Regulation of the large ( 1000 kb) imprinted murine Ube3a antisense transcript by alternative exons upstream of Snurf/Snrpn

Abstract

Most cases of Angelman syndrome (AS) result from loss or inactivation of ubiquitin protein ligase 3A (UBE3A), a gene displaying maternal-specific expression in brain. Epigenetic silencing of the paternal UBE3A allele in brain appears to be mediated by a non-coding UBE3A antisense (UBE3A-ATS). In human, UBE3A-ATS extends approximately 450 kb to UBE3A from the small nuclear ribonucleoprotein N (SNURF/SNRPN) promoter region that contains a cis-acting imprinting center (IC). The concept of a single large antisense transcript is difficult to reconcile with the observation that SNURF/SNRPN shows a ubiquitous pattern of expression while the more distal part of UBE3A-ATS, which overlaps UBE3A, is brain specific. To address this problem, we examined murine transcripts initiating from several alternative exons dispersed within a 500 kb region upstream of Snurf/Snrpn. Similar to Ube3a-ATS, these upstream (U) exon-containing transcripts are expressed at neuronal stages of differentiation in a cell culture model of neurogenesis. These findings suggest the novel hypothesis that brain-specific transcription of Ube3a-ATS is regulated by the U exons rather than Snurf/Snrpn exon 1 as previously suggested from human studies. In support of this hypothesis, we describe U-Ube3a-ATS transcripts where U exons are spliced to Ube3a-ATS with the exclusion of Snurf-Snrpn. We also show that the murine U exons have arisen by genomic duplication of segments that include elements of the IC, suggesting that the brain specific silencing of Ube3a is due to multiple alternatively spliced IC-Ube3a-ATS transcripts.

Angelman syndrome imprinting center encodes a transcriptional promoter

“…Instead, the mouse has nine highly repeated upstream promoters located over 530 kb (31). Although all nine repeats are unlikely to be active promoters, it is clear that some are used in both oocytes and somatic cells (10,(18)(19)(20)32). Removal of the three proximal promoters leads to a weak partial imprinting defect, consistent with functional complementation by the remaining upstream promoters (33).…”

“…Extrapolating from these paternally derived transcripts detected in brain tissue, they speculated the existence of similar transcripts in oocytes that would transit the PWS-SRO and thereby contribute to the imprint-setting process. Although lacking sequence homology, the mouse Snrpn locus also contains several upstream promoter exons that are alternatives to body exon 1 (10,18). These transcripts become detectable in mouse oocytes just before the application of DNA methylation at the PWS-IC (19,20).…”

Subcellular organization of UBE3A in neurons

“…Our results show that some neuronal UBE3A labeling remains in AS mice, perhaps reflecting the unusual imprinting mechanism (Landers et al, 2004;Le Meur et al, 2005). Rather than being uniformly distributed across the different neuronal compartments, UBE3A in AS mice concentrated in neuropil, especially in axon terminals, and was almost entirely absent from nuclei.…”

Narrowed abrogation of the Angelman syndrome critical interval on human chromosome 15 does not interfere with epigenotype maintenance in somatic cells

“…It was recently suggested, based on a cell culture model of neurogenesis, that Ube3a antisense transcripts contain upstream exons and that multiple alternatively spliced Ube3a antisense transcripts regulate brain-specific silencing of Ube3a (Rougeulle et al 1998;Chamberlain and Brannan 2001;Yamasaki et al 2003;Landers et al 2004). This possibility can be investigated for human chromosomes 15 using the DT40 cell shuttle system and a cell culture model of neurogenesis.…”