BMAL1 loss in oligodendroglia contributes to abnormal myelination and sleep
Myelination depends on maintenance of oligodendrocytes that arise from oligodendrocyte precursor cells (OPCs). We show that OPC-specific proliferation, morphology, and BMAL1 are time-of-day dependent. Knock out of Bmal1 in OPCs during development disrupts expression of genes associated with circadian rhythms, proliferation, density, morphology, and migration, leading to changes in OPC dynamics in a spatio-temporal manner. Furthermore, these deficits translate into thinner myelin, dysregulated cognitive and motor function, and increased sleep fragmentation.  OPC-specificBmal1loss in adulthood does not alter OPC density at baseline but impairs remyelination of a demyelinated lesion driven by changes in OPC morphology and migration. Lastly, we show sleep fragmentation is associated with increased prevalence of the demyelinating disorder multiple sclerosis (MS), suggesting a link between MS and sleep that requires further investigation. These findings have broad mechanistic and therapeutic implications for brain disorders that include both myelin and sleep phenotypes. Overall design: We developed a model to specifically knock out the Bmal1 DNA-binding domain in OPCs during embryonic development by crossing clock gene knock out (Bmal1fl/fl) and cell type-specific Cre driver (NG2::Cre) mice to generate a Bmal1 transcriptional hypomorph that significantly decreased BMAL1 levels specifically in OPCs (NG2:Cre+;Bmal1fl/fl or OPC-Bmal1-KO). To test the role of BMAL1 as a transcriptional regulator in OPCs, we isolated OPCs from P6-7 OPC-Bmal1-WT and OPC-Bmal1-KO mice at 6-hr intervals for 24 hrs. OPCs were isolated at 4 different zeitgeber times: ZT0 (lights on), ZT6, ZT12 (lights off), and ZT18. For each biological replicate, 2 mice from both sexes were pooled to obtain around one million OPCs. OPCs were purified using immunopanning through 2 negative selection plates for microglia and endothelial cells with BSL1 (Vector Laboratories L1100) and one positive selection plate with an anti-PDGFRa antibody (rat anti-mouse CD140A, BD Pharmingen 558774). OPCs were immediately lysed using QIAzol Lysis Reagent. RNA extraction was performed following manufacturer's instructions and RNA purity, integrity and concentration were assessed using a Bioanalyzer. Library prep and Bulk RNA sequencing was performed using Novogene Co., Ltd. sequencing service.