| RRC ID |
28643
|
| Author |
Kobayakawa S, Miike K, Nakao M, Abe K.
|
| Title |
Dynamic changes in the epigenomic state and nuclear organization of differentiating mouse embryonic stem cells.
|
| Journal |
Genes Cells
|
| Abstract |
Changes in nuclear organization and the epigenetic state of the genome are important driving forces for developmental gene expression. However, a strategy that allows simultaneous visualization of the dynamics of the epigenomic state and nuclear structure has been lacking to date. We established an experimental system to observe global DNA methylation in living mouse embryonic stem (ES) cells. The methylated DNA binding domain (MBD) and the nuclear localization signal (nls) sequence coding for human methyl CpG-binding domain protein 1 (MBD1) were fused to the enhanced green fluorescent protein (EGFP) reporter gene, and ES cell lines carrying the construct (EGFP-MBD-nls) were established. The EGFP-MBD-nls protein was used to follow DNA methylation in situ under physiological conditions. We also monitored the formation and rearrangement of methylated heterochromatin using EGFP-MBD-nls. Pluripotent mouse ES cells showed unique nuclear organization in that methylated centromeric heterochromatin coalesced to form large clusters around the nucleoli. Upon differentiation, the organization of these heterochromatin clusters changed dramatically. Time-lapse microscopy successfully captured a moment of dramatic change in chromosome positioning during the transition between two differentiation stages. Thus, this experimental system should facilitate studies focusing on relationships between nuclear organization, epigenetic status and cell differentiation.
|
| Volume |
12(4)
|
| Pages |
447-60
|
| Published |
2007-4-1
|
| DOI |
10.1111/j.1365-2443.2007.01063.x
|
| PII |
GTC1063
|
| PMID |
17397393
|
| MeSH |
Animals
Cell Differentiation / physiology*
Cell Nucleus* / metabolism
Cell Nucleus* / ultrastructure
Cells, Cultured
Chromosomes / metabolism
Chromosomes / ultrastructure
DNA (Cytosine-5-)-Methyltransferase 1
DNA (Cytosine-5-)-Methyltransferases / genetics
DNA (Cytosine-5-)-Methyltransferases / metabolism
DNA Methylation*
DNA-Binding Proteins / chemistry
DNA-Binding Proteins / genetics
DNA-Binding Proteins / metabolism
Embryonic Stem Cells* / cytology
Embryonic Stem Cells* / physiology
Epigenesis, Genetic*
Fluorescence Recovery After Photobleaching
Genes, Reporter
Humans
Mice
Mice, Knockout
Nuclear Localization Signals
Recombinant Fusion Proteins / chemistry
Recombinant Fusion Proteins / genetics
Recombinant Fusion Proteins / metabolism
Transcription Factors / chemistry
Transcription Factors / genetics
Transcription Factors / metabolism
|
| IF |
1.655
|
| Times Cited |
64
|
|
WOS Category
|
GENETICS & HEREDITY
CELL BIOLOGY
|
| Resource |
| DNA material |
pEGFP-MBD-nls (RDB08344) |
