UC Davis

Human cell line models, including LUHMES, are important for investigating developmental transcriptional dynamics within imprinted regions, particularly the 15q11-q13 Angelman (AS) & Prader-Willi (PWS) syndrome locus. AS is caused by a loss of maternal UBE3A, which is paternally silent in neurons. Silencing is mediated by the antisense UBE3A-ATS, which extends from SNRPN through UBE3A in neurons, as opposed to termination at PWAR1 in non-neurons.Quantitative (qPCR) analysis confirmed the exclusive & progressive increase in UBE3A-ATS in differentiating LUHMES neurons, validating their use for studying UBE3A silencing. Genome-wide transcriptome analyses revealed changes to 11,834 genes during neuronal differentiation, including the upregulation of genes within the 15q11-q13 locus. To identify dynamic changes to chromatin loops related to transcriptional changes we performed a HiChIP analysis validated by 4C which identified two neuron-specific CTCF chromatin loops between MAGEL2-SNRPN & PWAR1-UBE3A. To determine if allele-specific methylation patterns may be associated with CTCF loop anchors, whole genome long-read nanopore sequencing was performed. We identified a paternally hypomethylated DMR near the SNRPN loop anchor exclusive to neurons & a paternally hypermethylated DMR near the PWAR1 CTCF anchor exclusive to undifferentiated cells, suggesting its role in regulating the cell specific boundary that occurs there. Additionally, DMRs near CTCF binding sites, observed in both cell types, may also influence chromatin loop formation & gene regulation within this locus. This study provides an integrated view of the epigenetic landscape at the 15q11-q13 locus during LUHMES cell differentiation, underscoring the complex interplay of transcription, chromatin looping, & methylation, offering valuable insights for future therapeutic approaches for AS & PWS.