A developmental perspective on ZNF804A gene function: Understanding the neurobiological role of schizophrenia susceptibility gene ZNF804A in hiPSC-derived developing glutamatergic neurons

Student thesis: Doctoral ThesisDoctor of Philosophy

Abstract

Schizophrenia is a severe psychiatric disorder caused by a complex interaction of environmental risk factors and genetic susceptibility, yet its underlying cellular and molecular dysfunction remains poorly understood. Hence, it is crucial to uncover the neurobiological function of robust risk genes, such as ZNF804A, which has shown consistent associations with the disorder. While its function was linked to various cellular processes, including transcription factor activity, protein synthesis regulation, and synapse maintenance, its role in neurodevelopment remains unclear. The aim of this thesis was therefore, to analyse ZNF804A function in a developmental and cell type relevant model system, while hypothesising its involvement in forming glutamatergic synapses and mediating local translation in developing glutamatergic cells. To identify a suitable cellular system, ZNF804A transcript expression was tested using RT-qPCR in human induced pluripotent stem cell (hiPSC)-derived cortical neuroprogenitor cells (NPCs) and developing glutamatergic neurons at different stages of differentiation. The findings demonstrated gradually increasing expression throughout development, with highest levels in post-mitotic glutamatergic neurons. Consequently, a forward programming protocol, combined with SMAD inhibition, was validated to generate developing glutamatergic neurons to further characterise gene function. This cellular system was subjected to a CRISPR/Cas9-based precision functional genetics approach to create hiPSC lines with homozygotic and heterozygotic ZNF804A mutation. Homozygotic mutant neurons exhibited reduced ZNF804A protein expression and bulk RNA sequencing revealed differential gene expression, with an enrichment in genes involved in cell adhesion and synaptic functioning. Next, synaptic and translational processes were assessed in ZNF804A mutant lines. High-content confocal imaging showed an increase of pre- and post-synaptic proteins within ZNF804A-deficient neurites and enhanced putative synapse density. Moreover, up-regulated MAPK/ERK signalling and local protein synthesis as assessed by SUnSET coincided with an increase of key components of the translational machinery within neurites. Overall, this research provides insights into cellular and molecular mechanisms of ZNF804A during neurodevelopment, indicating a role for ZNF804A in local protein synthesis and synapse formation, thereby establishing a potential link between its known cellular functions. By unravelling ZNF804A’s neurodevelopmental function, this study contributes to our understanding of schizophrenia's aetiology and may aid to identify future therapeutic targets.
Date of Award1 Sept 2023
Original languageEnglish
Awarding Institution
  • King's College London
SupervisorDeepak Srivastava (Supervisor) & Anthony Vernon (Supervisor)

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