Patrick Emery

The cryb mutation identifies cryptochrome as a circadian photoreceptor in Drosophila.

A new rhythm mutation was isolated based on its elimination of per-controlled luciferase cycling. Levels of period or timeless clock gene products in the mutant are flat in daily light-dark cycles or constant darkness (although PER and TIM oscillate normally in temperature cycles). Consistent with the fact that light normally suppresses TIM, cryb is an apparent null mutation in a gene encoding Drosophilas version of the blue light receptor cryptochrome. Behaviorally, cryb exhibits poor synchronization to light-dark cycles in genetic backgrounds that cause external blindness or demand several hours of daily rhythm resets, and it shows no response to brief light pulses. cryb flies are rhythmic in constant darkness, correlating with robust PER and TIM cycling in certain pacemaker neurons.

CRY, a Drosophila Clock and Light-Regulated Cryptochrome, Is a Major Contributor to Circadian Rhythm Resetting and Photosensitivity

Light is a major environmental signal for circadian rhythms. We have identified and analyzed cry, a novel Drosophila cryptochrome gene. All characterized family members are directly photosensitive and include plant blue light photoreceptors. We show that cry transcription is under circadian regulation, influenced by the Drosophila clock genes period, timeless, Clock, and cycle. We also show that cry protein levels are dramatically affected by light exposure. Importantly, circadian photosensitivity is increased in a cry-overexpressing strain. These physiological and genetic data therefore link a specific photoreceptor molecule to circadian rhythmicity. Taken together with the data in the accompanying paper, we propose that CRY is a major Drosophila photoreceptor dedicated to the resetting of circadian rhythms.

Drosophila CRY is a deep brain circadian photoreceptor.

cry (cryptochrome) is an important clock gene, and recent data indicate that it encodes a critical circadian photoreceptor in Drosophila. A mutant allele, cry(b), inhibits circadian photoresponses. Restricting CRY expression to specific fly tissues shows that CRY expression is needed in a cell-autonomous fashion for oscillators present in different locations. CRY overexpression in brain pacemaker cells increases behavioral photosensitivity, and this restricted CRY expression also rescues all circadian defects of cry(b) behavior. As wild-type pacemaker neurons express CRY, the results indicate that they make a striking contribution to all aspects of behavioral circadian rhythms and are directly light responsive. These brain neurons therefore contain an identified deep brain photoreceptor, as well as the other circadian elements: a central pace-maker and a behavioral output system.

Cryptochromes Define a Novel Circadian Clock Mechanism in Monarch Butterflies That May Underlie Sun Compass Navigation

The circadian clock plays a vital role in monarch butterfly (Danaus plexippus) migration by providing the timing component of time-compensated sun compass orientation, a process that is important for successful navigation. We therefore evaluated the monarch clockwork by focusing on the functions of a Drosophila-like cryptochrome (cry), designated cry1, and a vertebrate-like cry, designated cry2, that are both expressed in the butterfly and by placing these genes in the context of other relevant clock genes in vivo. We found that similar temporal patterns of clock gene expression and protein levels occur in the heads, as occur in DpN1 cells, of a monarch cell line that contains a light-driven clock. CRY1 mediates TIMELESS degradation by light in DpN1 cells, and a light-induced TIMELESS decrease occurs in putative clock cells in the pars lateralis (PL) in the brain. Moreover, monarch cry1 transgenes partially rescue both biochemical and behavioral light-input defects in cryb mutant Drosophila. CRY2 is the major transcriptional repressor of CLOCK:CYCLE-mediated transcription in DpN1 cells, and endogenous CRY2 potently inhibits transcription without involvement of PERIOD. CRY2 is co-localized with clock proteins in the PL, and there it translocates to the nucleus at the appropriate time for transcriptional repression. We also discovered CRY2-positive neural projections that oscillate in the central complex. The results define a novel, CRY-centric clock mechanism in the monarch in which CRY1 likely functions as a blue-light photoreceptor for entrainment, whereas CRY2 functions within the clockwork as the transcriptional repressor of a negative transcriptional feedback loop. Our data further suggest that CRY2 may have a dual role in the monarch butterflys brain—as a core clock element and as an output that regulates circadian activity in the central complex, the likely site of the sun compass.

