Differential responses of clinically important gene classes to transient loss of DNA methylation in human differentiated cells.

Sarah-Jayne Mackin, Karla O'Neill, Colum P Walsh

Research output: Chapter in Book/Report/Conference proceedingConference contributionpeer-review

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Background: Methylation of DNA sequences at promoters, CpGislands and other elements plays a vital role in regulating gene activity. In human, loss of methylation is known to play a causativ erole in imprinting disorders and in inappropriate germline gene expression in cancers. While in mouse, loss of function mutants have given great insight into the targets of methylation, functional studies in human have been largely limited to cancer cells and more recently stem cells, not normal adult cells. Methods: Stable knockdown sof the maintenance methyltransferase DNMT1 were generated innormosomic hTERT-immortalised adult fibroblasts. Genome-wide methylation levels were assayed using the Illumina 450K bead array. Results were analysed using RnBeads and Galaxy. Locus specific methylation was verified using pyrosequencing and clonal analysis. Validation was achieved using transient siRNA. Results:Loss of function was poorly tolerated and all clonally-expanded cell lines had spontaneously restored DNMT1 levels by silencing of the shRNA. Evidence for a genome-wide methylation erasure event followed by a wave of remethylation could be clearly traced.Gene bodies and the shores of CpG islands showed the clearest loss of methylation overall. While most CpG islands are normally unmethylated and so unaffected, both imprints and germ line genes fall into the rarer category of normally methylated islands: ofthese two, lasting loss of methylation was much more common among imprints than germ line genes. Conclusions: 1: transient loss of methylation is poorly tolerated; 2: a robust mechanism for remethylation exists even in adult cells; 3: aberrant remethylationis frequent on recovery and 4: Imprints are particularly sensitive.
Original languageEnglish
Title of host publicationUnknown Host Publication
PublisherUlster Medical Journal
Number of pages1
Publication statusAccepted/In press - 3 Sept 2015
Event18th Meeting of the Irish Society of Human Genetics - Dublin City University
Duration: 3 Sept 2015 → …


Conference18th Meeting of the Irish Society of Human Genetics
Period3/09/15 → …


  • Methylation
  • DNMT1
  • 450k
  • Imprints.


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