Sleep the shutting of the eye: sleep duration and genes

Sleep duration and genes

Sleep is nearly ubiquitous throughout the animal kingdom and in general a great deal of variation exists in sleep duration among different animal phyla, with animals such as the African Elephant sleeping only 3–4 hrs per day while animals like armadillo sleeping over 18 hrs per day.

For example, two thirds of a cat’s life is spent asleep, a giraffe only needs 1.9 hrs of sleep per day, whereas a brown bat needs 19.9 hrs per day: The origins and evolution of sleep.

Even among humans, sleep times vary widely, ranging from less than 5 hrs to 10 hrs or more. This large difference in sleep duration among humans, suggests that an existing genetic variation among individuals affects sleep.

In fact, Ying-Hui Fu PhD  —  professor of neurology and a member of the UCSF Weill Institute for Neurosciences  —  led the research teams that discovered three of the “short sleep genes”, genes that allow people to sleep fewer than six hours per night and yet be able to function well throughout the next day.

In 2009, her team discovered that people who had inherited a particular mutation in a gene called DEC2  —  transcriptional repressor for the expression of orexin  —  averaged only 6.25 hrs of sleep per night, while participants lacking the mutation averaged 8.06 hrs The transcriptional repressor DEC2 regulates sleep length in mammals.

DEC2 — that was dubbed the “Thatcher gene” after UK Prime Minister Margaret Thatcher who famously lived on four hours of sleep per night — is a transcriptional repressor for orexin expression, and this particular DEC2 mutant discovered exerts less repressor activity than wild type-DEC2, resulting in increased orexin expression.

Orexin (from orexis meaning “appetite” in Greek), also known as hypocretin — because it is produced in the hypothalamus and bears a weak resemblance to peptide secretin — is a neuropeptide that regulates arousal, wakefulness and appetite. The most common form of narcolepsy (cataplexy), is caused by a lack of orexin in the brain due to destruction of the cells that produce it.

The discovery of orexin  —  the hypothalamic neuropeptide involved in the maintenance of wakefulness  —  indicated also the role of the orexinergic and other neural pathways in the regulation of sleep/wakefulness, so drug development followed and as a result two new drugs were FDA approved for treating insomnia.

In particular, the orexin receptor antagonist suvorexant  —  which specifically blocks the endogenous waking system  —  has been approved as a new drug (Belsomra made by Merck & Co) to treat insomnia (linked to the occurrence of specific variants on chromosome 7 close to AUTS2 gene, an activator of transcription and developmental regulator). And dayvigo (lemborexant), developed in-house by Japanese drug company Eisai, was also FDA approved for the treatment of insomnia. Lemborexant binds to wake-promoting orexin receptors OX1R and OX2R and acts as a competitive antagonist with stronger inhibition effect to OX2R, suppresing the wake drive.

Moreover, a 2014 study at the University of Pennsylvania Perelman School of Medicine found that a novel BHLHE41 variant is also associated with short sleep and resistance to sleep deprivation in humans. BHLHE41 (basic helix-loophelix family member e41) is the DEC2 gene or the “Thatcher gene”.

But all of these DEC2 mutations discovered are rare, so while they helped explain some natural short sleepers, they couldn’t account for all of them.

In fact, in 2019 Ying-Hui Fu’s team discovered also another single-letter mutation in a gene known as ADRB1  —  ß1-adrenergic receptor, a G-protein coupled receptor  —  that like the mutation in DEC2 was associated with natural short sleep: A Rare Mutation of β1-Adrenergic Receptor Affects Sleep/Wake Behaviours.

Her team found that ADRB1 receptor is highly expressed in the dorsal pons —  or pontine tegmentum is located within the brainstem  —  and that these ADRB1+ neurons are active during rapid eye movement (REM) sleep and wakefulness. So, activating these neurons can lead to wakefulness, and the activity of these neurons is affected by the single-letter mutation mutation in ADRB1.

The third short sleep gene that prevents also memory loss and discovered by the same team is Npsr1 (G protein–coupled Neuropeptide S receptor 1), which encodes a signaling protein that sits on the surface of neurons.

A rare missense mutation (occurring in fewer than 1 in 4 million people) in the Npsr1, led the team to associate Npsr1 with a natural short sleep phenotype in humans: Mutant neuropeptide S receptor reduces sleep duration with preserved memory consolidation.

Npsr1 was formerly an orphan receptor, GPR154, until the discovery of neuropeptide S as the endogenous ligand (NPS). The very rare NPSR mutation Y206H  —  which makes the receptor more sensitive to NPS  —  it has similar effects in transgenic mice, making them resistant to memory impairment caused by lack of sleep. Thus suggesting that the NPS/NPSR1 pathway determines sleep duration and links sleep homeostasis and memory consolidation.

Dr. Fu’s goal now is to find 10 more short-sleeping genes and to learn if there are any connections among them.

But achieving a full understanding of sleep function requires a detailed characterisation of the genetic, molecular and neuronal properties associated with sleep and wakefulness.

For this reason, over the past few decades, apart mammalian, also non-mammalian genetic models including the nematode worm Caenorhabditis elegans, the fruit fly Drosophila melanogaster and the zebrafish Danio rerio have also been particularly advantageous in elucidating genetic and molecular components underlying sleep. Let's see some examples now:

Drosophila melanogaster

The invertebrate whose sleep has been most intensively studied is the fruit fly Drosophila melanogaster. The fruit fly’s central nervous system has over 200,000 neurons (the human brain has billions of neurons) and does not have anatomic structures that clearly correspond to their vertebrate counterparts. But the fly genome has about 14,000 genes, many of which are highly conserved between flies and humans at the level of sequence and even function. In other words is easier to study sleep with Drosophila and yes, even the humble fruit fly needs sleep.

