| Přispěvatelé: |
Taylor, L, Ashton, A, George, C, Di Pretoro, S, Varga, N, Foster, A, Jagannath, A |
| Popis: |
Disruption to normal circadian rhythms and frequent periods of sleep deprivation (SD), are endemic in today’s 24 hour society; therefore understanding the fundamental biology underlying what happens in the suprachiasmatic nuclei (SCN) during entrainment, and other specific brain regions following SD has never been more important. Current understanding of entrainment and SD within the SCN at the molecular level is still very limited, despite SD already having been shown to have a profound effect on brain function. The published Assay for Transposase-Accessible Chromatin sequencing (ATAC-seq) protocol was successfully modified to allow the technique to be effective with small amounts of tissue collected from murine SCN tissue. ATAC-seq was effectively used to investigate chromatin accessibility within cells of the SCN following light pulsing at different times of the day, and following SD in the SCN, dentate gyrus, and cortex. Data from ATAC-seq light pulse experiments was compared to published gene expression data following light pulsing and determined changes in chromatin accessibility also affect the transcriptome. RNA-seq was used to investigate gene expression following SD, and gain insights into how chromatin affects the transcriptome following SD. Results obtained indicate, chromatin accessibility is highly dynamic within all brain regions investigated. ATAC-seq revealed chromatin accessibility changes in the SCN following light pulsing, but only at certain times of the day, and discovered chromatin modification across all brain regions following SD. Several genes identified close to differential chromatin are already known to play an important role within the molecular clock, and are recognized to be important for entrainment and SD. In addition several novel genes with differential chromatin nearby have been identified that also likely have an important role during entrainment and following SD. Several areas of differential chromatin were investigated using a luciferase reporter assay, and results obtained indicate the areas of chromatin are able to strongly drive gene expression, and respond to treatment using forskolin and horse serum, both known to have an effect on the molecular clock. |
