From Cell Type to Structure: A Multiscale Framework for Discovering Psychiatric Genes, Pathways and Mechanisms

Abstract

Human genetics is perhaps the most fundamental biomarker for neuropsychiatric disorders. While genetic studies have identified numerous risk loci for neuropsychiatric disorders, the biological mechanisms they perturb remain unclear. We generated single-nucleus RNA sequencing (snRNA-seq) data from across multiple human brain regions to weight neuronal co-expression patterns by polygenic heritability, enabling the identification of disease-relevant pathways from common variant architecture. This framework was robustly validated through convergence with rare variant signals from large-scale exome sequencing data across multiple neuropsychiatric disorders, revealing Ca2+ homeostasis as a central and recurrent axis of genetic vulnerability. Within this pathway, we identified ATP2B2 – a P-type ATPase responsible for Ca2+ extrusion – as consistently downregulated in the prefrontal cortex in donors with schizophrenia compared to controls, both synaptic proteomes and snRNA-seq. This reduction is specific to excitatory neurons, pointing to a cell type-specific loss-of-function mechanism linking ATP2B2 to schizophrenia risk. ATP2B2 displays a striking enrichment of missense variants implicated in schizophrenia, autism and neurodevelopmental disorders. To investigate the structural basis of this signal, we developed a 3D enrichment framework that leverages the AlphaFold 3-predicted structure to pinpoint structurally constrained mutational hotspots with likely functional impact. We did this by testing for an excess of case-derived variants within 15Å spherical neighborhoods around each residue, and identify compelling candidates for downstream mechanistic interrogation. We identified an enrichment of case-derived variants localized in close spatial proximity to both the Ca2+ permeation tunnel and binding site and the ATP:Mg2+ coordination site, suggesting two distinct mechanisms of ATP2B2 perturbation. In the Ca2+ binding neighborhood, substitution of the Ca2+-coordinating residue E457 with lysine (E457K) introduces a charge-reversal, suggesting disrupted binding as a focal mechanism of pathogenic variation in ATP2B2. We used AlphaFold 3 to simulate ATP2B2 and Ca2+ with and without E457K, and found it markedly reduced Ca2+ contact probabilities relative to wildtype, supporting an LoF effect. We validated E457K’s impact in two orthogonal assays. The variant abolished ATPase activity in recombinant ATP2B2 and in a cellular context it impaired Ca2+ extrusion in HEK293 cells using a GCaMP6s-based imaging assay - both consistent with a LoF mechanism aligned with the direction of genetic risk. This suggests that case variants in ATP2B2 very likely compromise its function and disrupt intracellular Ca2+ homeostatic equilibrium. Our study constitutes a significant contribution to the neurobiological elucidation of etiological genetic risk and advances mechanistic insight into the pathogenesis of neuropsychiatric disorders. First, in a biochemical assay, the variant completely abolished ATPase activity in recombinant ATP2B2. Second, in a cellular model, GCaMP6s-based Ca2+ imaging in HEK293 cells revealed a marked impairment in Ca2+ extrusion, indicative of disrupted Ca2+ clearance. Both findings converge on a loss-of-function mechanism, consistent with the direction of genetic risk observed in neuropsychiatric cases as well as the downregulation seen in functional genomics datasets. These results strongly suggest that pathogenic variants in ATP2B2 compromise its physiological role in maintaining intracellular Ca2+ homeostasis. Our study constitutes a significant contribution to the neurobiological elucidation of etiological genetic risk, and provides a mechanistic link between rare genetic variation and disrupted neuronal Ca2+ signaling, offering novel insight into the molecular pathogenesis of neuropsychiatric disorders