Report
Teaser, summary, work performed and final results
Periodic Reporting for period 2 - ProNeurons (Transcription Factor-mediated Neuronal Cell Fate Programming in Human Stem Cells)
Teaser
The use of human induced pluripotent stem cells (hiPSCs) facilitates studying the genesis of human cell types in an ethically approved setting. However, exploiting the full potency of stem cells is only possible with very few differentiated cell types. In particular, the...
Summary
The use of human induced pluripotent stem cells (hiPSCs) facilitates studying the genesis of human cell types in an ethically approved setting. However, exploiting the full potency of stem cells is only possible with very few differentiated cell types. In particular, the generation of neurons is in its infancy: of the many neuronal types present in the brain, only very few types have been generated in vitro. So far, neuronal differentiation protocols are multifaceted and tailored to individual cell types. The molecular events that occur during reprogramming remain enigmatic. Hence, we cannot confer these protocols easily on producing different neurons of interest. Therefore, we induce transcription factors as differentiation control buttons in hiPSCs and explore their potency to trigger in vitro neurogenesis systematically. Generating certain human neuronal cell types in high quality and quantity is of high interest for cell replacement therapies to repair brain function and vision. Furthermore, hiPSC-derived neurons represent potent model systems to study human pathologies but also basic cellular functions in a dish.
Work performed
To this end, we have composed and applied a human transcription factor library to conditional fluorescent hiPSC reporter lines, facilitating high-throughput isolation and analysis of induced neurons. We have already screened 2000 single cells and have identified novel neurogenic transcription factors, which we are currently validating. Towards generating a neuronal cell type of interest, we have identified the transcription factor cocktail by an unbiased screening approach to program human photoreceptors from hiPSCs. These cells can be applied to cell transplantation experiments in retinal degeneration diseases. Towards revealing the underlying gene regulatory networks using RNA sequencing over the entire differentiation period, we have successfully applied novel systems biological approaches. These computational methods will be useful to further characterize induced neurons but also to control their neurogenesis to specific neuronal cell types.
Final results
By transcription factor overexpression, we can simplify the induction of neurogenesis in hiPSCs regarding purity, speed and homogeneity. By revealing the biological rules of in vitro neurogenesis, our approach conceptually paves the way for targeted “forward†programming of hiPSCs to neurons.