Epigenetic Maintenance of Neural Cell Identity

Neurodevelopmental disorders (NDDs) comprise a group of genetically and phenotypically heterogeneous
pathologies commonly characterized by psychiatric impairment. The molecular basis of these
neuropathologies remains poorly understood. Recent whole-genome-sequencing studies revealed that
mutations in genes encoding heterochromatin modifiers are significantly associated with NDDs. This class of
transcriptional regulators is thought to stabilize neural cell identity and function by enforcing heritable silencing
of lineage non-specific genes through epigenetic chromatin modifications. However, since most heterochromatin
modifiers are ubiquitously expressed and lack sequence-specificity, (1) how precise targeting of repressive
chromatin is controlled and (2) how mutations in general heterochromatin modifiers contribute to NDD-
associated neuronal defects remains unclear. To gain experimental traction on these questions, we will examine
the mechanism by which a high-confidence NDD risk gene, ZNF462, recruits the heterochromatin
modifiers EHMT1/2. We will test whether and how ZNF462 restricts lineage non-specific gene expression
and maintains neural cell identity. ZNF462 haploinsufficiency causes Weiss-Kruszka syndrome, a complex
NDD characterized by neurodevelopmental defects including developmental delay and autism. However, the
neurodevelopmental role of the C2H2 zinc finger protein is unknown. We previously discovered that mouse
Zfp462, is required for endodermal gene repression, directing Ehmt1/2-dependent heterochromatin to
transposable element (TE)-derived enhancers in neural progenitor cells. We hypothesize that human ZNF462
controls facultative heterochromatin formation, by specifically restricting non-neural gene expression during
neurogenesis. However, we predict that due to rapid species-specific evolution of TEs, ZNF462 will have
novel human targets and control a distinct gene regulatory network. W e will therefore: (Aim 1) employ neural
differentiation of human embryonic stem cells (hESCs) coupled to epigenome and transcriptome profiling to
investigate the impact of ZNF462 heterozygosity on maintenance of neural gene expression, (Aim 2) perform
structure-function analysis and functional complementation in mESCs to identify ZNF462 protein domains
responsible for homodimerization, DNA binding and transcriptional repression and (Aim 3) profile CTCF
binding and three-dimensional chromosome conformation in neuroepithelial stem cells (NESCs) to investigate
the impact of ZNF462 heterozygosity on neuro-specific genome architecture. Our proposal provides a path to
novel insight into the molecular mechanism of ZNF462-dependent gene silencing, and enhance our
understanding of the etiology of Weiss-Kruszka syndrome. The following strategy will reveal new concepts in
gene regulation and neurobiology and elucidate the link between mutations in heterochromatin modifiers and
NDDs. Overall, our work will inform novel strategies to prevent and treat NDDs arising from epigenetic
dysregulation.

Grant Number: 5R01MH122565-05
NIH Institute/Center: NIH
Principal Investigator: Oliver Bell