International Journal of Biological Sciences

Impact factor
3.873

ISSN 1449-2288

News feeds of IJBS published articles
My Manuscript
My Account

Journal of Biomedicinenew

Theranostics

International Journal of Medical Sciences

Journal of Cancer

Oncomedicine

Journal of Genomics

Journal of Bone and Joint Infection (JBJI)

Nanotheranostics

Journal of Genomics now in PubMed/PubMed Central. Submit manuscript...

PubMed Central Indexed in Journal Impact Factor

Int J Biol Sci 2011; 7(2):193-208. doi:10.7150/ijbs.7.193

Research Paper

Interactions of MCP1 with Components of the Replication Machinery in Mammalian Cells

Elsa Bronze-da-Rocha1,2,4,✉, Chii-Mei Lin4, Tsutomu Shimura3,4, Mirit I. Aladjem4

1. Departamento de Ciências Biológicas, Laboratório de Bioquímica, Faculdade de Farmácia da Universidade do Porto, Portugal
2. Instituto de Biologia Molecular e Celular da Universidade do Porto, Portugal
3. Department of Pathology, Institute of Development Aging and Cancer, Tohoku University, Seiryo-machi, Sendai, Japan
4. Laboratory of Molecular Pharmacology, CCR/NCI/NIH, Bethesda, USA

Abstract

Eukaryotic DNA replication starts with the assembly of a pre-replication complex (pre-RC) at replication origins. We have previously demonstrated that Metaphase Chromosome Protein 1 (MCP1) is involved in the early events of DNA replication. Here we show that MCP1 associates with proteins that are required for the establishment of the pre-replication complex. Reciprocal immunoprecipitation analysis showed that MCP1 interacted with Cdc6, ORC2, ORC4, MCM2, MCM3 and MCM7, with Cdc45 and PCNA. Immunofluorescence studies demonstrated the co-localization of MCP1 with some of those proteins. Moreover, biochemical studies utilizing chromatin-immunoprecipitation (ChIP) revealed that MCP1 preferentially binds replication initiation sites in human cells. Interestingly, although members of the pre-RC are known to interact with some hallmarks of heterochromatin, our co-immunoprecipitation and immunofluorescence analyses showed that MCP1 did not interact and did not co-localize with heterochromatic proteins including HP1β and MetH3K9. These observations suggest that MCP1 is associated with replication factors required for the initiation of DNA replication and binds to the initiation sites in loci that replicate early in S-phase. In addition, immunological assays revealed the association of MCP1 forms with histone H1 variants and mass spectrometry analysis confirmed that MCP1 peptides share common sequences with H1.2 and H1.5 subtypes.

Keywords: MCP1, DNA replication, pre-replication complex, histone H1, interacting proteins

This is an open access article distributed under the terms of the Creative Commons Attribution (CC BY-NC) License. See http://ivyspring.com/terms for full terms and conditions.
How to cite this article:
Bronze-da-Rocha E, Lin CM, Shimura T, Aladjem MI. Interactions of MCP1 with Components of the Replication Machinery in Mammalian Cells. Int J Biol Sci 2011; 7(2):193-208. doi:10.7150/ijbs.7.193. Available from http://www.ijbs.com/v07p0193.htm