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2019

ANUÁRIO DO HOSPITAL
DONA ESTEFÂNIA

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LOSS OF HIERARCHICAL IMPRINTING REGULATION AT THE PRADER-WILLI-ANGELMAN SYNDROME LOCUS IN HUMAN iPSCs

Duarte Pólvora-Brandão1, Mariana Joaquim1, Inês Godinho1, Domenico Aprile1, Ana Rita Álvaro2,3 Isabel Onofre2, Ana Cláudia Raposo1, Luís Pereira de Almeida2,4, Sofia T Duarte1,5 and Simão T da Rocha1

1 - Instituto de Medicina Molecular João Lobo Antunes, Universidade de Lisboa, Lisboa, Portugal
2 - CNC - Center for Neurosciences and Cell Biology, University of Coimbra, Coimbra, Portugal
3 - Institute for Interdisciplinary Research (IIIUC), University of Coimbra, Coimbra, Portugal
4 - Faculty of Pharmacy, University of Coimbra,Coimbra, Portugal
5 - Hospital D. Estefânia, Centro Hospitalar Lisboa Central, Lisboa, Portugal

- Publicação em versão integral na revista Human Molecular Genetics
- Hum Mol Genet. 2018 Dec 1; 27(23): 3999–4011

The human chr15q11-q13 imprinted cluster is linked to several disorders, including Prader-Willi (PWS) and Angelman (AS) syndromes. Recently, disease modelling approaches based on induced pluripotent stem cells (iPSCs) have been used to study these syndromes. A concern regarding the use of these cells for imprinted disease modelling is the numerous imprinting defects found in many iPSCs. Here, by reprogramming skin fibroblasts from a control and an AS individuals, we generated several iPSC lines and addressed the stability of imprinting status across the PWS/AS domain. We focused on three important regulatory DNA elements which are all differentially methylated regions (DMRs), methylated on the maternal allele: the PWS imprinting center (PWS-IC), which is a germline DMR and the somatic NDN and MKRN3 DMRs, hierarchically controlled by PWS-IC. Normal PWS-IC methylation pattern was maintained in most iPSC lines, however, loss of maternal methylation in one out of five control iPSC lines resulted in a monoallelic to biallelic switch for many imprinted genes in this domain. Surprisingly, MKRN3 DMR was found aberrantly hypermethylated in all control and AS iPSCs, regardless of the methylation status of PWS-IC master-regulator. This suggests a loss of hierarchical control of imprinting at PWS/AS region. We confirmed these results in established iPSC lines derived using different reprogramming procedures. Overall, we show that hierarchy of imprinting control in somatic cells might not apply to iPSCs, accounting for their spectrum of the imprinting alterations. Such changes in imprinting regulation should be taken into consideration for the use of iPSCs in disease modelling.

Palavras Chave: Prader-Willi/Angelman syndrome; Imprinting; Human induced pluripotent stem cells