Imprinted expression of UBE3A is neuron-specific and occurs because the paternally-inherited allele is silenced. A long non-coding antisense transcript, UBE3A-ATS, is expressed in a neuron-specific manner and mediates UBE3A imprinting by an unknown mechanism. The overall goal of this proposal is to understand the processes underlying the regulation of neuron-specific expression of UBE3A-ATS and repression of paternal UBE3A in human neurons. We propose to investigate three different aspects regulating UBE3A imprinted expression: 1) the regulation of coding versus non-coding SNRPN RNAs in human neurons; 2) the tissue-specific regulation of UBE3A-ATS, the distal portion of the SNRPN ncRNA; and 3) the mechanism by which UBE3A-ATS leads to the repression of paternal UBE3A. We will use patient-specific induced pluripotent stem cells (iPSCs) derived from Angelman syndrome (AS) patients and their neuronal derivatives for these studies. We will dissect the functional elements of the cis-acting boundary element restricting UBE3A-ATS expression to neurons and determine the developmental timing of their removal. We will determine how SNRPN coding versus non-coding RNA is produced and manipulate expression levels of UBE3A-ATS and UBE3A to determine their effect on UBE3A imprinting. Finally, we will test the hypothesis that UBE3A-ATS silences paternal UBE3A via a transcriptional interference mechanism. A thorough understanding of the mechanisms underlying UBE3A imprinted expression may reveal novel gene regulatory mechanisms applicable elsewhere in the genome, and may identify novel therapeutic targets for treating AS.
These experiments propose to use human Angelman syndrome (AS) induced pluripotent stem cells to understand how imprinted expression of UBE3A is regulated in human neurons. This study benefits public health because it will 1) increase our understanding of how neurons express only one copy of UBE3A, 2) characterize a poorly-understood long non-coding RNA targeted by AS therapies currently under investigation, and 3) potentially identify new therapeutic targets to treat individuals with AS.
