To define the clinical spectrum and mechanisms of CDK19-related neurodevelopmental disorders (NDDs) by integrating human genotype–phenotype data with in vivo functional and transcriptomics.

CDK19 encodes a subunit of the Mediator complex, essential for RNA polymerase II–driven transcription during brain development. Pathogenic CDK19 variants have recently been implicated in epilepsy and NDDs, but the spectrum of clinical features and molecular mechanisms remains poorly characterized due to the absence of large-scale analyses.

An international consortium assembled a cohort of 28 individuals harboring rare heterozygous CDK19 variants (12 novel, 16 previously reported), encompassing 17 distinct alleles. Detailed clinical data were correlated with variant topology within the kinase and Mediator-binding domains. Single-cell RNAseq (scRNAseq) from the AllenBrain Atlas was used for cell-specific and temporal expression. Functional characterization employed Drosophila models expressing patient-derived alleles under ubiquitous and neuron-specific promoters. Viability, behavioral, and morphometric assays assessed functional severity, while RNA sequencing of larval brains showed transcriptional alterations.

All individuals exhibited global developmental and speech delay; 67% had epilepsy (predominantly infantile epileptic spasms), 50% hypotonia, and 57% brain MRI abnormalities, callosal thinning or white-matter loss. scRNAseq localized CDK19 expression to excitatory glutamatergic cortical projection neurons (SLC17A7) and oligodendrocyte lineages (MOG, OLIG1, OLIG2). In Drosophila, severe alleles (p.D18Y, p.T196K, p.G420R) caused complete lethality (0% viability), while p.K153R reduced survival to 27.3±3.8% (P<0.001). Neuron-specific expression shortened lifespan (median 5–15 days vs. 40 days; P<0.001) and increased climbing latency by 65±9% (P<0.01). Mitochondrial length doubled (2.1±0.3 µm vs. 1.0±0.2 µm; P<0.001), and nuclear CDK19 puncta dropped >80%. Transcriptomics revealed 209 dysregulated genes (115 upregulated, 94 downregulated), with downregulation of GluRIIA, Vgat, and Syn (P_adj=2.1×10?4).

Pathogenic CDK19 variants impair nuclear localization and transcriptional regulation, disrupting synaptic and neuromuscular gene networks. This study establishes allele-specific functional severity, expands the clinical spectrum of CDK19-related NDDs, and highlights a translational model for "Mediator complexopathies".