RRC ID 58389
Author Matsuzaki H, Kuramochi D, Okamura E, Hirakawa K, Ushiki A, Tanimoto K.
Title Recapitulation of gametic DNA methylation and its post-fertilization maintenance with reassembled DNA elements at the mouse Igf2/H19 locus.
Journal Epigenetics Chromatin
Abstract BACKGROUND:Paternal allele-specific DNA methylation of the H19 imprinting control region (ICR) regulates imprinted expression of the Igf2/H19 genes. The molecular mechanism by which differential methylation of the H19 ICR is established during gametogenesis and maintained after fertilization, however, is not fully understood. We previously showed that a 2.9-kb H19 ICR fragment in transgenic mice was differentially methylated only after fertilization, demonstrating that two separable events, gametic and post-fertilization methylation, occur at the H19 ICR. We then determined that CTCF/Sox-Oct motifs and the 478-bp sequence of the H19 ICR are essential for maintaining its maternal hypomethylation status and for acquisition of paternal methylation, respectively, during the post-fertilization period.
RESULTS:Using a series of 5'-truncated H19 ICR transgenes to dissect the 478-bp sequence, we identified a 118-bp region required for post-fertilization methylation activity. Deletion of the sequence from the paternal endogenous H19 ICR caused loss of methylation after fertilization, indicating that methylation activity of the sequence is required to protect endogenous H19 ICR from genome-wide reprogramming. We then reconstructed a synthetic DNA fragment in which the CTCF binding sites, Sox-Oct motifs, as well as the 118-bp sequence, were inserted into lambda DNA, and used it to replace the endogenous H19 ICR. The fragment was methylated during spermatogenesis; moreover, its allele-specific methylation status was faithfully maintained after fertilization, and imprinted expression of the both Igf2 and H19 genes was recapitulated.
CONCLUSIONS:Our results identified a 118-bp region within the H19 ICR that is required for de novo DNA methylation of the paternally inherited H19 ICR during pre-implantation period. A lambda DNA-based artificial fragment that contains the 118-bp sequence, in addition to the previously identified cis elements, could fully replace the function of the H19 ICR in the mouse genome.
Volume 13(1)
Pages 2
Published 2020-1-14
DOI 10.1186/s13072-019-0326-1
PII 10.1186/s13072-019-0326-1
PMID 31937365
PMC PMC6958606
MeSH Animals CCCTC-Binding Factor / chemistry CCCTC-Binding Factor / genetics DNA Methylation* Female Fertilization Genomic Imprinting* Insulin-Like Growth Factor II / genetics* Male Mice Mice, Inbred C57BL RNA, Long Noncoding / genetics* RNA, Long Noncoding / metabolism Spermatogonia / metabolism*
IF 4.237
Times Cited 0
Mice RBRC00639