Deconstructing in vivo glia-to-neuron conversion
Final Report Abstract
Direct lineage reprogramming of cell identity in the nervous system offers the prospect of remodeling diseased brain circuits. In recent years, substantial evidence has emerged, supporting the possibility of converting astrocytes and oligodendrocyte precursor cells (OPCs) into functional induced neurons in vivo. Yet, the process by which glial cells give up their original identity and adopt a neuronal fate remains by large enigmatic. In my work, I addressed questions regarding how specific neurogenic reprogramming factors, such as Ascl1 and its phospho-deficient counterpart (SA6, in which six serine residues have been altered to alanine) differentially remodel gene expression programs as glial cells convert into neurons in the mouse cortex. I also explored the contribution of glial cell heterogeneity to the variability in reprogramming success and aimed to identify potential candidate molecules that act as roadblocks for successful reprogramming. To achieve this, I injected MMLV-retroviruses encoding either Ascl1 or SA6 in combination with the cell death regulator Bcl2 and interneuronal transcription factor Dlx2 into the somatosensory cortex of postnatal mice, transducing primarily proliferating astrocytes and OPCs. Subsequently, I performed single-cell RNA sequencing of cells undergoing conversion in vivo and investigated the molecular trajectories to uncover the transcriptomic changes driving this process. Bioinformatical analysis of these cells reveal that Ascl1 and SA6 both induce neurogenic programmes in astrocytes; however, they both give rise to distinct neuronal cell populations characterised by specific transcription factors, which may orchestrate the respective reprogramming outcomes. Interestingly, Ascl1 did not seem to initiate neurogenic programs in OPCs, suggesting that cell type-specific molecular properties play a crucial role in reprogramming success. On the other hand, both astrocytes and OPCs initiated neurogenesis when overexpressing SA6, resulting in distinct neuronal cell fates despite expressing the same transcription factor. These findings provide valuable insights into the molecular underpinnings of glia-to-neuron conversion in vivo. We demonstrate that the phosphorylation state of the proneuronal factor Ascl1 plays a pivotal role in initiating differential gene expression programs and determining differential neuronal cell fates in cortical astrocytes and OPCs. Overall, this study may offer essential cues to further manipulate these reprogramming pathways and refine the efficacy of cell fate conversion.
Publications
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Gatekeeping astrocyte identity. eLife, 11.
Cooper, Alexis & Berninger, Benedikt
