The High-Affinity Interaction between ORC and DNA that Is Required for Replication Licensing Is Inhibited by 2-Arylquinolin-4-Amines

Gardner, N.J.; Gillespie, Peter J.; Carrington, Jamie T.; Shanks, Emma; McElroy, Stuart P.; Haagensen, Emma J.; Frearson, Julie A.; Woodland, Andrew; Blow, J. Julian

doi:10.1016/j.chembiol.2017.06.019

Cited by 18 publications

(17 citation statements)

References 66 publications

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“…In the presence of geminin, MCM loading is prevented before the second Cdc6 binding, allowing stabilization of ORC-Cdc6-Cdt1, and MCM3-C could promote the dissociation of ORC, Cdc6, and Cdt1. This effect of MCM3-C may be related to previous reports that ORC binding becomes salt-sensitive after the licensing reaction [46,47]. On the other hand, in the absence of geminin, MCM loading reaction could proceed to the second Cdc6 loading step.…”

Section: Possible Role Of Mcm3-c In Mediating the Autoinhibitory Actisupporting

confidence: 77%

MCM interference during licensing of DNA replication inXenopusegg extracts-Possible Role of a C-terminal region of MCM3-

2018

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The minichromosome maintenance (MCM) complex, consisting of six subunits, Mcm2-7, is loaded onto replication origins through loading factors (origin recognition complex [ORC], Cdc6, and Cdt1) and forms an MCM double hexamer that licenses the initiation of DNA replication. Previous studies with Xenopus egg extracts showed that loading factors, especially Cdc6, dissociate from chromatin on MCM loading, but the molecular mechanism and physiological significance remain largely unknown. Using a cell-free system for MCM loading onto plasmid DNA in Xenopus egg extracts, we found that MCM loaded onto DNA prevents DNA binding of the loading factors ORC, Cdc6, and Cdt1. We further report that a peptide of the C-terminal region of MCM3 (MCM3-C), previously implicated in the initial association with ORC/Cdc6 in budding yeast, prevents ORC/Cdc6/Cdt1 binding to DNA in the absence of MCM loading. ATP-γ-S suppresses inhibitory activities of both the MCM loaded onto DNA and the MCM3-C peptide. Other soluble factors in the extract, but neither MCM nor Cdt1, are required for the activity. Conservation of the amino acid sequences of MCM3-C and its activity in vertebrates implies a novel negative autoregulatory mechanism that interferes with MCM loading in the vicinity of licensed origins to ensure proper origin licensing.

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Section: Possible Role Of Mcm3-c In Mediating the Autoinhibitory Actisupporting

confidence: 77%

MCM interference during licensing of DNA replication inXenopusegg extracts-Possible Role of a C-terminal region of MCM3-

2018

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“…This is an exciting result, as it highlights a novel and essential role for Mcm2 C‐terminus in a late step of MCM2‐7 double‐hexamer formation (Barbon et al , in preparation), a process that is only poorly understood. Moreover, the CLMS data show that the Mcm2 C‐terminus is involved in a network of interactions with flexible domains of Orc6, Orc2 and Mcm5, indicating dynamics at the Mcm2‐Mcm5 DNA entry gate (Samel et al , ), which could represent an ideal target for the development of inhibitors with potential as anti‐cancer therapy (Gardner et al , ), as dynamic interactions have improved druggability characteristics over stable protein interactions (Ulucan et al , ; Jubb et al , ). Indeed, expressing Mcm2‐7ΔC2 causes dominant lethality (Fig C).…”

Section: Resultsmentioning

confidence: 99%

An integrated workflow for crosslinking mass spectrometry

Mendes

Fischer

Chen

et al. 2019

Molecular Systems Biology

148

150

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We present a concise workflow to enhance the mass spectrometric detection of crosslinked peptides by introducing sequential digestion and the crosslink identification software xiSEARCH. Sequential digestion enhances peptide detection by selective shortening of long tryptic peptides. We demonstrate our simple 12‐fraction protocol for crosslinked multi‐protein complexes and cell lysates, quantitative analysis, and high‐density crosslinking, without requiring specific crosslinker features. This overall approach reveals dynamic protein–protein interaction sites, which are accessible, have fundamental functional relevance and are therefore ideally suited for the development of small molecule inhibitors.

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“…One of the most important and wellknown factors is the MCM2-7 complex, which is a helical helix composed of 6 subunits of different MCM proteins. At the MCM2/5 unit, there is a gap which allows the entry of the DNA strand into the ring shape complex of MCM2-7 (3,4). This complex contributes to the formation of replication forks and determines where DNA duplication will proceed.…”

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confidence: 99%

MCM5 Expression Is Associated With the Grade of Malignancy and Ki-67 Antigen in LSCC

et al. 2019

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Background/Aim: The minichromosome maintenance proteins (MCMs) may be potential biomarkers of cancer cell proliferation. They are essential to initiate DNA replication. The aim of the study was to investigate the level of MCM5 expression in benign lesions (BLs) and laryngeal squamous cell cancer (LSCC). Materials and Methods: Immunohistochemical (IHC) analysis was carried out on 83 LSCCs and 10 BLs. Western-blot, immunofluorescence analysis (IF) and real-time PCR (RT-PCR) were performed using HEp-2 cancer cells and HaCaT keratinocytes. Results: The expression of MCM5 was higher in LSCC than in the BLs (p<0.0001) and was higher in subsequent malignancies of LSCC. Positive correlations were demonstrated between the expression levels of MCM5 and the Ki-67 antigen. In vitro studies have confirmed that the expression of MCM5 is elevated in cancer cells. Conclusion: MCM5 protein may be used as a potential marker of cancer cell proliferation in LSCC.

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The High-Affinity Interaction between ORC and DNA that Is Required for Replication Licensing Is Inhibited by 2-Arylquinolin-4-Amines

Cited by 18 publications

References 66 publications

MCM interference during licensing of DNA replication inXenopusegg extracts-Possible Role of a C-terminal region of MCM3-

MCM interference during licensing of DNA replication inXenopusegg extracts-Possible Role of a C-terminal region of MCM3-

An integrated workflow for crosslinking mass spectrometry

MCM5 Expression Is Associated With the Grade of Malignancy and Ki-67 Antigen in LSCC

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