Human Mcm proteins at a replication origin during the G1 to S phase transition

AUTOR(ES)

Schaarschmidt, Daniel

FONTE

Oxford University Press

RESUMO

Previous work with yeast cells and with Xenopus egg extracts had shown that eukaryotic pre-replication complexes assemble on chromatin in a step-wise manner whereby specific loading factors promote the recruitment of essential Mcm proteins at pre-bound origin recognition complexes (ORC with proteins Orc1p–Orc6p). While the order of assembly—Mcm binding follows ORC binding—seems to be conserved in cycling mammalian cells in culture, it has not been determined whether mammalian Mcm proteins associate with ORC-bearing chromatin sites. We have used a chromatin immunoprecipitation approach to investigate the site of Mcm binding in a genomic region that has previously been shown to contain an ORC-binding site and an origin of replication. Using chromatin from HeLa cells in G1 phase, antibodies against Orc2p as well as antibodies against Mcm proteins specifically immunoprecipitate chromatin enriched for a DNA region that includes a replication origin. However, with chromatin from cells in S phase, only Orc2p-specific antibodies immunoprecipitate the origin-containing DNA region while Mcm-specific antibodies immunoprecipitate chromatin with DNA from all parts of the genomic region investigated. Thus, human Mcm proteins first assemble at or adjacent to bound ORC and move to other sites during genome replication.

Documentos Relacionados

• G1 to S phase cell cycle transition in somatic and embryonic stem cells
• Human cytomegalovirus infection inhibits cell cycle progression at multiple points, including the transition from G1 to S.
• Transcription-dependent induction of G1 phase during the zebra fish midblastula transition.
• Spatial distribution and specification of mammalian replication origins during G1 phase
• Human cyclin E, a nuclear protein essential for the G1-to-S phase transition.