Boris Keren

De novo heterozygous variants in 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.

Disorders of GABRA3 , the only epilepsy-associated GABA _A receptor subunit gene on the X chromosome, have eluded clinical clarity due to ambiguous inheritance patterns and variable phenotypes. The long-standing assumption that all pathogenic variants cause loss of function further obscured genotype–phenotype relationships and hindered progress. Here, we curated a cohort of individuals with a GABRA3 variant, integrating deep phenotyping, genotyping, family history, and electrophysiology with a targeted mouse model. Among 43 individuals with 19 GABRA3 variants, functional analyses revealed gain- and loss-of-function effects, each linked to distinct clinical profiles. Gain-of-function variants were associated with severe, treatment-resistant epilepsy and profound intellectual disability, disproportionately affecting males, who were often nonambulant and had cortical visual impairment. Loss-of-function variants produced milder phenotypes, with epilepsy rarely observed; affected males showed behavioral issues and language delay, while females were unaffected carriers. Our gain-of-function ( Gabra3 ^Q242L/+ ) mouse model mirrored these sex-specific differences, showing increased seizure susceptibility, early death, and marked cortical hyperexcitability. These insights resolve longstanding uncertainties surrounding GABRA3 and redefine how X-linked disorders are interpreted. They demonstrate that it is the functional impact of a variant, not its mere presence, that determines whether a condition manifests dominantly or recessively. This distinction carries important implications for genetic counseling, precision medicine, and the broader interpretation of X-linked neurodevelopmental disorders.

Variants in RNU4-2 , encoding the small nuclear RNA (snRNA) U4, were recently identified as a major cause of neurodevelopmental disorders (ReNU syndrome). Here, we investigated de novo variants in 50 snRNAs in a French cohort of 23,649 individuals with rare disorders and collected data of additional patients through an international collaboration. Altogether, we identified 133 probands with pathogenic or likely pathogenic variants in RNU4-2 and 15 individuals with de novo and/or recurrent variants in constrained regions of RNU5B-1 , one of five genes encoding U5. These variants cluster in evolutionarily conserved regions of U4 and U5 essential for splicing. RNU4-2 variants affecting stem III are associated with milder phenotypes than those in the T-loop (quasi-pseudoknot). Phaseable variants associated with severe phenotypes occurred on the maternal allele. Individuals with RNU4-2 variants show specific defects in alternative 5’ splice site usage, correlating with variant location and clinical severity. Additionally, we report an episignature associated with severe ReNU syndrome. This study further highlights the importance of de novo variants in snRNAs and establishes RNU5B-1 as a new neurodevelopmental disorder gene.

Objective: De novo variants in cullin‐3 ubiquitin ligase (CUL3) have been strongly associated with neurodevelopmental disorders (NDDs), but no large case series have been reported so far. Here, we aimed to collect sporadic cases carrying rare variants in CUL3, describe the genotype–phenotype correlation, and investigate the underlying pathogenic mechanism.Methods: Genetic data and detailed clinical records were collected via multicenter collaboration. Dysmorphic facial features were analyzed using GestaltMatcher. Variant effects on CUL3 protein stability were assessed using patient‐derived T‐cells.Results: We assembled a cohort of 37 individuals with heterozygous CUL3 variants presenting a syndromic NDD characterized by intellectual disability with or without autistic features. Of these, 35 have loss‐of‐function (LoF) and 2 have missense variants. CUL3 LoF variants in patients may affect protein stability leading to perturbations in protein homeostasis, as evidenced by decreased ubiquitin‐protein conjugates in vitro. Notably, we show that 4E‐BP1 (EIF4EBP1), a prominent substrate of CUL3, fails to be targeted for proteasomal degradation in patient‐derived cells.Interpretation: Our study further refines the clinical and mutational spectrum of CUL3‐associated NDDs, expands the spectrum of cullin RING E3 ligase‐associated neuropsychiatric disorders, and suggests haploinsufficiency via LoF variants is the predominant pathogenic mechanism. ANN NEUROL 2025;97:76–89

Humans with monogenic inborn errors responsible for extreme disease phenotypes can reveal essential physiological pathways. We investigated germline mutations in GNAI2 , which encodes G _αi2 , a key component in heterotrimeric G protein signal transduction usually thought to regulate adenylyl cyclase–mediated cyclic adenosine monophosphate (cAMP) production. Patients with activating G _αi2 mutations had clinical presentations that included impaired immunity. Mutant G _αi2 impaired cell migration and augmented responses to T cell receptor (TCR) stimulation. We found that mutant G _αi2 influenced TCR signaling by sequestering the guanosine triphosphatase (GTPase)–activating protein RASA2, thereby promoting RAS activation and increasing downstream extracellular signal–regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K)–AKT S6 signaling to drive cellular growth and proliferation.

