A novel in vitro system for analyzing parental allele-specific histone acetylation in genomic imprinting.
One of the obstacles in studying human genomic imprinting is distinguishing the parental origin of alleles in diploid cells. To solve this problem, we have constructed a library of mouse A9 hybrids in which individual clones contain a single human chromosome of known parental origin. Here we extend this in vitro system to the analysis of the role of histone acetylation in the allelic expression of human imprinted genes. The levels of histone H4 acetylation of the imprinted human LIT1, H19, and SNRPN genes were examined by a chromatin immunoprecipitation (ChIP) assay in mouse A9 hybrids with a single human chromosome of known parental origin. We demonstrated that H4 histones associated with the actively expressed alleles of imprinted LIT1, H19, and SNRPN genes were highly acetylated, whereas they were hypoacetylated in the silent alleles. Furthermore, treatment of A9 hybrids with trichostatin A (TSA), an inhibitor of histone deacetylase, resulted in transcriptional reactivation of the silent alleles for LIT1 and SNRPN, suggesting that histone deacetylation is one of the key regulatory mechanisms in genomic imprinting. These results indicate that our monochromosomal hybrid system is a new technology for analyzing histone modifications between parental alleles in human imprinted genes.
Yoshioka
H
H
Department of Molecular and Cell Genetics, School of Life Sciences, Faculty of Medicine, Tottori University, Yonago, Japan.
Shirayoshi
Y
Y
Oshimura
M
M
D000123:Acetyltransferases / Q000235:genetics; D000818:Animals; D002460:Cell Line; D002843:Chromatin / Q000235:genetics; D002874:Chromosome Mapping; D002880:Chromosomes, Human, Pair 11; D002884:Chromosomes, Human, Pair 15; D017931:DNA Primers; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018392:Genomic Imprinting; D051548:Histone Acetyltransferases; D056572:Histone Deacetylase Inhibitors; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D051379:Mice; D016133:Polymerase Chain Reaction; D015183:Restriction Mapping; D020133:Reverse Transcriptase Polymerase Chain Reaction; D029701:Saccharomyces cerevisiae Proteins
D002843:Chromatin; D017931:DNA Primers; D004791:Enzyme Inhibitors; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D029701:Saccharomyces cerevisiae Proteins; C012589:trichostatin A; D000123:Acetyltransferases; D051548:Histone Acetyltransferases