Temporal profiling of the chromatin proteome reveals system-wide responses to replication inhibition.
Although the replication, expression, and maintenance of DNA are well-studied processes, the way that they are coordinated is poorly understood. Here, we report an analysis of the changing association of proteins with chromatin (the chromatin proteome) during progression through interphase of the cell cycle. Sperm nuclei were incubated in Xenopus egg extracts, and chromatin-associated proteins were analyzed by mass spectrometry at different times. Approximately 75% of the proteins varied in abundance on chromatin by more than 15%, suggesting that the chromatin proteome is highly dynamic. Proteins were then assigned to one of 12 different clusters on the basis of their pattern of chromatin association. Each cluster contained functional groups of proteins involved in different nuclear processes related to progression through interphase. We also blocked DNA replication by inhibiting either replication licensing or S phase CDK activity. This revealed an unexpectedly broad system-wide effect on the chromatin proteome, indicating that the response to replication inhibition extends to many other functional modules in addition to the replication machinery. Several proteins that respond to replication inhibition (including nuclear pore proteins) coprecipitated with the Mcm2-7 licensing complex on chromatin, suggesting that Mcm2-7 play a central role in coordinating nuclear structure with DNA replication.
Khoudoli
Guennadi A
GA
Wellcome Trust Centre for Gene Regulation and Expression, University of Dundee, Dundee DD1 5EH, United Kingdom.
Gillespie
Peter J
PJ
Stewart
Graeme
G
Andersen
Jens S
JS
Swedlow
Jason R
JR
Blow
J Julian
JJ
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism*; D004261:DNA Replication / Q000502:physiology; D004268:DNA-Binding Proteins / Q000378:metabolism*; D064248:Geminin; D007399:Interphase / Q000502:physiology*; D064111:Minichromosome Maintenance Complex Component 2; D064126:Minichromosome Maintenance Complex Component 3; D009687:Nuclear Proteins / Q000378:metabolism; D047428:Protein Kinase Inhibitors; D020543:Proteome; D011687:Purines; D000077546:Roscovitine; D013997:Time Factors; D014981:Xenopus; D029867:Xenopus Proteins / Q000378:metabolism*
D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; C577998:GMNN protein, Xenopus; D064248:Geminin; D009687:Nuclear Proteins; D047428:Protein Kinase Inhibitors; D020543:Proteome; D011687:Purines; D029867:Xenopus Proteins; D000077546:Roscovitine; C507762:MCM3 protein, Xenopus; C102186:Mcm2 protein, Xenopus; D064111:Minichromosome Maintenance Complex Component 2; D064126:Minichromosome Maintenance Complex Component 3