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PDF, English Download (17MB) | Lizenz: Investigating the Molecular Basis of Adult Neurogenesis: Single-Cell Multi-Omics Unveils DNA Methylation as a Key Regulator of Astrocyte Stemness by Kremer, Lukas Peter Maria underlies the terms of Creative Commons Attribution - ShareAlike 4.0

Abstract

The ventricular-subventricular zone (vSVZ) of adult mammalian brains harbors specialized astrocytes, called adult neural stem cells (NSCs), which are capable of generating both neurons and glial cells. In contrast, common parenchymal astrocytes perform a wide range of structural, metabolic, homeostatic and neurosupportive functions. Despite these distinct functions, studies employing immunostaining and single-cell RNA-sequencing have demonstrated that NSCs and astrocytes largely express the same set of genes, which raises the question how stem cell function is molecularly encoded. To address this question, I analyzed a single-cell triple-omic data set that contains information on gene expression, chromatin accessibility and DNA methylation for hundreds of cells of the adult NSC lineage, as well as common parenchymal astrocytes.

To enable this analysis, I developed "scbs", a Python software for the analysis of single-cell methylation data. I devised and implemented two major improvements over the current state of the art analysis workflow: First, instead of segmenting the genome into fixed intervals, I devised an approach to scan the entire genome for informative regions called variably methylated regions (VMRs). Second, instead of simply averaging methylation values within these tiles, I devised a more robust measure of DNA methylation.

By making use of these new methods, I demonstrated that adult NSCs possess a unique DNA methylation profile that is not found in common parenchymal astrocytes. This NSC methylome is characterized by hypomethylation of genes required for neurogenesis. I propose that this feature contributes to the neurogenic capabilities of NSCs by enabling the transcriptional activation of these genes. In contrast, common parenchymal astrocytes are locked in their current astrocyte fate by DNA methylation. To test this hypothesis, I analyzed single-cell multi-omic data from mice that were subjected to ischemia, as ischemia is known to induce a neurogenic response in common parenchymal astrocytes and NSCs. My analysis suggests that this gain of neurogenic capabilities is accompanied by gain of an NSC methylome, which supports the idea that a specific DNA methylome is required for stem cell function. Overall, my results demonstrate that DNA methylation is dynamic even in adult tissues and not just in embryonic development, and furthermore unveils DNA methylation as a crucial factor that constrains or enables alternative cellular fates.