Seongjin Seo, Jong‐Won Lim, Dhananjay Yellajoshyula, Li‐Wei Chang, Kristen L Kroll

Proneural basic helix-loop-helix proteins are key regulators of neurogenesis but their 'proneural' function is not well understood, partly because primary targets have not been systematically defined. Here, we identified direct transcriptional targets of the bHLH proteins Neurogenin and NeuroD and found that primary roles of these transcription factors are to induce regulators of transcription, signal transduction, and cytoskeletal rearrangement for neuronal differentiation and migration. We determined targets induced in both Xenopus and mouse, which represent evolutionarily conserved core mediators of Neurogenin and NeuroD activities. We defined consensus sequences for Neurogenin and NeuroD binding and identified responsive enhancers in seven shared target genes. These enhancers commonly contained clustered, conserved consensus-binding sites and drove neural-restricted transgene expression in Xenopus embryos. We then used this enhancer signature in a genome-wide computational approach to predict additional Neurogenin/NeuroD target genes involved in neurogenesis. Taken together, these data demonstrate that Neurogenin and NeuroD preferentially recognize neurogenesis-related targets through an enhancer signature of clustered consensus-binding sites and regulate neurogenesis by activating a core set of transcription factors, which build a robust network controlling neurogenesis.