Epigenetic regulation of mammalian imprinted genes: from primary to functional imprints.

Parental genomic imprinting was discovered in mammals some 20 years ago. This phenomenon, crucial for normal development, rapidly became a key to understanding epigenetic regulation of mammalian gene expression. In this chapter we present a general overview of the field and describe in detail the 'imprinting cycle'.

Genomic imprinting controls matrix attachment regions in the Igf2 gene.

Genomic imprinting at the Igf2/H19 locus originates from allele-specific DNA methylation, which modifies the affinity of some proteins for their target sequences. Here, we show that AT-rich DNA sequences located in the vicinity of previously characterized differentially methylated regions (DMRs) of the imprinted Igf2 gene are conserved between mouse and human. These sequences have all the characteristics of matrix attachment regions (MARs), which are known as versatile regulatory elements involved in chromatin structure and gene expression.

A real-time polymerase chain reaction assay for quantification of allele ratios and correction of amplification bias.

Allele-specific epigenetic modifications are crucial for several important biological functions, including genomic imprinting and X-inactivation in mammals. Consequently, an ever increasing number of investigations requires accurate quantification of the relative abundance of parental alleles of a specific sequence in a DNA sample. Here, combining the use of polymorphic restriction sites with real-time polymerase chain reaction (PCR) amplification, we describe a simple and quantitative assay to measure allele ratios.

Turnover of primary transcripts is a major step in the regulation of mouse H19 gene expression.

In the gene expression pathway, RNA biogenesis is a central multi-step process where both message fidelity and steady-state levels of the mature RNA have to be ascertained. An emerging question is whether RNA levels could be regulated at the precursor stage. Until recently, because it was technically very difficult to determine the level of a pre-mRNA, discrimination between changes in transcriptional activity and in pre-mRNA metabolism was extremely difficult.