CG methylated microarrays identify a novel methylated sequence bound by the CEBPB|ATF4 heterodimer that is active in vivo.
Identifieur interne : 002045 ( Main/Exploration ); précédent : 002044; suivant : 002046CG methylated microarrays identify a novel methylated sequence bound by the CEBPB|ATF4 heterodimer that is active in vivo.
Auteurs : Ishminder K. Mann [Canada] ; Raghunath Chatterjee ; Jianfei Zhao ; Ximiao He ; Matthew T. Weirauch ; Timothy R. Hughes ; Charles VinsonSource :
- Genome research [ 1549-5469 ] ; 2013.
Descripteurs français
- KwdFr :
- Animaux, Facteur de transcription ATF-4 (), Facteur de transcription ATF-4 (métabolisme), Facteurs de transcription (métabolisme), Femelle, Fibroblastes, Ilots CpG, Liaison aux protéines (), Matrices de scores, Motifs nucléotidiques, Multimérisation de protéines, Méthylation de l'ADN, Protéine bêta de liaison aux séquences stimulatrices de type CCAAT (), Protéine bêta de liaison aux séquences stimulatrices de type CCAAT (métabolisme), Sites de fixation, Souris, Séquence nucléotidique, Thapsigargine (immunologie).
- MESH :
- immunologie : Thapsigargine.
- métabolisme : Facteur de transcription ATF-4, Facteurs de transcription, Protéine bêta de liaison aux séquences stimulatrices de type CCAAT.
- Animaux, Facteur de transcription ATF-4, Femelle, Fibroblastes, Ilots CpG, Liaison aux protéines, Matrices de scores, Motifs nucléotidiques, Multimérisation de protéines, Méthylation de l'ADN, Protéine bêta de liaison aux séquences stimulatrices de type CCAAT, Sites de fixation, Souris, Séquence nucléotidique.
English descriptors
- KwdEn :
- Activating Transcription Factor 4 (chemistry), Activating Transcription Factor 4 (metabolism), Animals, Base Sequence, Binding Sites, CCAAT-Enhancer-Binding Protein-beta (chemistry), CCAAT-Enhancer-Binding Protein-beta (metabolism), CpG Islands, DNA Methylation, Female, Fibroblasts, Mice, Nucleotide Motifs, Position-Specific Scoring Matrices, Protein Binding (drug effects), Protein Multimerization, Thapsigargin (immunology), Transcription Factors (metabolism).
- MESH :
- chemical , chemistry : Activating Transcription Factor 4, CCAAT-Enhancer-Binding Protein-beta.
- chemical , immunology : Thapsigargin.
- chemical , metabolism : Activating Transcription Factor 4, CCAAT-Enhancer-Binding Protein-beta, Transcription Factors.
- drug effects : Protein Binding.
- Animals, Base Sequence, Binding Sites, CpG Islands, DNA Methylation, Female, Fibroblasts, Mice, Nucleotide Motifs, Position-Specific Scoring Matrices, Protein Multimerization.
Abstract
To evaluate the effect of CG methylation on DNA binding of sequence-specific B-ZIP transcription factors (TFs) in a high-throughput manner, we enzymatically methylated the cytosine in the CG dinucleotide on protein binding microarrays. Two Agilent DNA array designs were used. One contained 40,000 features using de Bruijn sequences where each 8-mer occurs 32 times in various positions in the DNA sequence. The second contained 180,000 features with each CG containing 8-mer occurring three times. The first design was better for identification of binding motifs, while the second was better for quantification. Using this novel technology, we show that CG methylation enhanced binding for CEBPA and CEBPB and inhibited binding for CREB, ATF4, JUN, JUND, CEBPD, and CEBPG. The CEBPB|ATF4 heterodimer bound a novel motif CGAT|GCAA 10-fold better when methylated. The electrophoretic mobility shift assay (EMSA) confirmed these results. CEBPB ChIP-seq data using primary female mouse dermal fibroblasts with 50× methylome coverage for each strand indicate that the methylated sequences well-bound on the arrays are also bound in vivo. CEBPB bound 39% of the methylated canonical 10-mers ATTGC|GCAAT in the mouse genome. After ATF4 protein induction by thapsigargin which results in ER stress, CEBPB binds methylated CGAT|GCAA in vivo, recapitulating what was observed on the arrays. This methodology can be used to identify new methylated DNA sequences preferentially bound by TFs, which may be functional in vivo.
