Distinct differentiation trajectories leave lasting impacts on gene regulation and function of V2a neurons [Bulk RNA-seq]

Single nucleus multiomic analysis showed that different V2a lineages were enriched for different transcription factor motifs and differentially expressed hundreds of genes relevant to neuron function, such as axon growth and calcium handling. Attempting to ‘skip’ developmental patterning by induced transcription factor expression yielded a population unlike either developmentally relevant population, highlighting the importance of following developmental steps in establishing cell identities in vitro.

Lastly, we employed the CellOracle tool and lentiviral knockdown to perturb lineage-specific gene regulatory networks and identify CREB5 and TCF7L2 as regulators unique to a spinal-like lineage. Overall, this work highlights how small differences in differentiation protocols can influence mature cell fate and uncovers new regulators of neural diversity along the anterior-posterior axis of the central nervous system.