Pathological disruption of CELF2 shuttling causes neuronal hyperactivity, learning deficits, and seizures
Michelle Hua, et al.
Michelle Hua, et al.
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Research Article Clinical Research Development Genetics

Pathological disruption of CELF2 shuttling causes neuronal hyperactivity, learning deficits, and seizures

Abstract

De novo heterozygous variants in CUGBP Elav-like family member 2 (CELF2) have recently been associated with a rare neurodevelopmental disorder, yet the mechanisms linking specific variants to distinct clinical phenotypes remain poorly understood. Here, we reported a cohort of 18 individuals and provided evidence that variants causing CELF2 mislocalization, but not protein-null variants, were associated with seizures. Using proband-derived human cortical neurons and transgenic mouse models, we demonstrated that CELF2 underwent activity-dependent nucleocytoplasmic shuttling in excitatory neurons and that its cytoplasmic retention caused neuronal hyperactivity, elevated seizure susceptibility, and learning and memory deficits. We further found that cytoplasmic CELF2 regulated mRNAs critical for synaptic function and neuronal excitability and implicated in epileptic seizures and intellectual disability. Drug screening further identified AKT signaling as a key regulator of CELF2 nucleocytoplasmic shuttling and a candidate target for reversing neuronal hyperactivity. Together, our findings expand the clinical and genetic spectrum of CELF2-related neurodevelopmental disorders and establish a variant-specific mechanism that links CELF2 mislocalization to neuronal hyperactivity, seizures, and cognitive impairment.

Authors

Michelle Hua, Mohamad-Reza Aghanoori, Melissa J. MacPherson, Yi Ren, Shehani V. Siripala, Yifan Yang, Yvonne Yan Yan Or, Malea Nguyen, Robert Duba-Kiss, Daniel Feng, Laura Williams, Christopher J. Gafuik, GengYi Wang, Chloe Quelin, Boris Keren, Sarah Schuhmann, Georgia Vasileiou, Alexia Bourgois, Antonio Vitobello, Christophe Philippe, Zornitza Stark, Richard J. Leventer, George McGillivray, Frederic Tran Mau-Them, Marine Tessarech, Clément Prouteau, Phillis Lakeman, Mahdi M. Motazacker, Donald R. Latner, Raymond C. Caylor, Yvette van Ierland, Eloise Prijoles, Angie Lichty, Evangelos Theodorou, David A. Sweetser, Edward Steel, Jan Cobben, Majed J. Dasouki, Daniel G. Calame, Bertrand Isidor, Benjamin Cogné, Mitchell Kesler, Brooke Rackel, Isabel Clark, Deborah M. Kurrasch, G. Campbell Teskey, James Ellis, Guiqiong He, Scott D. Ryan, Douglas J. Mahoney, A. Micheil Innes, Jonathan R. Epp, Guang Yang

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Figure 6

Neuronal hyperactivity caused by CELF2 mislocalization elevates seizure susceptibility and disrupts learning and memory.

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Neuronal hyperactivity caused by CELF2 mislocalization elevates seizure ...
(A and B) Racine score progression following PTZ injection in female (A) and male (B) WT and KI mice. Shaded areas represent SEM. Mantel-Cox log-rank test with Benjamini-Hochberg correction. **P = 0.008. (C) Schematic of the experimental setup for fiber photometry and contextual fear conditioning (CFC). Three weeks after AAV-mediated GCaMP8f expression and fiber implantation in the CA1 region, neuronal responses were recorded during foot shock trials and subsequent retrieval tests. (D) Contextual fear memory retention in mice. (E and F) The fractional change in fluorescence relative to a baseline signal (ΔF/F) traces recorded during CFC acquisition. (G) Quantification of photometry amplitude in response to foot shocks. (H and I) Photometry ΔF/F traces during memory retrieval, aligned to freezing-to-moving transitions (±2 seconds from freezing offset). (J and K) Quantification of peak frequency (J) and AUC (K) of the photometry signal during freezing-to-moving transitions. (L) Schematic showing sequential context exploration and catFISH with 4 nuclear expression profiles of immediate-early genes (IEGs) Arc and H1a mRNA in response to contexts I and II. (M) Confocal images of H1a (red) and Arc (green) mRNA in hippocampal CA1 neurons. (N) Confocal images of hippocampal CA1 regions following catFISH after context exploration. Red, green, white, and empty arrows indicate H1a-only, Arc-only, double-positive, and IEG-negative neurons, respectively. Nuclei were counterstained with Hoechst 33258 and outlined with dashed white lines. (O) Quantification of IEG expression in CA1 neurons from N. n ≥ 9 each. Means ± SEM. Each dot represents 1 animal. Two-way ANOVA, Tukey’s post hoc test (D, G, J, and K), and Mann-Whitney U test (O). Scale bars: 5 μm (M), 10 μm (N).