DOI: 10.1101/gr.146654.112
PubMed: 23590861
Affiliations:
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Mann</name><affiliation wicri:level="4"><nlm:affiliation>Donnelly Centre, University of Toronto, Toronto, Canada.</nlm:affiliation><country xml:lang="fr">Canada</country><wicri:regionArea>Donnelly Centre, University of Toronto, Toronto</wicri:regionArea><orgName type="university">Université de Toronto</orgName><placeName><settlement type="city">Toronto</settlement><region type="state">Ontario</region></placeName></affiliation></author><author><name sortKey="Chatterjee, Raghunath" sort="Chatterjee, Raghunath" uniqKey="Chatterjee R" first="Raghunath" last="Chatterjee">Raghunath Chatterjee</name></author><author><name sortKey="Zhao, Jianfei" sort="Zhao, Jianfei" uniqKey="Zhao J" first="Jianfei" last="Zhao">Jianfei Zhao</name></author><author><name sortKey="He, Ximiao" sort="He, Ximiao" uniqKey="He X" first="Ximiao" last="He">Ximiao He</name></author><author><name sortKey="Weirauch, Matthew T" sort="Weirauch, Matthew T" uniqKey="Weirauch M" first="Matthew T" last="Weirauch">Matthew T. Weirauch</name></author><author><name sortKey="Hughes, Timothy R" sort="Hughes, Timothy R" uniqKey="Hughes T" first="Timothy R" last="Hughes">Timothy R. Hughes</name></author><author><name sortKey="Vinson, Charles" sort="Vinson, Charles" uniqKey="Vinson C" first="Charles" last="Vinson">Charles Vinson</name></author></analytic><series><title level="j">Genome research</title><idno type="eISSN">1549-5469</idno><imprint><date when="2013" type="published">2013</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Activating Transcription Factor 4 (chemistry)</term><term>Activating Transcription Factor 4 (metabolism)</term><term>Animals</term><term>Base Sequence</term><term>Binding Sites</term><term>CCAAT-Enhancer-Binding Protein-beta (chemistry)</term><term>CCAAT-Enhancer-Binding Protein-beta (metabolism)</term><term>CpG Islands</term><term>DNA Methylation</term><term>Female</term><term>Fibroblasts</term><term>Mice</term><term>Nucleotide Motifs</term><term>Position-Specific Scoring Matrices</term><term>Protein Binding (drug effects)</term><term>Protein Multimerization</term><term>Thapsigargin (immunology)</term><term>Transcription Factors (metabolism)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>Animaux</term><term>Facteur de transcription ATF-4 ()</term><term>Facteur de transcription ATF-4 (métabolisme)</term><term>Facteurs de transcription (métabolisme)</term><term>Femelle</term><term>Fibroblastes</term><term>Ilots CpG</term><term>Liaison aux protéines ()</term><term>Matrices de scores</term><term>Motifs nucléotidiques</term><term>Multimérisation de protéines</term><term>Méthylation de l'ADN</term><term>Protéine bêta de liaison aux séquences stimulatrices de type CCAAT ()</term><term>Protéine bêta de liaison aux séquences stimulatrices de type CCAAT (métabolisme)</term><term>Sites de fixation</term><term>Souris</term><term>Séquence nucléotidique</term><term>Thapsigargine (immunologie)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>Activating Transcription Factor 4</term><term>CCAAT-Enhancer-Binding Protein-beta</term></keywords><keywords scheme="MESH" type="chemical" qualifier="immunology" xml:lang="en"><term>Thapsigargin</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Activating Transcription Factor 4</term><term>CCAAT-Enhancer-Binding Protein-beta</term><term>Transcription Factors</term></keywords><keywords scheme="MESH" qualifier="drug effects" xml:lang="en"><term>Protein Binding</term></keywords><keywords scheme="MESH" qualifier="immunologie" xml:lang="fr"><term>Thapsigargine</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Facteur de transcription ATF-4</term><term>Facteurs de transcription</term><term>Protéine