Holoprosencephaly (HPE) is a genetically heterogeneous disorder with multiple underlying causes and modes of inheritance. Approximately 50% of cases are attributable to chromosomal abnormalities, 25% occur as part of a defined syndrome, and the remaining 25% present as isolated, de novo (non-syndromic) HPE. Although HPE is typically classified as autosomal dominant, inheritance is non-Mendelian with incomplete penetrance, as only ~70% of individuals carrying pathogenic variants in HPE-associated genes manifest clinical features.
To date, fourteen genes, including ZIC2have been implicated in non-syndromic classic and microform HPE. In classic HPE the two most commonly mutated genes are ZIC2and SHH. Probands with ZIC2mutations have been identified across the entire HPE phenotypic spectrum, the vast majority resulting in classic HPE; however, mutations in ZIC2can also cause microform HPE.
Analysis of HPE cohorts reveals allelic heterogeneity whereby different mutations in the same gene give rise to similar phenotypes. There are more than 118 unique ZIC2variants associated with HPE that have been reported. The vast majority of these occur in the ZIC2coding region and are predicted to substantially alter the ZIC2transcript rather than a single amino acid.
Of the reported non-coding ZIC2variants, most are splice-site mutations predicted to significantly alter transcript structure. The remaining non-coding variants are single-nucleotide changes located in a conserved region of the 3′ untranslated region (UTR). Overall, approximately 98% of ZIC2-associated HPE variants are predicted loss-of-function mutations.
The zinc finger domain (ZFD) is the primary location of ZIC2mutations, with nearly half of all mutations occurring in this region. 73% of ZFD mutations are missense, demonstrating that single amino acid changes in the ZFD are sufficient to cause disease. In contrast, few missense variants occur outside of the ZFD where 86% of known variants are predicted to alter the ZIC2transcript. Mutations further occur in the c-terminal alanine repeat, a region that influences the strength of ZIC2 DNA binding and its transcriptional activity.
The brain and midline defects characteristic of HPE originate during embryonic gastrulation as a result of abnormal neural patterning. Ventral forebrain patterning depends on inductive interactions between the anterior neurectoderm and the underlying prechordal plate (PrCP), as demonstrated in mouse models.
Anterior midline patterning is exceptionally conserved between humans and mice, meaning mouse models can be used to understand the molecular mechanisms underpinning HPE. Current murine model evidence indicates that ZIC2 has at least two critical roles during forebrain development. During mid-gastrulation, ZIC2 functions at the embryonic node to shape the NODAL signalling gradient. Severe Zic2 loss-of-function mutations lead to a transient failure in the production of the anterior notochord, and, consequently, PrCP fate is not stabilised. As such, the SHH signal is not sent from the PrCP and the ventral forebrain signalling centre is not established, resulting in gross perturbations in forebrain D–V pattern and classic HPE.
ZIC2