Single-cell proteomic landscape of the developing human brain

Profiling protein abundance and dynamics at single-cell resolution in complex human tissues is challenging. Given the discordance between transcript and protein abundance observed in studies of the human cerebral cortex, we developed an optimized workflow that combines label-free single-cell mass spectrometry with precise sample preparation to resolve quantitative proteomes of individual cells from the developing human brain. Our method achieves deep proteomic coverage (~800 proteins per cell) even in small immature prenatal human neurons (diameter ~7-10 μm, ~50 pg protein), capturing major brain cell types and enabling proteome-wide characterization at single-cell resolution. We document extensive transcriptome-proteome discordance across cell types, particularly in genes associated with neurodevelopmental disorders. Proteins exhibit markedly higher cell-type specificity than their mRNA counterparts, underscoring the importance of proteomic-level analysis. By reconstructing developmental trajectories from radial glia to excitatory neurons at the proteomic level, we identify dynamic, stage-specific protein co-expression modules and pinpoint the intermediate progenitor-to-neuron transition as a genetically vulnerable phase associated with autism.