SUMMARY: Variable expressivity of disease-associated variants implies a role for secondary variants that modify clinical features. We assessed the effects of modifier variants towards clinical outcomes of 2,252 individuals with primary variants. Among 132 families with the 16p12.1 deletion, distinct rare and common variant classes conferred risk for specific developmental features, including short tandem repeats for neurological defects and SNVs for microcephaly, while additional disease-associated variants conferred multiple genetic diagnoses. Within disease and population cohorts of 773 individuals with the 16p12.1 deletion, we found opposing effects of secondary variants towards clinical features across ascertainments. Additional analysis of 1,479 probands with other primary variants, such as 16p11.2 deletion and CHD8 variants, and 1,084 without primary variants, showed that phenotypic associations differed by primary variant context and were influenced by synergistic interactions between primary and secondary variants. Our study provides a paradigm to dissect the genomic architecture of complex disorders towards personalized treatment.

Autophagy is a fundamental and evolutionary conserved biological pathway with vital roles in intracellular quality control and homeostasis. The process of autophagy involves the engulfment of intracellular targets by autophagosomes and their delivery to the lysosome for digestion and recycling. We have previously reported recessive variants in EPG5 , encoding for the ectopic P-granules 5 autophagy protein with a crucial role in autophagosome-lysosome fusion, as the cause of Vici syndrome (VS), a severe multisystem neurodevelopmental disorder defined by a combination of distinct clinical features including callosal agenesis, cataracts, cardiomyopathy, immunodeficiency, and hypopigmentation. Here, we present extensive novel genetic, clinical, neuroradiological and pathological features from the largest cohort of EPG5 -related disorders reported to date, complemented by experimental findings from patient cells and models of EPG5 defects in Caenorhabditis elegans and Mus musculus . We identified 200 patients with recessive EPG5 variants, 86 of them previously unpublished. The associated phenotypic spectrum ranged from antenatally lethal presentations and the classic VS phenotype (n=60) to much milder neurodevelopmental disorders with less specific manifestations (n=140). Myopathic features and epilepsy with variable progression were frequently observed. Novel manifestations included early-onset parkinsonism and dystonia with cognitive decline during adolescence, hereditary spastic paraplegia (HSPP), and myoclonus. Radiological findings included previously recognized EPG5 -related features with callosal abnormalities and pontocerebellar hypoplasia, and a range of novel features suggesting an emerging continuum with disorders of brain iron accumulation or copper metabolism as well as HSPPs. Genotype-phenotype studies suggested a correlation between predicted residual EPG5 expression and clinical severity, especially regarding disease progression and survival. The Epg5 p.Gln331Arg knock-in mouse, a model of milder EPG5 -related disorders, showed an age-related motor phenotype and impaired autophagic clearance in several brain regions mirroring those also affected in humans. In Caenorhabditis elegans, epg-5 knockdown gave rise to neurodevelopmental features and motor impairment comparable to defects in parkinsonism-related genes, abnormal mitochondrial respiration, and impaired mitophagic clearance early in life. Cellular assays revealed impaired PINK1-Parkin dependent mitophagic clearance in patient fibroblasts. Our findings expand the phenotypic spectrum of EPG5 -related disorders and indicate a life time continuum of disease that overlaps with other disorders of defective autophagy and intracellular trafficking. Our observations also suggest close links between early-onset neurodevelopmental and neurodegenerative conditions of later onset due to EPG5 defects, in particular dystonia and parkinsonism, highlighting the fundamental importance of dysfunctional autophagy in the pathophysiology of common neurodegenerative disorders.