Multiple sclerosis (MS) is a debilitating disorder that affects 2.8 million people worldwide. MS is characterized by loss of myelin, the structure surrounding nerves necessary for efficient communication between neurons and critical to neurodevelopment, maintenance, and plasticity. The severe symptoms that arise as MS progresses are exacerbated by the loss of oligodendrocytes—the myelin-producing cells—and impaired differentiation of their precursor, oligodendrocyte precursor cells (OPCs). To understand why remyelination fails in MS, we first need to comprehend the mechanisms driving the proliferation and differentiation of myelin-forming precursors. OPCs are the most abundantly mitotic cells in the brain and maintain strict, homeostatic boundaries, indicative of a precision in cell cycle control but also an ability to maintain elaborate tiling throughout the brain despite an ever-changing microenvironment. Oligodendroglia dynamics require the precise timing of transcription factors (TFs) expression, that is essential for efficiently remyelinating lesions. This suggests a large range in dynamic plasticity, a characteristic afforded to cells by the circadian (~24 hour) clock system, a transcriptional-translational negative feedback loop driven by the transcription factors BMAL1 and CLOCK that regulates up to 50% of the mammalian transcriptome. While much is known about the vital role of circadian rhythms in neurons, comparatively little is known about their role in oligodendroglial cells. There is still a significant gap in our knowledge of the genetic mechanisms through which BMAL1 and other TFs control OPC differentiation during myelination. My central hypothesis is that the dynamic nature of myelin-forming glia fostered by BMAL1 and other master transcriptional regulators can be used to enhance myelination. This is based on my data in which Bmal1 loss in OPCs results in transcriptional dysregulation, aberrant OPC dynamics and myelination. These data strongly suggest the necessity of BMAL1 in OPC differentiation and myelination. To test this hypothesis, my approach will be to: 1) Characterize the role of BMAL1—the only single clock factor necessary for circadian rhythmicity—in OPC transcriptional regulation during neurodevelopment through single-cell RNAseq and CUT&Tag using our established conditional clock knockout that lacks Bmal1 in OPCs; 2) Evaluate the recovery of the differentiation potential of OPCs that lack Bmal1 by modifying signaling pathways that act downstream of BMAL1; 3) Study the transcriptional control of regulators of human OPC differentiation through a CRISPR screen in human OPCs to discover enhancers. My goal is to identify new regulatory mechanisms of OPC differentiation into myelin-forming cells, starting with the role of BMAL1 in OPC dynamics, and continuing with genomic elements that control OPC differentiation. With the K99/R00 Award, I will obtain the training to prepare me for a lifelong independent research career in...