Clockwork orange beats time for the body

A RIKEN-led team of researchers from Japan and the US has used an innovative combination of genome survey techniques in live Drosophila fruit flies to reveal a previously unknown master gene involved in setting circadian rhythms. It is the tenth of a series of genes which generate proteins that interact in complex interlocking feedback loops to measure the 24-hour day.

This network of genes ensures that the rhythms of vertebrate animals—sleep and wakefulness, changes in body temperature and blood pressure, the secretion of hormones and regulation of fertility—are attuned to daily and seasonal cycles. The importance of the network’s role is demonstrated not only by the fact that mutations in clock genes have been linked to cancer and obesity, but also that they have been highly conserved during evolution and are similar in organisms from fruit flies to mammals. The regulation of the network is intricate and complex because, in addition to the simple measurement of time, it has to respond to changing daily patterns of light and dark and seasonal patterns of temperature and the environment.

In humans, such common problems as jet lag and lack of alertness of shiftworkers arise when the body’s circadian rhythms are not properly adjusted to the external environment. Permanent disruption of the body’s clock can lead to much more serious disorders, such as delayed sleep phase syndrome, and has also been implicated in the depressive mental illness known as seasonal affective disorder (SAD). Work on clock genes could well have relevance to treating these conditions.

In a recent issue of the journal Genes & Development ¹, the research group—from RIKEN’s Center for Developmental Biology in Kobe, Kyushu University, Japan’s National Institute of Genetics and two universities in Texas—detailed how it found the new master gene. The gene exhibits rhythmic daily activity and codes for a protein that represses other clock genes through the regulatory DNA sequence known as E-box. Because it carries a characteristic DNA domain known as ORANGE, the research team named the gene clockwork orange (cwo ) referring to the well-known Anthony Burgess novel and Stanley Kubrick film.

The researchers uncovered cwo using a novel partnering of micro-array technology that shows which genes are cyclically switched on, with RNA interference techniques, whereby individual genes can be switched off at will. In previous work, using micro-arrays they had followed the daily activity pattern of all genes in the Drosophila genome in cells of the head region of the fruit fly, where the circadian pacemaker is located. They found about 200 genes showed a rhythmic activity pattern over the daily cycle both under normal conditions of light and dark, and also in constant darkness.

In the recently reported study, using RNA interference techniques the researchers then switched off each of about 130 genes which showed such regular daily cycles of activity. They were looking for dramatic disturbances of overall rhythmic behavior which, they hypothesized, would only happen when they disrupted the core genes involved in setting the body’s clock.

In addition to genes already known to be an integral part of the circadian mechanism in Drosophila, the researchers found five other candidates for such clock genes. Of these, cwo was the gene with the most pronounced and stable impact.