Molecular Sciences I.
Orsolya Sárkány, Department of Physiology, Semmelweis University, Budapest
Introduction: The circadian rhythm is a nearly 24-hour time-measuring mechanism driven by molecular oscillators that helps organisms adapt to daily environmental fluctuations. Endogenous time measuring is based on an evolutionarily conserved transcription-translation feedback loop (TTFL) which is present in almost all eukaryotes including the Neurospora crassa filamentous fungus which is widely used as a model system in the circadian research.
The circadian clock helps maintain and optimize homeostasis by coordinating various metabolic processes. The metabolic compensation of the clock allows the oscillators to operate with nearly constant period at different nutrient levels.
The RAS pathway plays a key role in different cellular events, including cell proliferation, differentiation and nutrient sensing. According to literature data there is a close connection between the RAS pathway and the circadian rhythm, however, the underlying mechanism is still unclear.
Aims: Mapping the relationship between the RAS pathway and the circadian rhythm by characterizing the role of a RasGEF protein in the metabolic compensation of the circadian clock.
Method: Since the conidiation rhythm of Neurospora is a well-examined output of the circadian rhythm, the phase and period length of the clock were measured using a race tube test. A rescue strain was generated by transforming the rasgef-ko strain with a construct encoding a flag-tagged version of RasGEF. The RNA and protein levels of the main clock components and rasgef were analyzed using qRT-PCR and Western blot.
Results and Conclusion: In the rasgef-ko strain the phase of conidiation is delayed and under constant conditions the period is longer compared to the wild type strain. At the molecular level, RasGEF affects the starvation-dependent reorganization of the oscillator. Expression of the rasgef RNA shows circadian oscillation.
We propose that rasgef is a novel clock-controlled gene that plays an important role in the metabolic regulation of the circadian clock.
Funding: SE250+ and FE0301ELET TKP-EGA-25