PATIENT-SPECIFIC MODELLING OF SYNDROMIC AUTISM: UNCOVERING THE ROLE OF ADNP IN CHROMATIN DYSREGULATION

ADNP encodes Activity-Dependent Neuroprotective Protein, whose de novo heterozygous mutations cause Helsmoortel-Van der Aa Syndrome (HVDAS), a rare developmental syndrome affecting brain formation and neuronal functions, involving autism spectrum disorder and intellectual disability. Although ADNP is one of the single-gene most frequently mutated in ASD, its precise role in the syndrome onset has yet to be clarified. ADNP is the DNA-binding component of the newly identified chromatin remodeler complex ChAHP in mESC. It recognizes euchromatin regions to establish less accessible local chromatin domains and has also been recently identified as a new player in the regulation of genomic topology, competing with CTCF in the organization of chromatin architecture. Our aim is to understand the genetic and epigenetic implications of ADNP underlying this neurodevelopmental condition; we harnessed cell reprogramming to establish a highly informative cohort of patient-specific iPSCs and use it as a platform to develop meaningful model for the pathology, thus enabling the assessment of the ADNP pivotal relevance in both pluripotent and neuronally-patterned stages. We discovered an altered gene expression program associated with cell fate decision and neuronal lineage commitment, highlighting a neurodevelopmental disruption elicited by ADNP mutations already at the pluripotent stage. Employing CRISPR/Cas9-engineering, we FLAG-tagged the endogenous ADNP to assess its genomic occupancy and revealed a genome-wide distribution of ADNP at gene-regulatory elements and a predominant presence at transposable elements, Alu sequences in particular. We decoupled ADNP and CTCF interplay in our human iPSCs model, and found a global redistribution of active enhancer histone marks signature, which sustain upregulation with the intervention of EZH2-mediated derepression. Finally, HVDAS cortical organoid models show morpho-functional impairment in the early stages of neuronal differentiation, with decreased size and lower mitotic activity, coupled with accelerated maturation phenotype assessed through single-cell transcriptomic analysis. Altogether, with these results we delineate how ADNP deficiency affects pluripotent regulatory landscape and disease-relevant mechanisms that ultimately impact neuronal development and functionality.