Peripheral circadian clock for the cuticle deposition rhythm in Drosophila melanogaster

Abstract

Insect endocuticle thickens after adult emergence by daily alternating deposition of two chitin layers with different orientation. Although the cuticle deposition rhythm is known to be controlled by a circadian clock in many insects, the site of the driving clock, the photoreceptor for entrainment, and the oscillatory mechanism remain elusive. Here, we show that the cuticle deposition rhythm is regulated by a peripheral oscillator in the epidermis in Drosophila melanogaster. Free-running and entrainment experiments in vitro reveal that the oscillator for the cuticle deposition rhythm is independent of the central clock in the brain driving the locomotor rhythms. The cuticle deposition rhythm is absent in null and dominant-negative mutants of clock genes (i.e., period, timeless, cycle, and Clock), indicating that this oscillator is composed of the same clock genes as the central clock. Entrainment experiments with monochromatic light-dark cycles and cry b flies reveal that a blue light-absorbing photoreceptor, cryptochrome (CRY), acts as a photoreceptor pigment for the entrainment of the cuticle deposition rhythm. Unlike other peripheral rhythms in D. melanogaster, the cuticle deposition rhythm persisted in cry b and cry OUT mutant flies, indicating that CRY does not play a core role in the rhythm generation in the epidermal oscillator.circadian rhythm ͉ clock genes ͉ cryptochrome ͉ entrainment ͉ epidermal cell C ircadian clocks control daily rhythms at the molecular, physiological, and behavioral levels. Like most organisms, the central circadian oscillator controlling the locomotor activity rhythm in Drosophila is proposed to consist of molecular feedback loops. The feedback loops comprise several core clock genes, such as period (per), timeless (tim), Clock (Clk), and cycle (cyc) (1). These genes encode PER, TIM, CLK, and CYC proteins, respectively, and loss-of-function or dominant-negative mutations in these genes result in loss of normal circadian rhythmicity (2-5). CLK and CYC form a heterodimer to activate transcription of per and tim (2, 4, 6). PER and TIM form another heterodimer, are transferred into the nucleus, and then inactivate the transcriptional activity of the CLK/CYC heterodimer (6). The feedback loops are entrained and reset by the lightinduced degradation of TIM mediated by a photoreceptor pigment, cryptochrome (CRY) (7-10).In addition to the central oscillator, peripheral oscillators exist in many tissues, including the compound eyes, antennae, wings, legs, and Malpighian tubules in adults and the ring gland in pupae (11)(12)(13)(14). These oscillators are self-sustained and directly light-entrainable even in vitro (11-13). In addition, the oscillators are clearly independent of the central circadian oscillator(s) in the brain (15, 16). Previous reports revealed that the peripheral oscillator shares the same genes with the central clock in the brain (12, 17). However, there is a remarkable difference that, in the peripheral oscillator, CRY functions as a core component besides a photorecepto...

Genome-wide profiling of diel and circadian gene expression in the malaria vector Anopheles gambiae

“…Daily cuticle deposition in the inner apodemes has been noted in An. gambiae, forming discrete layers that are reliable indicators of mosquito age in days (93), and in many insect species this deposition is under circadian regulation (94,95). In fact, in Drosophila this rhythm is regulated locally by a peripheral oscillator in the epidermis (95).…”

A comparative view of insect circadian clock systems

“…These results indicate that, instead of a photoreceptor in the central pacemaker, CRY1 functions as a core component in these peripheral oscillators [112]. Unlike these, however, another peripheral rhythm, which was recently found in the cuticle deposition, persisted in cry mutant flies [113]. Thus, there are at least two kinds of circadian molecular machinery in peripheral oscillators: one that requires CRY1 as an essential component, and the other that includes it just as a photoreceptor.…”

How Light Resets Circadian Clocks

“…Even the rhythms of cultured body parts are synchronized by LD cycles , showing that the underlying peripheral oscillators are cell autonomous and photo responsive. Light entrains these oscillators directly, probably via CRY (Ivanchenko et al 2001;Ito et al 2008) (see Sect. 18.6.4).…”