concerted Loading of Mcm2–7 Double Hexamers around DNA during DNA Replication Origin Licensing

Cited by 608 publications

References 74 publications

Spatio-temporal regulation of the human licensing factor Cdc6 in replication and mitosis

Spatio-temporal regulation of the human licensing factor Cdc6 in replication and mitosis

Nucleosome occupancy as a novel chromatin parameter for replication origin functions

KLF15 negatively regulates estrogen-induced epithelial cell proliferation by inhibition of DNA replication licensing

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