The role of EZH1 in neuronal differentiation and neurological disorders

PROJECT SUMMARY
Intellectual disabilities (ID) are lifelong conditions caused by neurodevelopmental errors. We recently identified
gain and loss of function (GOF and LOF) mutations in the chromatin modifier Enhancer of Zeste Homologue 1
(EZH1) as the cause a previously undiagnosed intellectual disability syndrome in ten children. EZH1 is one of
the two Histone H3 Lysine 27 (H3K27) methyltransferases of the Polycomb Repressive Complex 2 (PRC2). The
other one, EZH2, has long been considered the main responsible for H3K27 di and trimethylation (H3K27me2/3)
and PRC2 mediated transcriptional repression, in part owing to a weaker catalytic activity of EZH1. EZH2 is
highly expressed in dividing cells, and its dysfunction leads to defects in neural progenitor proliferation and fate
specification, and neurodevelopmental disease. Despite evidence indicating that EZH1 is also expressed in the
developing and adult nervous system, its relevance and function in neural development and homeostasis remain
unknown. In preliminary work, we found that EZH1 is expressed constantly across human cerebral cortex
development and becomes the predominant paralogue by the late neurogenesis period owing to a rapid decline
of EZH2 expression. Using human embryonic stem cells (hESC) carrying EZH1 LOF and GOF mutations, and
their differentiation to cortical neurons in monolayer and organoid cultures, we found signs of delayed neuronal
differentiation in EZH1 LOF and premature differentiation in EZH1 GOF. However, amounts of H3K27me3
measured by WB, showed similar levels across all the mutant and control cell lines. Thus, we hypothesize that
EZH1 regulates cortical neurogenesis timing through a non EZH2 redundant mechanism that becomes dominant
as neurogenesis progresses and EZH2 expression declines. To test this hypothesis in Aim 1 we will determine
differentiation stage specific molecular functions of EZH1 during neurogenesis by defining the genomic
binding profile of EZH1 (SubAim1.1), its effects on H3K27 methylation and transcriptional regulation
(SubAim1.2), and the composition of PRC2-EZH1 subcomplexes (SubAim1.3) over time during neuronal
differentiation. In Aim 2 we will dissect the effect of EZH1 mutations in cortical neurogenesis timing and
the ability of EZH1/2 inhibitors to them we will dissect the origin and consequences of EZH1 mutation driven
dysregulated neurogenesis timing by extending our organoid analysis to 30, 40, 60 and 100 days (SubAim2.1),
by unbiased quantification of the composition and cell type specific differential gene expression using scRNAseq
of the organoids (SubAim2.2), and assessing the potential of EZH1/2 inhibitors on restoring the cellular and
molecular alterations caused by EZH1LOF and GOF mutations (SubAim2.3). These studies will uncover a
currently disregarded role of EZH1 in the regulation of cortical neurogenesis and neurodevelopmental diseases
and may provide new therapeutic targets for IDs.

Grant Number: 5R01NS119699-04
NIH Institute/Center: NIH
Principal Investigator: Naiara Akizu