Sparse Neuronal C4 Elevation Induces Network-Wide Transcriptomic and Lipid-Droplet Remodeling in the Cortical Microenvironment

The complement component C4 regulates synaptic refinement and plasticity in the brain and has been implicated in multiple neurological and psychiatric disorders, with especially strong genetic evidence linking elevated C4A expression to schizophrenia. Although altered C4 expression has been associated with disease risk, it remains unclear whether localized C4 elevation in a small neuronal population is sufficient to reshape broader cortical gene-regulatory programs and cellular phenotypes. To address this question, we overexpressed mouse C4 in approximately 2% of prefrontal cortical neurons using in utero electroporation. Bulk RNA sequencing of microdissected prefrontal cortex revealed widespread transcriptional changes across tissue composed predominantly of untransfected cells, indicating non-cell-autonomous effects of sparse neuronal C4 elevation. C4-OE induced coordinated changes in genes involved in cholesterol biosynthesis, axon guidance, synaptic plasticity, cytoprotection, neurodevelopment, and inflammatory regulation. Co-expression analysis identified a C4-containing co-expression module enriched for dendritic development, synapse organization, and cell-cycle-associated programs, while comparison with human schizophrenia synaptic proteomic datasets identified selective gene- and pathway-level overlap in pathology-relevant processes. Reference-based projection of directional DEG signatures onto cortical single-cell transcriptomic atlases further identified cell-type-associated transcriptional structure, including a prominent astrocyte-associated axis within the C4-OE-downregulated signature and structured lipid-metabolic, vascular-immune, and immune/stress correlation domains. Spatial validation with MFISH showed increased Hmgcr expression in neighboring GFP-negative cells, and PLIN2 imaging revealed increased DAPI-associated/perinuclear lipid-droplet burden within the local C4-OE field. Together, these findings demonstrate that sparse neuronal C4 elevation is sufficient to reprogram the cortical microenvironment through network-wide transcriptomic remodeling and neighboring-cell lipid-droplet accumulation. This work expands the interpretation of C4 risk biology beyond synapse elimination alone, suggesting that focal immune-gene dysregulation can engage lipid-homeostatic, stress-regulatory, and cell-type-associated transcriptional programs that may contribute to cortical circuit vulnerability. Author SummaryComplement component C4 is best known for its role in immune defense, but increasing evidence suggests that abnormal C4 levels can also influence brain development and disease-relevant neural circuits. In this study, we asked what happens when C4 is increased in only a sparse subset of cortical neurons. Using a mouse model in which C4 was elevated in a small population of prefrontal cortical neurons, we found that this local manipulation was associated with broad molecular changes in the surrounding cortical microenvironment. Transcriptomic analyses revealed coordinated changes in genes related to neuronal activity, synaptic signaling, immune and stress responses, vascular-associated pathways, and lipid metabolism. Importantly, these effects were not restricted to the directly manipulated neurons, suggesting that focal complement dysregulation can influence neighboring-cell and local microenvironmental states. We further validated a lipid-associated component of this response by PLIN2 immunostaining, which revealed increased lipid droplet burden in the local C4-elevated cortical field. Together, these findings support the idea that altered levels of an immune-linked gene can modify local cortical microenvironments and engage lipid- and metabolic-stress pathways relevant to brain vulnerability.