Single-cell transcriptomics and analysis for new molecular regulators of sleep

The molecular mechanisms governing sleep are largely unknown. Here, we used a combination of single-cell RNA sequencing to interrogate the molecular and functional underpinnings of sleep. Different cell types in three important brain regions for sleep (brainstem, cortex and hypothalamus) had a similar transcriptional response to sleep need, with a large proportion of cells changing during recovery sleep. In contrast, sleep deprivation regulated expression of different functions in each brain region. This includes antigen processing, synaptic transmission and cellular metabolism in brainstem, cortex and hypothalamus, respectively. Increased sleep need enhances expression of the transcription factors Sox2, Mafb, and Zic1 in brainstem; Hlf, Cebpb and Sox9 in cortex, and Atf3, Fosb and Mef2c in hypothalamus. In turn, these transcription factors regulate downstream gene expression during sleep deprivation and recovery. In cortex, we also interrogated the proteome of two major cell types: neurons and astrocytes. We found surprising functional overlap of proteins that mediate vesicle and neurotransmitter transport in both cell types. In contrast, other functions were specific to each cell type.