Circadian clocks are self-sustaining molecular oscillators that regulate daily changes in behavioral activity and physiology. For functional reliability and precision, the frequency of these molecular oscillations must be stable at different ambient temperatures, known as “temperature compensation”. Despite being an intrinsic property of all circadian clocks, this phenomenon is not well understood at the molecular level.
A team of biologists led by Prof. Ralf Stanewsky of the University of Münster, and in collaboration with teams from Dalhousie University in Canada and the University of Mainz in Germany, have now found a vital piece in the puzzle that may provide an answer to this demand offers.
The scientists found a point mutation in the fruit fly Drosophila melanogaster that causes circadian clock periods to lengthen in response to temperature. It is found in a ‘period’, a central ‘clock gene’. Flies with this perI530A mutation show a typical sleep-wake cycle of 24 hours at 18 degrees Celsius. The indoor clock, on the other hand, works five hours slower and lasts for 29 hours at 29 degrees Celsius. The activity or expression of the menstrual gene in the clock neurons of the brain is similarly affected by the prolongation of menstruation.
Usually, the protein in question (PERIOD) undergoes a slow chemical change over 24 hours; more precisely, it is phosphorylated. It is broken down when phosphorylation is at its peak. When fruit flies are active between 18 and 29 degrees Celsius, this mechanism is usually the same. The researchers showed that at 18 degrees Celsius, phosphorylation normally occurs in the perI530A mutant, but decreases as the temperature increases. The “PERIOD” protein is then stabilized at higher temperatures.
The researchers used fruit fly mutants that had been modified in the period gene they had created using modern molecular genetics techniques (CRISPR/Cas9 mutagenesis and homologous recombination). It was then determined whether the sleep-wake cycles of the animals and, consequently, their running activity varied with ambient temperature.
The so-called nuclear export signal (NES), which also occurs in this form in mammalian menstrual genes and is involved in moving the PERIOD proteins out of the nucleus, is affected by the mutation the team analyzed. This export from the cell nucleus had no previously known biological purpose. The latest work shows that, again, only at higher temperatures does the mutation result in prolonged retention of the PERIOD protein in the nucleus of central clock neurons.
Ralf Stanewsky said: “We therefore assume that the export of the protein from the cell nucleus plays an important role in temperature compensation – at least as far as the fruit fly is concerned.”
- Astrid Giesecke, Peter S. Johnstone, Angelique Lamaze, Johannes Landskron, Ezgi Atay, Ko-Fan Chen, Eva Wolf, Deniz Top, and Ralf Stanewsky (2022): A new period mutation, implicating nuclear export in temperature compensation of the Drosophila circadian clock. Current Biology; DOI: 10.1016/j.cub.2022.12.011