VRILLE Feeds Back to Control Circadian Transcription of Clock in the Drosophila Circadian Oscillator

Abstract

The Drosophila circadian oscillator consists of interlocked period (per)/timeless (tim) and Clock (Clk) transcriptional/translational feedback loops. Within these feedback loops, CLK and CYCLE (CYC) activate per and tim transcription at the same time as they repress Clk transcription, thus controlling the opposite cycling phases of these transcripts. CLK-CYC directly bind E box elements to activate transcription, but the mechanism of CLK-CYC-dependent repression is not known. Here we show that a CLK-CYC-activated gene, vrille (vri), encodes a repressor of Clk transcription, thereby identifying vri as a key negative component of the Clk feedback loop in Drosophila's circadian oscillator. The blue light photoreceptor encoding cryptochrome (cry) gene is also a target for VRI repression, suggesting a broader role for VRI in the rhythmic repression of output genes that cycle in phase with Clk.

Modelling of circadian rhythms in Drosophila incorporating the interlocked PER/TIM and VRI/PDP1 feedback loops

“…These results are consistent with observations that per and tim mRNA levels oscillate in phase to one another and they reach peak levels early in the evening at CT12 -CT16 (Hardin, Hall et al 1990); clk mRNA levels oscillate in anti-phase to per and tim mRNA levels and clk mRNA levels peak at late night to early in the morning (CT23 -CT4) ; the simulated concentration of vri mRNA reaches a peak at CT11.5 and that of pdp1 mRNA reaches its maximum at CT13.5 with a 2 h delay. This agrees with experimental data that vri mRNA oscillates in anti-phase with clk mRNA, and pdp1 mRNA oscillates with a similar phase to vri mRNA after several hours delay (Cyran, Buchsbaum et al 2003;Glossop, Houl et al 2003). Figure 3B illustrates the oscillations in concentrations of proteins.…”

“…This loop starts with activation of vri and pdp1 transcription by CLK/CYC during the late day and early night. Like per and tim genes, E-boxes are also found in the promoters of the vri and pdp1 genes and CLK/CYC dimers have been shown to activate vri and pdp1 expression in vitro in an E-box-dependent manner (Cyran, Buchsbaum et al 2003;Glossop, Houl et al 2003). VRI accumulates first in phase with its mRNA then PDP1 accumulates during the mid to late evening.…”

Clk post-transcriptional control denoises circadian transcription both temporally and spatially

“…The latter is constitutively expressed, whereas CLK is present in limiting amounts 4,10 . The CLK-CYC heterodimer also activates the expression of VRILLE (VRI) and PAR-DOMAIN-PROTEIN 1 (PDP1), which have been postulated to be responsible for the oscillation of Clk mRNA 11,12 . Although the role of VRI in repressing Clk transcription is well established, recent reports suggest that PDP1 might not directly regulate Clk 13 .…”

“…The current model states that most (if not all) of this control is through post-translational regulation such as phosphorylation, ubiquitination and other types of modifications [21][22][23][24][25] . Indeed, although Clk mRNA levels display strong transcriptional oscillations 11,12 , CLK protein levels are nearly constant though the day 26 . Moreover, expression of CLK under the tim or per promoter in ClkARK fly strains (in which transcription has the opposite daily phase relative to that under control of the Clk promoter 21 ) does not disrupt circadian behaviour, indicating that flies can adapt or compensate for high levels of CLK 21 .…”

Even a stopped clock tells the right time twice a day: circadian timekeeping in Drosophila

“…Here, CLK/CYC activates transcription of vrille (vri) and PAR-domain protein 1ɛ (Pdp1ɛ), which encode related basic leucine zipper transcription factors [39][40][41]. VRI and PDP1ɛ regulate Clk transcription, with VRI acting as a repressor and PDP1ɛ an activator of transcription, causing Clk RNA levels to cycle in opposite phase to per, tim, vri and Pdp1ɛ [39,40]. vri RNA and protein levels peak ∼3-6 h before those of Pdp1ɛ, presumably underpinning the daily rhythm in Clk expression [40].…”