Neuronal Basic Helix–Loop–Helix Proteins Neurod2/6 Regulate Cortical Commissure Formation before Midline Interactions

Establishment of long-range fiber tracts by neocortical projection neuronsis fundamental for higher brain functions. The molecular control of axon tract formation, however, is still poorly understood. Here, we have identified basic helix–loop–helix(bHLH) transcription factors Neurod2and Neurod6 as key regulators of fasciculationand targeted axogenesis in the mouse neocortex. In Neurod2/6 double-mutant mice, callosalaxons lack expression of the cell adhesion moleculeContactin2, defasciculate in the subventricular zone, and fail to grow toward the midline without forming Probst bundles. Instead, mutant axons overexpress Robo1and follow random trajectories into the ipsilateral cortex. In contrast to long-range axogenesis, generation and maintenance of pyramidal neurons and initial axon outgrowth are grossly normal, suggesting that these processes are under distinct transcriptional control. Our findings define a new stage in corpus callosumdevelopment and demonstrate that neocortical projection neuronsrequire transcriptional specification by neuronal bHLH proteins to execute an intrinsic program of remote connectivity.