Shank3 Modulates Sleep and Expression of Circadian Transcription Factors

Background: Autism Spectrum Disorder (ASD) is the most prevalent neurodevelopmental disorder in the United States and often co-represents with sleep problems. Sleep problems in ASD predict the severity of ASD core diagnostic symptoms and have a considerable impact on the quality of life of caregivers. Little is known, however, about the underlying molecular mechanisms. We investigated the role of Shank3, a high confidence ASD gene candidate, in sleep architecture and regulation.

Objectives: To better understand the mechanisms underlying sleep problems in ASD, we use an animal model that can closely recapitulate sleep phenotypes observed in clinical populations.

Methods: Sleep questionnaire data from Phelan-McDermid syndrome (PMS) patients with Shank3 mutations was obtained through the PMSIR foundation. To assess the sleep-wake behavior in Shank3^ΔC mice, we surgically implanted mutant mice and their wild type littermates with electroencephalographic (EEG) and electromyographic (EMG) electrodes. The EEG/EMG data was used to determine sleep architecture. Genome wide gene expression was performed to assess the differences in gene expression sleep deprivation induces in the prefrontal cortex of Shank3C mice and wild type littermates. Continuous running wheel behavior of Shank3^ΔC mice and wild type mice was monitored to assess circadian rhythms in constant darkness.

Results: Through examination of sleep in PMS patients with Shank3 mutations we show that PMS patients have trouble falling and staying asleep similar to what is observed in the general ASD population. We also show that mice lacking exon 21 of Shank3 have problems falling asleep even when sleepy and sleep less deeply. Using RNA-seq we show that sleep deprivation increases the differences in gene expression between mutants and wild types, downregulating circadian transcription factors Per3, Dec2, Hlf, Tef, and Reverbα. Shank3 mutants also have trouble regulating wheel-running activity in constant darkness. Overall our study shows that Shank3 is an important modulator of sleep and clock gene expressions.

Conclusions: We show that both PMS patients and Shank3^ΔC mutant mice have trouble falling asleep. Shank3^ΔC mice have problems falling asleep when sleep pressure is high at the end of their active period or following sleep deprivation. They also sleep less deeply, but show no gross abnormalities in sleep homeostasis or circadian rhythms which suggests that the sleep deficit is due to difficulty with sleep onset. Our molecular studies show that sleep deprivation increases differences in gene expression between Shank3^ΔC mice and wild type mice, with differences pointing to the downregulation of circadian transcription factors. Shank3^ΔC mice do not show a disruption in circadian rhythmicity but a large reduction in wheel-running activity in response to constant darkness. This suggests the mutant sleep phenotype involves clock gene function outside their central time-keeping role.