The earliest sleep screen performed in Drosophila (Reduced sleep in Drosophila Shaker mutants) gave about 9,000 mutagenized lines that were phenotyped for extreme short sleep time. Flies carrying loss-of-function mutations for Shaker  —  a gene encoding a voltage-dependent potassium channel  —  were found to sleep exceptionally little. In Drosophila, the shaker gene is located on the X chromosome and the closest human homolog is KCNA3.

Surprisingly, after screening several thousand additional mutagenized lines, a later Drosophila sleep screen identified the gene Sleepless (SSS), which proved to be a regulator of Shaker: Identification of SLEEPLESS, a sleep-promoting factor.

SSS encodes a brain-enriched, glycosyl-phosphatidylinositol (GPI)-anchored membrane protein. SSS and Shaker exhibit similar expression patterns in the brain and specifically affect each other’s expression levels. Together, the two studies suggested neuronal membrane excitability as a core feature of homeostatic sleep drive.

Loss of the Sleepless protein causes an extreme (>80%) reduction in sleep. Genetic and molecular analyses revealed that quiver, a mutation that impairs Shaker-dependent K+ current, is an allele of sleepless. Consistent with this finding, Shaker protein level is reduced in sleepless mutants. It is proposed that Sleepless is a signaling molecule that connects sleep drive to lowered membrane excitability.

By phenotyping about 12,000 fly lines for sleep duration, Nemuri (Japanese for “sleep,” abbreviated nur, encoding an uncharacterized antimicrobial peptide) was also identified: A sleep-inducing gene, nemuri, links sleep and immune function in Drosophila.

The Nemuri protein is an antimicrobial peptide that can be secreted ectopically to drive prolonged sleep and to promote survival after infection. Loss of nemuri increased arousability during daily sleep and attenuated the acute increase in sleep induced by sleep deprivation or bacterial infection.

Caenorhabditis elegans

Another organism whose sleep has been also intensively studied is the transparent nematode Caenorhabditis elegans. The ~100 MB genome of C. elegans has ~20,000 protein-coding genes many of which are required for the function of the nervous system, composed of 302 neurons in the adult hermaphrodite and of 383 neurons in the adult male.

When the C. elegans proteome was used as an alignment template to assist in novel human gene identification from human nucleotide databases, among the available 18,452 C. elegans protein sequences, it came out that at least 83% (15,344 sequences) of C. elegans proteome has human homologous genes, with 7,954 records of C. elegans proteins matching known human gene transcripts. Only 11% or less of C. elegans proteome contains nematode-specific genes.

In C. elegans, when forward genetic screens for sleep was performed, results indicated that upon stress the release of the neuropeptide Flp-13 (FMRFamide-like peptide-13) and its receptor Dmsr-1 — a G protein–coupled receptor — can mediate the sleep-promoting effect: Genome-Wide Screen for Genes Involved in Caenorhabditis elegans Developmentally Timed Sleep.

The G-protein that is coupled to Dmsr-1 may be the G i/o alpha subunit Goa-1, which was identified in another mutagenesis screen for genes regulating developmentally timed sleep, along with Gpb-2, encoding another G-protein subunit. Together, these results confirm the importance of G-protein signaling pathways in worm sleep.

Danio rerio

The zebrafish (Danio rerio)  —  a freshwater fish belonging to the minnow family — is another organism whose sleep has been also intensively studied. The zebrafish genome is strikingly similar to humans. According to a paper published in Nature, 70% of protein-coding human genes are related to genes found in the zebrafish (Danio rerio), and 84% of genes known to be associated with human disease have a zebrafish counterpart. As a vertebrate, the zebrafish has the same major organs and tissues as humans.

In a zebrafish-overexpression screen for secreted proteins, the neuropeptide Neuromedin U (Nmu) was identified to regulate sleep-wake behavior: A Zebrafish Genetic Screen Identifies Neuromedin U as a Regulator of Sleep/Wake States.

Interestingly, anatomical and functional analyses found that Nmu-induced arousal is mediated by corticotropin-releasing hormone (CRH) signaling in the brainstem.

Neuromedin U is a highly conserved neuropeptide present in many species, and found also in the brain of humans, and is a multifunctional neuropeptide with pleiotropic effects, including the mediation of intestinal peristalsis and the modulation of the sense of satiety, body weight, circadian oscillation, bone formation, insulin production, cancer development, energy balance and metabolism.

Also, overexpression of the neuropeptide Neuropeptide Y (NPY) was identified in this screen. Neuropeptide Y is one of the most abundant peptides present in the mammalian central nervous system, and it is secreted alongside other neurotransmitters such as GABA and glutamate. In humans, NPY was found to have hypnotic properties, possibly acting as a physiological antagonist of corticotropin-releasing hormone (CRH).

Of course, the list of these genes goes on!

Despite its essential role in maintaining optimal performance, health and well-being, and despite an increasing number of studies and mining large database for sleep genes, the genetic mechanisms underlying sleep remain poorly understood and coherent picture on “sleep” genes has yet to emerge.

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