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| Status |
Public on Jan 01, 2008 |
| Title |
Origin of replication for Pyrococcus abyssi by ChIP-chip |
| Organism |
Pyrococcus abyssi |
| Experiment type |
Genome binding/occupancy profiling by genome tiling array
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| Summary |
Recent advances in the study of archaeal DNA replication have uncovered defined replication origins (oriC) and demonstrated specific binding of the Cdc6/Orc1 protein and Mcm helicase to oriC in vivo and/or in vitro. The oriC of the hyperthermophilic archaeon Pyrococcus abyssi is characterized by 13 bp repeats, AT-rich regions and an inverted repeat whose precise roles remain unclear. We report here that the 13 bp repeats are widespread in the three Pyrococcus genomes. Nevertheless, by means of chromatin immunoprecipitation coupled with hybridization on a whole genome microarray (ChIP-chip), we found that binding of P. abyssi Cdc6/Orc1 to oriC in vivo was highly specific both in exponential and stationary phases, allowing oriC to be distinguished in the 1.8 M bp genome. ChIP-chip analysis also indicated that a single 13 bp repeat is not sufficient for stable binding of Cdc6/Orc1. Purified Cdc6/Orc1 binds a DNA fragment containing the inverted repeat of oriC with a relatively low affinity, suggesting that multiple clusters of the 13 bp repeat discovered in this study contribute to the stable binding of Cdc6/Orc1 to oriC. Our ChIP-chip analysis revealed that Mcm also binds oriC only in proliferating cells, consistent with its role as a licensing factor. Finally, we found that both Cdc6/Orc1 and Mcm have one additional target site. Notably, Mcm binds constitutively to a GC-rich region containing two rRNA genes and a tRNA gene, suggesting a role of archaeal Mcm in DNA replication and/or transcription of this peculiar region.
Keywords: ChIP-chip
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| Overall design |
Experiments were done using at least three (Cdc6 in exponential and stationary phases, MCM in stationary phase) or four (MCM in exponential phase) independently prepared samples.
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| Contributor(s) |
Matsunaga F, Glatigny A, Mucchielli M, Agier N, Delacroix H, Marisa L, Meno T, Ishino Y, Aggerbeck L, Forterre P |
| Citation(s) |
17452353 |
| |
| Submission date |
Jan 26, 2006 |
| Last update date |
Mar 16, 2012 |
| Contact name |
LAWRENCE AGGERBECK |
| E-mail(s) |
[email protected], [email protected], [email protected]
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| Phone |
33-(0)169823752
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| Fax |
33-(0)169823740
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| Organization name |
Centre National de la Recherche Scientifique
|
| Department |
Centre de Génétique Moléculaire
|
| Lab |
GODMAP
|
| Street address |
Avenue de la Terrasse
|
| City |
Gif-sur-Yvette |
| ZIP/Postal code |
91198 |
| Country |
France |
| |
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| Platforms (1) |
| GPL3384 |
Origin of replication for Pyrococcus abyssi by ChIP-chip |
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| Samples (14)
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| GSM93618 |
Cdc 6/stationary phase replica 4 |
| GSM93619 |
Cdc 6/exponential phase replica 1 |
| GSM93620 |
Cdc 6/exponential phase replica 2 |
| GSM93621 |
Cdc 6/exponential phase replica 4 |
| GSM93622 |
Mcm/ stationary phase/ replica 1 |
| GSM93623 |
Mcm/ stationary phase/ replica 2 |
| GSM93624 |
Mcm/ stationary phase/ replica 3 |
| GSM93625 |
Mcm/ stationary phase/ replica 4 |
| GSM93626 |
Mcm/ exponential phase/ replica 1 |
| GSM93627 |
Mcm/ exponential phase/ replica 2 |
| GSM93628 |
Mcm/ exponential phase/ replica 4 |
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| Relations |
| BioProject |
PRJNA95265 |
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