TIMELESS-dependent positive and negative autoregulation in the Drosophila circadian clock

The timeless protein (TIM) is a central component of the circadian pacemake r machinery of the fruitfly Drosophila melanogaster. Both TIM and its partn er protein, the period protein PER, show robust circadian oscillations in m RNA and protein levels. Yet the role of TIM in the rhythm generation mechan ism is largely unknown. To analyze TIM function, we constructed transgenic flies that carry a heat shock-inducible copy of the timeless gene (tim) in an arrhythmic tim loss-of-function genetic background. When heat shocked, T IM levels in these flies rapidly increased and initiated a molecular cycle of PER accumulation and processing with dynamics very similar to the PER cy cle observed in wild-type flies. Analysis of period (per) mRNA levels and t ranscription uncovered a novel role for TIM in clock regulation: TIM increa ses per mRNA levels through a post-transcriptional mechanism. Our results s uggest positive as well as negative autoregulation in the Drosophila circad ian clock.