Phosphorylation and dephosphorylation enzyme groups regulate sleep and wakefulness in mammals

A research group, Professor Hiroki Ueda, Doctoral Student Yimeng Wang, Doctoral Student Siyu Cao, and Lecturer Koji Ode et al. at the Graduate School of Medicine of the University of Tokyo, has discovered that protein kinase A (PKA)^*2) promotes wakefulness, while protein phosphatase 1 (PP1) and calcineurin^*3), dephosphorylation enzymes, promote sleep in mammals.

Focusing on PKA and dephosphorylation enzymes, the research group created comprehensive gene knockout mice and conducted further experiments inducing the expression of functionally modified enzymes using viral vectors^*4). Consequently, they found that PKA activation decreased sleep duration and delta power, and indicator of sleep needs. On the other hand, PP1 and calcineurin activation conversely increased sleep duration and delta power. In these sleep-wake promoting activities, it is essential that PKA, PP1, and calcineurin act at post-synapses responsible for information transmission between neurons. In addition, they demonstrated that PKA and PP1/calcineurin may work competitively to regulate the daily sleep duration.

This study has revealed that the balance between sleep and wakefulness is regulated by the action of multiple enzymes, which is an important finding when considering how to control sleep duration and sleepiness on the molecular level.

This result was achieved in the Ueda Biological Timing Project, a research area of the Exploratory Research for Advanced Technology (ERATO) by the Japan Science and Technology Agency (JST). Under this project, JST is developing "systems biology for understanding humans" using sleep-wake rhythms as a model system in this project, aiming to understand "biological time" information that runs from molecules to individual humans living in society.

(*1) Protein phosphorylation

After proteins are produced by transcription and translation, their activity may be adjusted by various chemical modifications. Phosphorylation is the most ubiquitous modification found in cells. Enzymes that catalyze the reaction of transferring phosphate groups to proteins using adenosine triphosphate as a substrate are called protein kinases, and enzymes that catalyze the reaction of removing phosphorylation modifications from phosphoproteins are called protein phosphatases.

(*2) Protein kinase A (PKA)

A protein kinase activated by cyclic adenosine monophosphate, an intracellular signaling molecule. PKA consists of a catalytic subunit responsible for kinase activity and a regulatory subunit that inhibits enzyme activity.

(*3) Protein phosphatase 1 (PP1) and calcineurin

Among the protein phosphatases, PP1, PP2A, and calcineurin are expressed at high levels in the brain. This study revealed that PP1 and calcineurin have sleep-regulating functions. These dephosphorylation enzymes consist of a catalytic subunit responsible for dephosphorylation activity and a regulatory subunit controlling the enzyme's subcellular localization and enzyme activity. Unlike other dephosphorylation enzymes, calcineurin is characterized by its activation by calcium.

(*4) Viral vectors

A tool for introducing a gene into a cell, taking advantage of the viral ability of cellular infection adeno-associated virus (AAV)-PHP.eB, a type of viral vector modified from AAV, was used in this study. This viral vector allows for highly efficient gene transfer, especially to the central nervous system.