Studying the genetic and epigenetic alterations in Imprinting Disorders

Imprinting disorders (ImpDis) are a group of congenital diseases caused by dysregulation of genomic imprinting and affecting prenatal and postnatal growth, neurocognitive development, and metabolism. Silver-Russell syndrome (SRS) and Beckwith-Wiedemann syndrome (BWS) represent two clinically opposite imprinting disorders caused primarily by the dysregulation of the imprinted gene cluster on chr11p15.5. Some patients with ImpDis show multi-locus imprinting disturbances (MLID), characterised by methylation abnormalities affecting multiple imprinted regions, although the underlying mechanisms and health impact of this epigenetic disturbance remain unclear. In my PhD thesis, I investigated the genetic and epigenetic alterations in imprinting disorders, with particular focus on SRS and BWS. Through integrated multi-omics approaches, this work provides a comprehensive overview of imprinting disorders and reveals their molecular complexity and heterogeneity. The thesis comprises the following four main studies: ❖ Chapter 3 identifies a novel link between PLAG1 damaging variants and methylation changes at the MEG8 locus in SRS patients, showing that PLAG1 can influence methylation maintenance at the chr14q32 imprinted region. These findings expand the classical PLAG1-HMGA2-IGF2 pathway and highlight crosstalk between imprinted and non-imprinted genes. ❖ Chapter 4 presents the molecular characterisation of an Italian cohort of SRS patients, revealing the heterogeneity of Silver-Russell syndrome. The integration of genomic and methylation analyses uncovered both pathogenic variants and methylation changes affecting genes involved in interconnected regulatory pathways that govern growth and morphogenesis. ❖ Chapter 5 describes a rare case of adult-onset β-thalassemia caused by a nonsense HBB mutation inherited in heterozygosity from his father and becoming homozygous in the bone marrow because of somatic and segmental paternal uniparental isodisomy of chr11p15. This study provides direct evidence that somatic imprinting dysregulation can cooperate with inherited recessive mutations to drive an acquired, tissue-specific disease. ❖ Chapter 6 focuses on BWS patients and MLID. It demonstrates that MLID patients exhibit highly variable methylation changes at imprinted and non-imprinted loci, seemingly in a stochastic manner across the genome. These findings support a model in which maternaleffect mutations and oocyte ageing compromise methylation maintenance during early embryogenesis, contributing to imprinting instability. Overall, this thesis provides a comprehensive overview of two imprinting disorders, demonstrating that their molecular bases involve converging genetic, epigenetic and environmental mechanisms. These studies reveal how dysregulation of imprinted networks contributes to both congenital and acquired phenotypes, redefining imprinting disorders as dynamic and complex conditions.