Nuclear distribution and chromatin association of DNA polymerase α-primase is affected by TEV protease cleavage of Cdc23 (Mcm10) in fission yeast
<p style="text-align:justify;"> <b>Background:</b> Cdc23/Mcm10 is required for the initiation and elongation steps of DNA replication but its biochemical function is unclear. Here, we probe its function using a novel approach in fission yeast, involving Cdc23 cleavage by...
| Main Authors: | , , , , |
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| Formato: | Journal article |
| Idioma: | English |
| Publicado: |
BioMed Central
2005
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| _version_ | 1826303954382749696 |
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| author | Yang, X Gregan, J Lindner, K Young, H Kearsey, S |
| author_facet | Yang, X Gregan, J Lindner, K Young, H Kearsey, S |
| author_sort | Yang, X |
| collection | OXFORD |
| description | <p style="text-align:justify;"> <b>Background:</b> Cdc23/Mcm10 is required for the initiation and elongation steps of DNA replication but its biochemical function is unclear. Here, we probe its function using a novel approach in fission yeast, involving Cdc23 cleavage by the TEV protease.<br/><br/> <b>Results:</b> Insertion of a TEV protease cleavage site into Cdc23 allows in vivo removal of the C-terminal 170 aa of the protein by TEV protease induction, resulting in an S phase arrest. This C-terminal fragment of Cdc23 is not retained in the nucleus after cleavage, showing that it lacks a nuclear localization signal and ability to bind to chromatin. Using an in situ chromatin binding procedure we have determined how the S phase chromatin association of DNA polymerase α-primase and the GINS (Sld5-Psf1-Psf2-Psf3) complex is affected by Cdc23 inactivation. The chromatin binding and sub-nuclear distribution of DNA primase catalytic subunit (Spp1) is affected by Cdc23 cleavage and also by inactivation of Cdc23 using a degron allele, implying that DNA polymerase α-primase function is dependent on Cdc23. In contrast to the effect on Spp1, the chromatin association of the Psf2 subunit of the GINS complex is not affected by Cdc23 inactivation.<br/><br/> <b>Conclusion:</b> An important function of Cdc23 in the elongation step of DNA replication may be to assist in the docking of DNA polymerase α-primase to chromatin. </p> |
| first_indexed | 2024-03-07T06:10:31Z |
| format | Journal article |
| id | oxford-uuid:ef52eb52-e460-42c9-8e22-656121b3571a |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-07T06:10:31Z |
| publishDate | 2005 |
| publisher | BioMed Central |
| record_format | dspace |
| spelling | oxford-uuid:ef52eb52-e460-42c9-8e22-656121b3571a2022-03-27T11:39:25ZNuclear distribution and chromatin association of DNA polymerase α-primase is affected by TEV protease cleavage of Cdc23 (Mcm10) in fission yeastJournal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:ef52eb52-e460-42c9-8e22-656121b3571aEnglishSymplectic Elements at OxfordBioMed Central2005Yang, XGregan, JLindner, KYoung, HKearsey, S <p style="text-align:justify;"> <b>Background:</b> Cdc23/Mcm10 is required for the initiation and elongation steps of DNA replication but its biochemical function is unclear. Here, we probe its function using a novel approach in fission yeast, involving Cdc23 cleavage by the TEV protease.<br/><br/> <b>Results:</b> Insertion of a TEV protease cleavage site into Cdc23 allows in vivo removal of the C-terminal 170 aa of the protein by TEV protease induction, resulting in an S phase arrest. This C-terminal fragment of Cdc23 is not retained in the nucleus after cleavage, showing that it lacks a nuclear localization signal and ability to bind to chromatin. Using an in situ chromatin binding procedure we have determined how the S phase chromatin association of DNA polymerase α-primase and the GINS (Sld5-Psf1-Psf2-Psf3) complex is affected by Cdc23 inactivation. The chromatin binding and sub-nuclear distribution of DNA primase catalytic subunit (Spp1) is affected by Cdc23 cleavage and also by inactivation of Cdc23 using a degron allele, implying that DNA polymerase α-primase function is dependent on Cdc23. In contrast to the effect on Spp1, the chromatin association of the Psf2 subunit of the GINS complex is not affected by Cdc23 inactivation.<br/><br/> <b>Conclusion:</b> An important function of Cdc23 in the elongation step of DNA replication may be to assist in the docking of DNA polymerase α-primase to chromatin. </p> |
| spellingShingle | Yang, X Gregan, J Lindner, K Young, H Kearsey, S Nuclear distribution and chromatin association of DNA polymerase α-primase is affected by TEV protease cleavage of Cdc23 (Mcm10) in fission yeast |
| title | Nuclear distribution and chromatin association of DNA polymerase α-primase is affected by TEV protease cleavage of Cdc23 (Mcm10) in fission yeast |
| title_full | Nuclear distribution and chromatin association of DNA polymerase α-primase is affected by TEV protease cleavage of Cdc23 (Mcm10) in fission yeast |
| title_fullStr | Nuclear distribution and chromatin association of DNA polymerase α-primase is affected by TEV protease cleavage of Cdc23 (Mcm10) in fission yeast |
| title_full_unstemmed | Nuclear distribution and chromatin association of DNA polymerase α-primase is affected by TEV protease cleavage of Cdc23 (Mcm10) in fission yeast |
| title_short | Nuclear distribution and chromatin association of DNA polymerase α-primase is affected by TEV protease cleavage of Cdc23 (Mcm10) in fission yeast |
| title_sort | nuclear distribution and chromatin association of dna polymerase α primase is affected by tev protease cleavage of cdc23 mcm10 in fission yeast |
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