Genetic and phenotypic outputs of the circadian clock in Per3 knock-out mice and humanised Per3 knock-in mice

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Context The circadian clock enables animals to synchronise their biology with the light/dark cycle. This allows animals to anticipate dawn and to prepare the body to emerge from sleep into alertness, a big evolutionary advantage with respect to prey detection or predator avoidance. In mammals, the master clock is in the suprachiasmatic nuclei (SCN) in the brain, which receive timekeeping signals from light detection in the eyes. The SCN sends out signals to subservient clocks in other organs and tissues to keep them synchronised. The oscillations of the circadian clock are created by specific clock genes. These are expressed into proteins, which interact in positive and negative feedback loops. In one central negative feedback loop, three groups of clock proteins called PER, CRY, and CK I combine into a 'timesome' in the cell's cytoplasm, and enter the nucleus together. Inside the nucleus, they inhibit the expression of the Per and Cry genes, preventing the reformation of PER and CRY protein. This eventually removes the feedback, and the cycle starts again. The time taken to complete this cycle determines the period of the clock, which in humans is just over 24 hours. In the cytoplasm, phosphate groups are added to the PER and CRY proteins by CK I (phosphorylation). This may cause the timesome to enter the nucleus faster, shortening the period of the clock. Our research has shown that human PER3 has a region to which many phosphate groups could potentially be added through phosphorylation. But humans have two versions of this region, one longer and one shorter. Because we carry two versions of each chromosome (apart from the sex chromosome), one from each parent, people may have two copies of one or one of each. The long and short versions have different numbers of phosphorylation sites. We have shown that the long form is associated with morning people, while the short form is associated with evening people and also a condition called delayed sleep phase syndrome (DSPS). Aims & Objectives Although mice have been used as a model animal for mammalian circadian studies, the mouse PER3 does not contain this phosphorylation region. We propose to investigate further the role of PER3 in the circadian clock. This will be done by studying mice in which the Per3 gene has been disrupted (so there is no PER3 protein) and also mice whose Per3 gene has been modified to resemble the two forms of the human gene. A comparison will then be made of how different genes are turned on or off in mice with a normal or humanised Per3 gene, and in knock-out mice which lack the Per3 gene altogether. The project will also investigate how this affects the output of the SCN through circadian behaviour and sleep. Potential Applications & Benefits This research will help us to understand more fully the role of PER3 in the circadian clock and how the phosphorylation can affect circadian function at the genetic, neural and behavioural levels. This should help us to understand why people have differences in morning and evening preference, and even why some of them suffer from sleep disorders such as DSPS. An association has also been found between human PER3 length and incidence of breast cancer. The results from the genetic work may identify genes that are involved in this association, and the mice with the human PER3 region added will provide a model system for further research in this area.