Developmental consequences of epileptic encephalopathy- and intellectual disability-associated EEF1A2 mutations in human neurons

PROJECT SUMMARY Epilepsy affects over 50 million people worldwide, with developmental and epileptic encephalopathies (DEEs) representing the most severe and treatment-resistant forms. DEEs are characterized by early-onset seizures and developmental delays, often caused by pathogenic variants in genes critical for brain function. Understanding these genetic underpinnings has revolutionized epilepsy research, paving the way for precision medicine—therapeutic strategies tailored to specific genetic and molecular mechanisms. Among these disorders, EEF1A2-related neurodevelopmental disorder (EEF1A2 syndrome) stands out as a rare yet compelling example, highlighting the need for targeted research to understand and treat epilepsy. Mutations in the EEF1A2 gene, which encodes the eukaryotic elongation factor 1A2 (eEF1A2), have been identified as a cause of severe neurodevelopmental disorders, including DEEs. eEF1A2 facilitates protein synthesis by delivering aminoacyl-tRNAs to ribosomes in a GTP-dependent manner. Approximately 50 distinct missense mutations have been reported, with recurrent variants such as G70S, E122K, and D252H associated with severe intellectual disability. These mutations map to key functional domains of eEF1A2—GTP binding (G70S), tRNA binding (E122K), and actin binding (D252H)—and exhibit distinct clinical phenotypes. While G70S and E122K are linked to early-onset epilepsy, D252H is associated with milder or no epilepsy. The role of eEF1A2 in actin dynamics through its third functional domain is also unclear, suggesting potential additional roles in neurons. This project aims to address these gaps through two specific aims: 1. Determining how EEF1A2 mutations alter protein synthesis in human neurons. 2. Determining how EEF1A2 mutations alter actin dynamics, neuronal morphology, and development. Previous studies have been limited by the species-specific isoform switch between eEF1A1 and eEF1A2 that occur in humans but are not fully replicated in mouse models. By utilizing human-induced pluripotent stem cells (hiPSCs) differentiated into cortical neurons, this research will overcome these limitations by determining the alterations due to these mutations on protein synthesis, elongation rate, translatome profile, and translational efficiency (Aim 1). Additionally, alterations in actin mobility and subcellular structure, as well as neuronal morphology, synapse number, and neuronal electrophysiological properties will be determined (Aim 2). The findings will advance our understanding of eEF1A2's neuron-specific roles and inform precision medicine strategies for EEF1A2 syndrome treatment.