A Morpho-Proteomic Atlas of Mitosis at Sub-Minute Resolution

Precise spatiotemporal protein organization is critical for fundamental biological processes including cell division ^1,2 . Indeed, aberrant mitosis and mitotic factors are involved in diverse diseases, including various cancers ^3,4 , Alzheimer’s disease ⁵ , and rare diseases ⁶ . During mitosis, complex spatial rearrangements and regulation ensure the accurate separation of replicated sister chromatids to produce genetically identical daughter cells ^7–9 . Previous studies employed high-throughput methodologies to follow specific proteins during mitosis ^10–15 . Still a temporally refined systems-level approach capable of monitoring morphological and proteomic changes throughout mitosis has been lacking. Here, we achieved unprecedented resolution by phenotypically decomposing mitosis into 40 subsections of a regression plane for proteomic analysis using deep learning and regression techniques. Our deep visual proteomics (DVP) workflow ¹⁶ , revealed rapid, dynamic proteomic changes throughout mitosis. We quantified 4,350 proteins with high confidence, demonstrating that 147 show significant dynamic abundance changes during mitotic progression. Clustering revealed coordinated patterns of protein regulation, while network analysis uncovered tight regulation of core cell cycle proteins and a link between cell cycle and cancer-linked mutations. Immunofluorescence validated abundance changes and linked previously uncharacterised proteins, like C19orf53, to mitosis. To facilitate data navigation, we developed Mito-Omix, a user-friendly online platform that integrates intricate morphological and molecular data. Our morphological and proteomic dataset spans mitosis at high resolution, providing a rich resource for understanding healthy and aberrant cell division.