Function of WWP1 Variant in Undiagnosed Neurodevelopmental Disease

Abstract

Developing effective treatment plans for rare undiagnosed diseases poses a significant challenge due to the inherent difficulty in diagnosing these conditions. Consequently, treatment strategies often center on managing symptoms, but the results are frequently temporary and insufficient. Recent advancements in sequencing techniques have facilitated the diagnosis of patients with rare undiagnosed diseases by employing sequencing to identify pathogenic variants. Although genomic studies offer insights into potential causal genes, functional studies through disease modeling are necessary to validate the direct involvement of the gene in the disease.

In an undiagnosed clinical case, WWP1 D793N, a de novo heterozygous mutation has been identified as the top disease-associated mutation candidate by whole exome sequencing. The patient exhibits symptoms including diffuse brain atrophy, corpus callosum thinning, epilepsy, and no independent walking at 10 years old. Notably, around 13% of E3 ligase genes were found to have mutations in a neurological disorder, underscoring the importance of E3 ligase in neurological disorders. WWP1 has also been implicated in cancers, infectious diseases, and neurological disorders. However, its specific role in neurodevelopment remains understudied.

In this study, our primary objectives include characterizing the functionality of WWP1 D793N and elucidating the underlying cellular mechanism contributing to the pathology in the patient. In our investigations, WWP1 D793N overexpression in developing mouse brains via IUE resulted in migration defects and cell death in neighboring cells during early development. To further elucidate the underlying pathological mechanisms, we established the WWP1 variant overexpressing HeLa cell lines for phenotyping and transcriptomic analysis. Transcriptomic profiling of WWP1 D793N overexpressing cell lines indicated an upregulation in apoptosis and downregulation in neurogenesis and cell adhesion-related gene expression. However, limitations inherent to the WWP1-HeLa cell lines precluded the characterization of significant changes in cellular activities such as apoptosis. To overcome this, we employed transient gene expression in human neural progenitor cells, uncovering morphological defects that ultimately led to apoptosis upon WWP1 D793N overexpression. Furthermore, biochemical functionality tests via ubiquitination assays indicated that WWP1 D793N promotes increased autopolyubiquitination of the protein. This suggests a gain-of- function characteristic for this mutant, providing valuable insights into the altered cellular dynamics associated with the D793N mutation in WWP1.

Overall, our study aims to provide significant insights into the comprehension of WWP1- related neurodevelopmental disorders and subsequently offers valuable perspectives for the development of treatments for rare diseases.