Evaluation of Allelic Expression of Imprinted Genes in Adult Human Blood

Jennifer M Frost,Gudrun E Moore,Lyn S Chitty,Kathryn Woodfine,Dave Monk,Taita Stojilkovic-Mikic,Adele Murrell,Philip Stanier

doi:10.1371/journal.pone.0013556

Abstract

BackgroundImprinted genes are expressed from only one allele in a parent-of-origin dependent manner. Loss of imprinted (LOI) expression can result in a variety of human disorders and is frequently reported in cancer. Biallelic expression of imprinted genes in adult blood has been suggested as a useful biomarker and is currently being investigated in colorectal cancer. In general, the expression profiles of imprinted genes are well characterised during human and mouse fetal development, but not in human adults.Methodology/Principal FindingsWe investigated quantitative expression of 36 imprinted genes in adult human peripheral blood leukocytes obtained from healthy individuals. Allelic expression was also investigated in B and T lymphocytes and myeloid cells. We found that 21 genes were essentially undetectable in adult blood. Only six genes were demonstrably monoallelic, and most importantly, we found that nine genes were either biallelic or showed variable expression in different individuals. Separated leukocyte populations showed the same expression patterns as whole blood. Differential methylation at each of the imprinting control loci analysed was maintained, including regions that contained biallelically expressed genes. This suggests in some cases methylation has become uncoupled from its role in regulating gene expression.Conclusions/SignificanceWe conclude that only a limited set of imprinted genes, including IGF2 and SNRPN, may be useful for LOI cancer biomarker studies. In addition, blood is not a good tissue to use for the discovery of new imprinted genes. Finally, lymphocyte DNA methylation status in the adult may not always be a reliable indicator of monoallelic gene expression.

Highlights

For the majority of expressed human genes there is both a maternal and a paternal contribution in the offspring
Tests based on analysis in adult blood samples of allelic expression of the imprinted IGF2 gene in the diagnosis of colorectal cancer have become the subject of commercial development [6]
Analysis of imprinted gene expression levels in adult Peripheral blood leukocytes (PBL) In order to study imprinting in adult blood, we first investigated the mRNA expression levels of 36 gene transcripts by quantitative (q)RT-PCR (Fig. 1), which was performed in two unrelated healthy adult individuals

Summary

Introduction

For the majority of expressed human genes there is both a maternal and a paternal contribution in the offspring. For other genes, imprinted expression is found in the majority of tissues during fetal development, as in the case of IGF2, but becomes biallelic in some tissues in the adult [15] Regulation of this temporal and tissue specificity is likely to be brought about through transcription factor binding to alternative promoters and enhancers. Imprinting expression patterns and related mechanisms in the human have largely been extrapolated from experiments in the mouse, it is clear that there are important differences between the two [11,17] To fill this knowledge gap, with the growing importance of LOI as a marker in cancer diagnostics, this study was designed to investigate genomic imprinting in adult human PBL, analysing 36 gene transcripts, including representatives from all of the known human imprinted cluster loci and several isolated imprinted genes. DNA methylation at ICRs was measured in order to verify DMR maintenance and potentially identify the source of any imprinting errors

Materials and Methods

Results

Discussion