Drosophila cryptochromes: A unique circadian-rhythmphotoreceptor

Cryptochrome proteins are critical for circadian rhythms, but their function(s) is uncertain. Here we show that a mutation in a cryptochrome (dCRY) from the fruitfly Drosophila blocks an essential photoresponse of circadian rhythms, namely arrhythmicity under constant light conditions. We conclude that dCRY acts as a key photoreceptor for circadian rhythms and that there is probably no other comparable photoreceptor in this species.

A unique circadian-rhythm photoreceptor.

Cryptochrome proteins are critical for circadian rhythms, but their function(s) is uncertain. Here we show that a mutation in a cryptochrome (dCRY) from the fruitfly Drosophila blocks an essential photoresponse of circadian rhythms, namely arrhythmicity under constant light conditions. We conclude that dCRY acts as a key photoreceptor for circadian rhythms and that there is probably no other comparable photoreceptor in this species.

Light and Temperature Control the Contribution of Specific DN1 Neurons to Drosophila Circadian Behavior

The brain of Drosophila melanogaster contains approximately 150 circadian neurons [1] functionally divided into morning and evening cells that control peaks in daily behavioral activity at dawn and dusk, respectively [2, 3]. The PIGMENT DISPERSING-FACTOR (PDF)-positive small ventral lateral neurons (sLN(v)s) promote morning behavior, whereas the PDF-negative sLN(v) and the dorsal lateral neurons (LN(d)s) generate evening activity. Much less is known about the approximately 120 dorsal neurons (DN1, 2, and 3). Using a Clk-GAL4 driver that specifically targets a subset of DN1s, we generated mosaic per(0) flies with clock function restored only in these neurons. We found that the Clk4.1M-GAL4-positive DN1s promote only morning activity under standard (high light intensity) light/dark cycles. Surprisingly, however, these circadian neurons generate a robust evening peak of activity under a temperature cycle in constant darkness. Using different light intensities and ambient temperatures, we resolved this apparent paradox. The DN1 behavioral output is under both photic and thermal regulation. High light intensity suppresses DN1-generated evening activity. Low temperature inhibits morning behavior, but it promotes evening activity under high light intensity. Thus, the Clk4.1M-GAL4-positive DN1s, or the neurons they target, integrate light and temperature inputs to control locomotor rhythms. Our study therefore reveals a novel mechanism contributing to the plasticity of circadian behavior.

A rhythmic ror

The circadian clock mechanism in mammals involves two interlocking transcriptional feedback loops. Rev-erb alpha, through its role as a transcriptional repressor, was thought to be the primary determinant of the feedback loop that regulates Bmal1 transcription. Results reported by Sato et al. in this issue of Neuron now show that the transactivator Rora acts coordinately with Rev-erb alpha and that their competing activities on the same promoter element drive the rhythm in Bmal1 transcription. This finding defines the second feedback loop in mammals.

A SUBSET OF DORSAL NEURONS MODULATES CIRCADIAN BEHAVIOR AND LIGHT RESPONSES IN DROSOPHILA

A fundamental property of circadian rhythms is their ability to persist under constant conditions. In Drosophila, the ventral Lateral Neurons (LNvs) are the pacemaker neurons driving circadian behavior under constant darkness. Wild-type flies are arrhythmic under constant illumination, but flies defective for the circadian photoreceptor CRY remain rhythmic. We found that flies overexpressing the pacemaker gene per or the morgue gene are also behaviorally rhythmic under constant light. Unexpectedly, the LNvs do not drive these rhythms: they are molecularly arrhythmic, and PDF--the neuropeptide they secrete to synchronize behavioral rhythms under constant darkness--is dispensable for rhythmicity in constant light. Molecular circadian rhythms are only found in a group of Dorsal Neurons: the DN1s. Thus, a subset of Dorsal Neurons shares with the LNvs the ability to function as pacemakers for circadian behavior, and its importance is promoted by light.

Drosophila Clock Can Generate Ectopic Circadian Clocks

Circadian rhythms of behavior, physiology, and gene expression are present in diverse tissues and organisms. The function of the transcriptional activator, Clock, is necessary in both Drosophila and mammals for the expression of many core clock components. We demonstrate in Drosophila that Clock misexpression in nai;ve brain regions induces circadian gene expression. This includes major components of the pacemaker program, as Clock also activates the rhythmic expression of cryptochrome, a gene that CLOCK normally represses. Moreover, this ectopic clock expression has potent effects on behavior, radically altering locomotor activity patterns. We propose that Clock is uniquely able to induce and organize the core elements of interdependent feedback loops necessary for circadian rhythms.