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Defining the sequence requirements for the positioning of base J in DNA using SMRT sequencing
Base J (β-D-glucosyl-hydroxymethyluracil) replaces 1% of T in the Leishmania genome and is only found in telomeric repeats (99%) and in regions where transcription starts and stops. This highly restricted distribution must be co-determined by the thymidine hydroxylases (JBP1 and JBP2) that catalyze...
Autores principales: | , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Oxford University Press
2015
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4344527/ https://www.ncbi.nlm.nih.gov/pubmed/25662217 http://dx.doi.org/10.1093/nar/gkv095 |
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author | Genest, Paul-Andre Baugh, Loren Taipale, Alex Zhao, Wanqi Jan, Sabrina van Luenen, Henri G.A.M. Korlach, Jonas Clark, Tyson Luong, Khai Boitano, Matthew Turner, Steve Myler, Peter J. Borst, Piet |
author_facet | Genest, Paul-Andre Baugh, Loren Taipale, Alex Zhao, Wanqi Jan, Sabrina van Luenen, Henri G.A.M. Korlach, Jonas Clark, Tyson Luong, Khai Boitano, Matthew Turner, Steve Myler, Peter J. Borst, Piet |
author_sort | Genest, Paul-Andre |
collection | PubMed |
description | Base J (β-D-glucosyl-hydroxymethyluracil) replaces 1% of T in the Leishmania genome and is only found in telomeric repeats (99%) and in regions where transcription starts and stops. This highly restricted distribution must be co-determined by the thymidine hydroxylases (JBP1 and JBP2) that catalyze the initial step in J synthesis. To determine the DNA sequences recognized by JBP1/2, we used SMRT sequencing of DNA segments inserted into plasmids grown in Leishmania tarentolae. We show that SMRT sequencing recognizes base J in DNA. Leishmania DNA segments that normally contain J also picked up J when present in the plasmid, whereas control sequences did not. Even a segment of only 10 telomeric (GGGTTA) repeats was modified in the plasmid. We show that J modification usually occurs at pairs of Ts on opposite DNA strands, separated by 12 nucleotides. Modifications occur near G-rich sequences capable of forming G-quadruplexes and JBP2 is needed, as it does not occur in JBP2-null cells. We propose a model whereby de novo J insertion is mediated by JBP2. JBP1 then binds to J and hydroxylates another T 13 bp downstream (but not upstream) on the complementary strand, allowing JBP1 to maintain existing J following DNA replication. |
format | Online Article Text |
id | pubmed-4344527 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | Oxford University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-43445272015-03-17 Defining the sequence requirements for the positioning of base J in DNA using SMRT sequencing Genest, Paul-Andre Baugh, Loren Taipale, Alex Zhao, Wanqi Jan, Sabrina van Luenen, Henri G.A.M. Korlach, Jonas Clark, Tyson Luong, Khai Boitano, Matthew Turner, Steve Myler, Peter J. Borst, Piet Nucleic Acids Res Gene regulation, Chromatin and Epigenetics Base J (β-D-glucosyl-hydroxymethyluracil) replaces 1% of T in the Leishmania genome and is only found in telomeric repeats (99%) and in regions where transcription starts and stops. This highly restricted distribution must be co-determined by the thymidine hydroxylases (JBP1 and JBP2) that catalyze the initial step in J synthesis. To determine the DNA sequences recognized by JBP1/2, we used SMRT sequencing of DNA segments inserted into plasmids grown in Leishmania tarentolae. We show that SMRT sequencing recognizes base J in DNA. Leishmania DNA segments that normally contain J also picked up J when present in the plasmid, whereas control sequences did not. Even a segment of only 10 telomeric (GGGTTA) repeats was modified in the plasmid. We show that J modification usually occurs at pairs of Ts on opposite DNA strands, separated by 12 nucleotides. Modifications occur near G-rich sequences capable of forming G-quadruplexes and JBP2 is needed, as it does not occur in JBP2-null cells. We propose a model whereby de novo J insertion is mediated by JBP2. JBP1 then binds to J and hydroxylates another T 13 bp downstream (but not upstream) on the complementary strand, allowing JBP1 to maintain existing J following DNA replication. Oxford University Press 2015-02-27 2015-02-06 /pmc/articles/PMC4344527/ /pubmed/25662217 http://dx.doi.org/10.1093/nar/gkv095 Text en © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research. http://creativecommons.org/licenses/by-nc/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com |
spellingShingle | Gene regulation, Chromatin and Epigenetics Genest, Paul-Andre Baugh, Loren Taipale, Alex Zhao, Wanqi Jan, Sabrina van Luenen, Henri G.A.M. Korlach, Jonas Clark, Tyson Luong, Khai Boitano, Matthew Turner, Steve Myler, Peter J. Borst, Piet Defining the sequence requirements for the positioning of base J in DNA using SMRT sequencing |
title | Defining the sequence requirements for the positioning of base J in DNA using SMRT sequencing |
title_full | Defining the sequence requirements for the positioning of base J in DNA using SMRT sequencing |
title_fullStr | Defining the sequence requirements for the positioning of base J in DNA using SMRT sequencing |
title_full_unstemmed | Defining the sequence requirements for the positioning of base J in DNA using SMRT sequencing |
title_short | Defining the sequence requirements for the positioning of base J in DNA using SMRT sequencing |
title_sort | defining the sequence requirements for the positioning of base j in dna using smrt sequencing |
topic | Gene regulation, Chromatin and Epigenetics |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4344527/ https://www.ncbi.nlm.nih.gov/pubmed/25662217 http://dx.doi.org/10.1093/nar/gkv095 |
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