FMR1 and its paralog FXR1 encode the RNA-binding proteins FMRP and FXR1P, respectively, which can both homodimerize as well as heterodimerize to regulate mRNA stability, transport and translation. A broad body of literature implicates FMRP or FXR1P either directly or indirectly in Autism Spectrum Disorder (ASD), Fragile X Syndrome (FXS) and Schizophrenia (SCZ). However, the mechanisms by which they produce dysfunction in the human brain, and the convergence of these mechanisms across distinct neurodevelopmental disorders have not been fully elucidated. Importantly, our existing understanding of FMRP and FXR1P biology has not translated into clinical utility. This application is therefore focused on dissecting mechanisms of FMRP and FXR1P function and dysfunction in human cortical neurons. Several fundamental questions remain about the neurobiological functions of FMRP and FXR1P in both the normal and disease states. Importantly, the mRNA cargoes of FMRP and FXR1P have not been defined in human brain cell types, and it remains unclear whether they regulate different mRNA cargoes in different cell types or at different stages of neuronal development. The confirmation of previously identified mechanisms from mouse models, as well as the identification of novel mechanisms in human neurons would both have important clinical implications. With the recent surge in human psychiatric genetic data, it is also unclear to what extent FMRP and FXR1P regulate emerging ASD and SCZ susceptibility genes. In order to address these important questions, we will define the mRNA cargoes of FMRP and FXR1P in in vitro derived human cortical neurons and integrate this information with existing mouse brain datasets and emerging human psychiatric genetic data (Specific Aim I). The extent to which FMRP and FXR1P have distinct vs. overlapping functions and targets is also an open question. In order to address this fundamental question and to better understand disease mechanisms, we will analyze the mRNA cargoes of FXR1P in the context of FMRP loss, and the mRNA cargoes of FMRP in the context of FXR1P loss (Specific Aim II). Finally, we will assess how loss of FMRP or FXR1P impacts the expression, localization and translation of specific mRNA cargoes (Specific Aim II). In order to carry out the aims described above, we have engineered human pluripotent stem cells (hPSCs) with gene-disrupting mutations in FMR1 and FXR1, and we are now separately adding affinity tags to the endogenous loci. We will couple these resources with a highly efficient neuronal differentiation paradigm to generate excitatory cortical neurons from hPSCs in vitro. Based on extensive molecular and functional analyses, these neurons most closely resemble excitatory neurons from superficial cortical layers, a population which post mortem studies implicate in the pathogenesis of ASD and SCZ. Overall, this project will define functions of FMRP and FXR1P in human neurons in both the normal and perturbed states to better understand normal neurobiological functions as well as potentially convergent disease mechanisms.

Public Health Relevance

RNA-binding protein dysfunction is strongly implicated in multiple neurodevelopmental disorders including Autism Spectrum Disorder (ASD), Fragile X Syndrome (FXS) and Schizophrenia (SCZ). The goal of this project is to elucidate the functions of two related RNA-binding proteins, FMRP and FXR1P, in human cortical neurons, and how their dysfunction may contribute to disease. These molecular and cellular analyses are essential for understanding neurobiological mechanisms and improving therapeutic strategies.

National Institute of Health (NIH)
National Institute of Mental Health (NIMH)
Exploratory/Developmental Grants (R21)
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Developmental Brain Disorders Study Section (DBD)
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Beckel-Mitchener, Andrea C
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Broad Institute, Inc.
United States
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