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| Status |
Public on Dec 01, 2023 |
| Title |
Unified analysis of DNA methylation, hydroxymethylation and chromatin accessibility through targeted DNA labeling [TOP-Seq] |
| Organism |
Mus musculus |
| Experiment type |
Other
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| Summary |
Epigenetic regulation depends on complex interactions between different epigenetic factors, such as DNA methylation and chromatin accessibility. As the gene activation potential cannot be predicted by looking at chromatin accessibility or DNA modification alone, methods capable of relating multiple factors are highly desirable. Existing methods for combined chromatin accessibility and DNA modification analysis rely on bisulfite sequencing, which has a number of well known drawbacks. We developed a novel cost-efficient method for simultaneous profiling of DNA methylation and hydroxymethylation within open chromatin loci. Our approach is based on chemo-enzymatic covalent tagging of unmodified (uCG) and hydroxymethylated (5hmC) CG sites along with GC sites in native chromatin, which are then mapped by tag-selective sequencing. Validation of the technology on model DNA systems and mammalian cells demonstrated a great applicability of this method for epigenomic studies. We employed the technology for tracking chromatin dynamics combined with changes of two different cytosine states, unmodified and hydroxymethylated, during differentiation of embryonic stem cells (ESC) which revealed extensive chromatin dynamics and DNA modification changes across gene bodies of development-associated genes at early transition from ESC. Strong DNA demethylation in the ESC-to-neuron differentiation pathway proceeds independently from cell transcriptional programs and highlights the value of separate 5hmC and 5mC analysis in understanding cell fate decisions.
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| Overall design |
We performed differentiation of mouse ESC (mESC) into a population of radial-glial neuronal progenitor cells (NPC) and later into terminally differentiated glutamatergic pyramidal neurons (NC). 1*105 nuclei extracted from three developmental stages cells were used for double uCG- and GC-, 5hmC- and GC-azide and triple CG-, GC-alkyne, and 5hmC-azide labeling reactions.
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| Contributor(s) |
Skardžiūtė K, Kvederavičiūtė K, Pečiulienė I, Narmontė M, Ličytė J, Klimašauskas S, Kriukienė E |
| Citation(s) |
38154461 |
| |
| Submission date |
May 08, 2023 |
| Last update date |
Apr 04, 2024 |
| Contact name |
Kotryna Kvederaviciute |
| E-mail(s) |
[email protected]
|
| Organization name |
Vilnius University
|
| Street address |
Universiteto str. 3
|
| City |
Vilnuis |
| ZIP/Postal code |
01513 |
| Country |
Lithuania |
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| Platforms (2) |
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| Samples (9)
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| This SubSeries is part of SuperSeries: |
| GSE231929 |
Unified analysis of DNA methylation, hydroxymethylation and chromatin accessibility through targeted DNA labeling |
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| Relations |
| BioProject |
PRJNA970253 |
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