Tissue-Specific and Minor Inter-Individual Variation in Imprinting of IGF2R Is a Common Feature of Bos taurus Concepti and Not Correlated with Fetal Weight

Daniela Bebbere,Stefan Hiendleder,Eckhard Wolf,Horst-Dieter Reichenbach,Susanne E Ulbrich,Rainer W Fürst,Myriam Reichenbach,Sergio Ledda,Stefan Bauersachs,Ivica Medugorac,Robert Feil

doi:10.1371/journal.pone.0059564

Abstract

The insulin-like growth factor 2 receptor (IGF2R) is essential for prenatal growth regulation and shows gene dosage effects on fetal weight that can be affected by in-vitro embryo culture. Imprinted maternal expression of murine Igf2r is well documented for all fetal tissues excluding brain, but polymorphic imprinting and biallelic expression were reported for IGF2R in human. These differences have been attributed to evolutionary changes correlated with specific reproductive strategies. However, data from species suitable for testing this hypothesis are lacking. The domestic cow (Bos taurus) carries a single conceptus with a similar gestation length as human. We identified 12 heterozygous concepti informative for imprinting studies among 68 Bos taurus fetuses at Day 80 of gestation (28% term) and found predominantly maternal IGF2R expression in all fetal tissues but brain, which escapes imprinting. Inter-individual variation in allelic expression bias, i.e. expression of the repressed paternal allele relative to the maternal allele, ranged from 4.6−8.9% in heart, 4.3−10.2% in kidney, 6.1−11.2% in liver, 4.6−15.8% in lung and 3.2−12.2% in skeletal muscle. Allelic bias for mesodermal tissues (heart, skeletal muscle) differed significantly (P<0.05) from endodermal tissues (liver, lung). The placenta showed partial imprinting with allelic bias of 22.9−34.7% and differed significantly (P<0.001) from all other tissues. Four informative fetuses were generated by in-vitro fertilization (IVF) with embryo culture and two individuals displayed fetal overgrowth. However, there was no evidence for changes in imprinting or DNA methylation after IVF, or correlations between allelic bias and fetal weight. In conclusion, imprinting of Bos taurus IGF2R is similar to mouse except in placenta, which could indicate an effect of reproductive strategy. Common minor inter-individual variation in allelic bias and absence of imprinting abnormalities in IVF fetuses suggest changes in IGF2R expression in overgrown fetuses could be modulated through other mechanisms than changes in imprinting.

Highlights

The multifunctional mannose 6 phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R), hereafter referred to as IGF2R, mediates endocytosis and subsequent clearance or activation of a variety of ligands involved in the regulation of cell growth and motility, including insulin-like growth factor 2 and transforming growth factor b [1,2]
Indirect evidence for such an effect was obtained in the sheep model where fetal overgrowth induced by embryo culture was associated with hypomethylation at a CpG site in an intronic sequence element implicated in IGF2R imprinting and down regulated IGF2R expression [22]
Contrary to expectations, imprinting and DNA methylation were not affected by in-vitro fertilization (IVF) with in-vitro embryo culture and there was no correlation between minor variation in inter-individual allele-specific expression bias and fetal weight

Summary

Introduction

The multifunctional mannose 6 phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R), hereafter referred to as IGF2R, mediates endocytosis and subsequent clearance or activation of a variety of ligands involved in the regulation of cell growth and motility, including insulin-like growth factor 2 and transforming growth factor b [1,2].The IGF2R gene shows developmental stage specific expression levels which are highest in the fetus and decline rapidly after birth [3]. These features make the cow an excellent model for comparisons with human and mouse, but current Bos taurus IGF2R imprinting data is limited to two fetal liver samples of unspecified age [12]. We determined the tissue-specific imprinting status of IGF2R in first trimester Bos taurus concepti generated in-vivo or in-vitro.

Results

Conclusion