Epigenetic reprogramming of the human H19 gene in mouse embryonic cells does not erase the primary parental imprint.
BACKGROUND
Genomic imprinting in mammals is thought to result from epigenetic modifications to chromosomes during gametogenesis, which leads to differential allelic expression during development. There is a requirement for an appropriate experimental system to enable the analysis of the mechanisms of genomic imprinting during embryogenesis.


RESULTS
To develop a novel in vitro system for studying the molecular basis of genomic imprinting, we constructed mouse cell lines containing either a paternal or maternal human chromosome 11, by microcell-mediated chromosome transfer. Allele-specific expression and DNA methylation studies revealed that the imprinting status of the human H19 gene was maintained in mouse A9 mono-chromosomal hybrids. Each parental human chromosome was introduced independently into mouse near-diploid immortal fibroblasts (m5S) and two embryonal carcinoma (EC) cell lines (OTF9-63 and P19). The paternal allele of human H19 remained in a repressed state in m5S cells, but was de-repressed in both EC cells. The paternal H19 allele was demethylated extensively in OTF9-63 cells, whereas the only alteration in P19 hybrids was de novo methylation on both alleles in the 3' region. Following in vitro differentiation, the expressed paternal H19 allele was selectively repressed in differentiated derivatives of EC hybrids.


CONCLUSIONS
These results indicated that human imprint marks could function effectively in mouse cells, and that the imprinting process was epigenetically reprogrammed in embryonal carcinoma cells, without erasure of the primary imprint that marked the parental origin. Therefore, these mono-chromosomal hybrids could provide a valuable in vitro system to study the mechanisms involved in the regulation of imprinted gene expression.
Mitsuya
K
K
Department of Molecular and Cell Genetics, School of Life Sciences, Faculty of Medicine, Tottori University, Yonago, Japan.
Meguro
M
M
Sui
H
H
Schulz
T C
TC
Kugoh
H
H
Hamada
H
H
Oshimura
M
M
D000818:Animals; D002454:Cell Differentiation; D002459:Cell Fusion; D002880:Chromosomes, Human, Pair 11 / Q000235:genetics; D019175:DNA Methylation; D005260:Female; D005786:Gene Expression Regulation; D018392:Genomic Imprinting; D006801:Humans; D006822:Hybrid Cells; D008297:Male; D051379:Mice; D009124:Muscle Proteins / Q000235:genetics*; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated; D013045:Species Specificity
C094335:H19 long non-coding RNA; D009124:Muscle Proteins; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated