Data for: Doublecortin reinforces microtubules to promote growth cone advance in soft environments

Doublecortin (DCX) is a microtubule (MT)-associated protein in immature neurons. DCX is essential for early brain development, and DCX mutations account for nearly a quarter of all cases of lissencephaly-spectrum brain malformations that arise from a neuronal migration failure through the developing cortex. By analyzing pathogenic DCX missense mutations in non-neuronal cells, we show that disruption of MT-binding is central to DCX pathology. In human induced pluripotent stem cell (hiPSC)-derived cortical i3Neurons genome-edited to express DCX-mEmerald from the endogenous locus, DCX-MT interactions polarize very early during neuron morphogenesis. DCX interacts with MTs through two conserved DCX domains that bind between protofilaments and adjacent tubulin dimers, a site that changes conformation during GTP hydrolysis. Consequently and consistent with our previous results, DCX specifically binds straight growth cone MTs and is excluded from the GTP/GDP-Pi cap recognized by EB1. Comparing MT-bound DCX fluorescence to mEmerald-tagged nanocage standards, we measure approximately one hundred DCX molecules per micrometer growth cone MT. DCX is required for i3Neuron growth cone advance in soft microenvironments that mimic the viscoelasticity of brain tissue, and using high resolution traction force microscopy, we find that growth cones produce comparatively small and transient traction forces. Given our finding that DCX stabilizes MTs in the growth cone periphery by inhibiting MT depolymerization, we propose that DCX contributes to growth cone biomechanics and reinforces the growth cone cytoskeleton to counteract actomyosin generated contractile forces in soft physiological environments in which weak and transient adhesion-mediated traction may be insufficient for productive growth cone advance.

Methods
This dataset contains raw data and analysis used to produce figures and conclusions associated with the referenced Current Biology manuscript. Detailed description of the data collection and analysis methodology can be found in the associated manuscript.

Raw microscopy data are provided as 16-bit Nikon ND2 files that are native to Nikon Elements microscopy software or TIFF files. ND2 files can also be opened with ImageJ/Fiji with the Bioformats plugin (Note: metadata may not be imported correctly into ImageJ), or with a freely available Nikon Elements Viewer.

Data analysis on which manuscript figures are based are provided as Microsoft Excel spreadsheets.