General population ZBTB18 missense variants influence DNA binding and transcriptional regulation

Genetic variation to the multi-zinc finger BTB domain transcription factor ZBTB18 can cause a spectrum of human neurodevelopmental disorders, but the underlying mechanisms are not well understood. Recently, we reported that pathogenic, de novo ZBTB18 missense mutations alter its DNA-binding specificity and gene regulatory functions, leading to human neurodevelopmental disease. However, the functional impact of general population ZBTB18 missense variants is unknown. Here, we investigated such variants documented in the Genome Aggregation Database (gnomAD) to discover that ZBTB gene family members are intolerant to loss-of-function and missense mutations, but not synonymous mutations. We studied ZBTB18 as a protein-DNA complex to find that general population missense variants are rare, and disproportionately map to non DNA-contact residues, in contrast to the majority of disease-associated variants that map to DNA-contact residues, essential to motif-binding. We studied a selection of variants (n=12), which spans the multi-zinc finger region to find 58.3% (7/12) of variants displayed altered DNA-binding, 41.6% (5/12) exhibited altered transcriptional activity in a luciferase reporter assay, 33.3% (4/12) exhibited altered DNA-binding and transcriptional activity, while 33.3% (4/12) displayed negligible functional impact. Our results demonstrate that general population variants, while rare, can influence ZBTB18 function, with potential consequences on neurodevelopment, homeostasis and disease. This article is protected by copyright. All rights reserved.