Selective instability of Orc1 protein accounts for the absence of functional origin recognition complexes during the M-G1 transition in mammals

被引:65
作者
Natale, DA [1 ]
Li, CJ [1 ]
Sun, WH [1 ]
DePamphilis, ML [1 ]
机构
[1] NICHHD, NIH, Bethesda, MD 20892 USA
关键词
DNA replication; initiation; origin recognition complex;
D O I
10.1093/emboj/19.11.2728
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
To investigate the events leading to initiation of DNA replication in mammalian chromosomes, the time when hamster origin recognition complexes (ORCs) became functional was related to the time when Orc1, Orc2 and Mcm3 proteins became stably bound to hamster chromatin, Functional ORCs, defined as those able to initiate DNA replication, were absent during mitosis and early G(1) phase, and reappeared as cells progressed through G(1) phase. Immunoblotting analysis revealed that hamster Orc1 and Orc2 proteins were present in nuclei at equivalent concentrations throughout the cell cycle, but only Orc2 was stably bound to chromatin, Orc1 and McmS were easily eluted from chromatin during mitosis and early G(1) phase, but became stably bound during mid-G(1) phase, concomitant with the appearance of a functional pre-replication complex at a hamster replication origin. Since hamster Ore proteins are closely related to their human and mouse homologs, the unexpected behavior of hamster Orc1 provides a novel mechanism in mammals for delaying assembly of prereplication complexes until mitosis is complete and a nuclear structure has formed.
引用
收藏
页码:2728 / 2738
页数:11
相关论文
共 56 条
[11]  
DePamphilis ML, 1999, BIOESSAYS, V21, P5, DOI 10.1002/(SICI)1521-1878(199901)21:1<5::AID-BIES2>3.0.CO
[12]  
2-6
[13]   2 STEPS IN THE ASSEMBLY OF COMPLEXES AT YEAST REPLICATION ORIGINS IN-VIVO [J].
DIFFLEY, JFX ;
COCKER, JH ;
DOWELL, SJ ;
ROWLEY, A .
CELL, 1994, 78 (02) :303-316
[14]   Mcm2, but not RPA, is a component of the mammalian early G1-phase prereplication complex [J].
Dimitrova, DS ;
Todorov, IT ;
Melendy, T ;
Gilbert, DM .
JOURNAL OF CELL BIOLOGY, 1999, 146 (04) :709-722
[15]   Cyclin A-dependent kinase activity affects chromatin binding of ORC, Cdc6, and MCM in egg extracts of Xenopus laevis [J].
Findeisen, M ;
El-Denary, M ;
Kapitza, T ;
Graf, R ;
Strausfeld, U .
EUROPEAN JOURNAL OF BIOCHEMISTRY, 1999, 264 (02) :415-426
[16]   Cell cycle dependent topological changes of chromosomal replication origins in Saccharomyces cerevisiae [J].
Fujita, M ;
Hori, Y ;
Shirahige, K ;
Tsurimoto, T ;
Yoshikawa, H ;
Obuse, C .
GENES TO CELLS, 1998, 3 (11) :737-749
[17]  
Fujita M, 1997, J BIOL CHEM, V272, P10928
[18]  
GILBERT DM, 1995, MOL CELL BIOL, V15, P2942
[19]   MAPPING REPLICATION UNITS IN ANIMAL-CELLS [J].
HANDELI, S ;
KLAR, A ;
MEUTH, M ;
CEDAR, H .
CELL, 1989, 57 (06) :909-920
[20]  
Harlow E., 1999, Using Antibodies: A Laboratory Manual