Doublecortin mutation disrupts lattice-dependent proximity to reveal cytoskeletal networks in developing neurons

Doublecortin (DCX) is a microtubule-associated protein (MAP) required for neuronal migration and cortical development. However, DCX is not sufficient to direct these processes on its own. Instead, nucleokinesis, growth cone advancement, collateral branching, and polarity depend on the coordination of different MAPs and cytoskeletal proteins, including DCX, to define distinct polymer properties and cytoskeletal interfaces. While many of these proteins have been investigated for their mechanistic cytoskeletal engagement, the spatiotemporal proteomic landscape of developing neurons remains largely uncharacterized. Here, we explore DCX-centric cytoskeletal networks using BioID accumulated throughout the differentiation of cortical neurons from induced pluripotent stem cells (iPSCs). Using the morphological staging of these cells as temporal landmarks, we generate a time-resolved proteomic atlas that charts DCX's molecular environment throughout differentiation. From this atlas, we identify a cluster of cytoskeletal proteins whose persistent proximity to DCX is disrupted with the disease-associated mutation, DCX p.Arg178Leu (R178L). Consistent with R178L's diminished cooperativity, we find many of these species are implicated in modulating microtubule stability and intracellular transport, including proteins EML4 and MAP7D1. Together, our findings place DCX at the interface of distinct microtubule populations and cytoskeletal systems throughout neuronal development.