bêta de liaison aux séquences stimulatrices de type CCAAT</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Animals</term><term>Base Sequence</term><term>Binding Sites</term><term>CpG Islands</term><term>DNA Methylation</term><term>Female</term><term>Fibroblasts</term><term>Mice</term><term>Nucleotide Motifs</term><term>Position-Specific Scoring Matrices</term><term>Protein Multimerization</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Animaux</term><term>Facteur de transcription ATF-4</term><term>Femelle</term><term>Fibroblastes</term><term>Ilots CpG</term><term>Liaison aux protéines</term><term>Matrices de scores</term><term>Motifs nucléotidiques</term><term>Multimérisation de protéines</term><term>Méthylation de l'ADN</term><term>Protéine bêta de liaison aux séquences stimulatrices de type CCAAT</term><term>Sites de fixation</term><term>Souris</term><term>Séquence nucléotidique</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">To evaluate the effect of CG methylation on DNA binding of sequence-specific B-ZIP transcription factors (TFs) in a high-throughput manner, we enzymatically methylated the cytosine in the CG dinucleotide on protein binding microarrays. Two Agilent DNA array designs were used. One contained 40,000 features using de Bruijn sequences where each 8-mer occurs 32 times in various positions in the DNA sequence. The second contained 180,000 features with each CG containing 8-mer occurring three times. The first design was better for identification of binding motifs, while the second was better for quantification. Using this novel technology, we show that CG methylation enhanced binding for CEBPA and CEBPB and inhibited binding for CREB, ATF4, JUN, JUND, CEBPD, and CEBPG. The CEBPB|ATF4 heterodimer bound a novel motif CGAT|GCAA 10-fold better when methylated. The electrophoretic mobility shift assay (EMSA) confirmed these results. CEBPB ChIP-seq data using primary female mouse dermal fibroblasts with 50× methylome coverage for each strand indicate that the methylated sequences well-bound on the arrays are also bound in vivo. CEBPB bound 39% of the methylated canonical 10-mers ATTGC|GCAAT in the mouse genome. After ATF4 protein induction by thapsigargin which results in ER stress, CEBPB binds methylated CGAT|GCAA in vivo, recapitulating what was observed on the arrays. This methodology can be used to identify new methylated DNA sequences preferentially bound by TFs, which may be functional in vivo.</div></front></TEI><affiliations><list><country><li>Canada</li></country><region><li>Ontario</li></region><settlement><li>Toronto</li></settlement><orgName><li>Université de Toronto</li></orgName></list><tree><noCountry><name sortKey="Chatterjee, Raghunath" sort="Chatterjee, Raghunath" uniqKey="Chatterjee R" first="Raghunath" last="Chatterjee">Raghunath Chatterjee</name><name sortKey="He, Ximiao" sort="He, Ximiao" uniqKey="He X" first="Ximiao" last="He">Ximiao He</name><name sortKey="Hughes, Timothy R" sort="Hughes, Timothy R" uniqKey="Hughes T" first="Timothy R" last="Hughes">Timothy R. Hughes</name><name sortKey="Vinson, Charles" sort="Vinson, Charles" uniqKey="Vinson C" first="Charles" last="Vinson">Charles Vinson</name><name sortKey="Weirauch, Matthew T" sort="Weirauch, Matthew T" uniqKey="Weirauch M" first="Matthew T" last="Weirauch">Matthew T. Weirauch</name><name sortKey="Zhao, Jianfei" sort="Zhao, Jianfei" uniqKey="Zhao J" first="Jianfei" last="Zhao">Jianfei Zhao</name></noCountry><country name="Canada"><region name="Ontario"><name sortKey="Mann, Ishminder K" sort="Mann, Ishminder K" uniqKey="Mann I" first="Ishminder K" last="Mann">Ishminder K. Mann</name></region></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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