Post-doctoral Research Fellow Emory University Atlanta, Georgia, United States
Abstract: Fragile X syndrome (FXS), a leading genetic cause of intellectual disability and autism spectrum disorder, is typically caused by CGG trinucleotide repeat expansion in the FMR1 gene that results in gene silencing and complete loss of its protein product, fragile X messenger ribonucleoprotein (FMRP). FMRP is a multifunctional protein that binds to selective mRNAs and regulates their stability, editing, transport, and translation. As a key brain region for learning, memory, and emotional processing – and a major site of FMR1 expression – the human hippocampus is of particular interest, especially in understanding how FMRP loss affects its development. Indeed, anxiety and aggressive behaviors are common in FXS patients, and hippocampal enlargement has been reported in younger patients, suggesting hippocampal abnormalities. However, the regulatory role of FMRP and the impacts of its loss during human hippocampal development remain unexplored. To address these questions, we generated human hippocampal organoids (HOs) from FXS patient-derived and healthy control-derived iPSCs. Through transcriptomic, cellular, and electrophysiological analyses, we observed altered developmental trajectories, specifically increased neurogenesis and decreased gliogenesis in FXS HOs, which may contribute synergistically to neural network hyperexcitability in them. To investigate potential underlying mechanisms, we performed eCLIP-seq on HOs at the early and late stages and identified stage-specific FMRP targets that correspond to the predominant biological processes at each stage. Particularly, we demonstrated, for the first time, a switch in FMRP binding targets from mRNAs involved in cell cycle and neurogenesis at early stage to those involved in gliogenesis at late stage, highlighting a dynamic temporal regulatory role of FMRP during hippocampal development. Further construction of gene regulatory networks in HOs by single-cell transcriptomic analyses revealed key regulons targeted by FMRP that may drive altered developmental trajectories in FXS HOs. Together, our study delineates the molecular, cellular, and functional impacts of FMRP loss on hippocampal development and provides new insights into its regulatory role and RNA binding dynamics, offering potential avenues for therapeutic advancement.
