Imprinting regulation of the murine Meg1/Grb10 and human GRB10 genes; roles of brain-specific promoters and mouse-specific CTCF-binding sites

Abstract

The imprinted mouse gene Meg1/Grb10 is expres sed from maternal alleles in almost all tissues and organs, except in the brain, where it is expressed biallelically, and the paternal allele is expressed preferentially in adulthood. In contrast, the human GRB10 gene shows equal biallelic expression in almost all tissues and organs, while it is almost always expressed paternally in the fetal brain. To elucidate the molecular mechanisms of the complex imprinting patterns among the different tissues and organs of humans and mice, we analyzed in detail both the genomic structures and tissue-specific expression profiles of these species. Experiments using 5'-RACE and RT-PCR demonstrated the existence in both humans and mice of novel brain- specific promoters, in which only the paternal allele was active. The promoters were located in the primary differentially methylated regions. Interest ingly, CTCF-binding sites were found only in the mouse promoter region where CTCF showed DNA methylation-sensitive binding activity. Thus, the insulator function of CTCF might cause reciprocal maternal expression of the Meg1/Grb10 gene from another upstream promoter in the mouse, whereas the human upstream promoter is active in both parental alleles due to the lack of the corresponding insulator sequence in this region.

Tissue-specific regulation and function of Grb10 during growth and neuronal commitment

“…Subsequent experiments in mice demonstrated that Grb10 is transcribed in neurons from a series of downstream alternative promoters, exclusively from the paternal allele (8,47). This paternal expression is conserved in the human brain (48).…”

“…1). The region surrounding one of these alternative promoters has been identified as an ICR, which exhibits DNA methylation only on the maternal allele in all examined tissues (8). On the paternal allele, the unmethylated Grb10 ICR is bound by CCCTC-binding factor (CTCF), a multifunctional transcription factor, which is recruited in a DNA methylation-sensitive manner and has been implicated in the regulation of imprinted expression at other loci (Fig.…”

Genome-wide and parental allele-specific analysis of CTCF and cohesin DNA binding in mouse brain reveals a tissue-specific binding pattern and an association with imprinted differentially methylated regions

“…CTCF has long been associated with genomic imprinting due to its selective binding of the unmethylated maternal allele of the Igf2/H19 ICR resulting in parent-of-origin-specific expression of Igf2 and H19 in mouse and human (Bell and Felsenfeld 2000;Hark et al 2000;Kanduri et al 2000;Fedoriw et al 2004;Szabo et al 2004). CTCF has been studied at several other imprinted loci, and it binds the unmethylated allele at the gDMRs of Rasgrf1, Peg13, Kcnq1ot1 (Yoon et al 2005;Fitzpatrick et al 2007;Singh et al 2011), and Grb10 (Hikichi et al 2003;Mukhopadhyay et al 2004). CTCF-mediated regulation is postulated to be one of two major control mechanisms operating at ICRs (Lewis and Reik 2006;Kim et al 2009).…”

Genomic Imprinting and Epigenetic Control of Development

“…The differential use of alternative promoters appears to be a common theme. For example, the Grb10 gene is maternally expressed in most tissues, yet predominantly paternal in the brain (Arnaud et al 2003;Hikichi et al 2003). This is accomplished through the use of a hypomethylated paternal DMR that is capable of serving as a promoter in this state (Arnaud et al 2003;Hikichi et al 2003;Yamasaki-Ishizaki et al 2007;Sanz et al 2008).…”