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Molecular genetic analysis indicates that rhythmic changes in the abundance of the Drosophila lark RNA-binding protein are important for circadian regulation of adult eclosion (the emergence or ecdysis of the adult from the pupal case). To define the tissues and cell types that might be important for lark function, we have characterized the spatial and developmental patterns of lark protein expression. Using immunocytochemical or protein blotting methods, lark can be detected in late embryos and throughout postembryonic development, from the third instar larval stage to adulthood. At the late pupal (pharate adult) stage, lark protein has a broad pattern of tissue expression, which includes two groups of crustacean cardioactive peptide (CCAP)-containing neurons within the ventral nervous system. In other insects, the homologous neurons have been implicated in the physiological regulation of ecdysis. Whereas lark has a nuclear distribution in most cell types, it is present in the cytoplasm of the CCAP neurons and certain other cells, which suggests that the protein might execute two different RNA-binding functions. Lark protein exhibits significant circadian changes in abundance in at least one group of CCAP neurons, with abundance being lowest during the night, several hours prior to the time of adult ecdysis. Such a temporal profile is consistent with genetic evidence indicating that the protein serves a repressor function in mediating the clock regulation of adult ecdysis. In contrast, we did not observe circadian changes in CCAP neuropeptide abundance in late pupae, although CCAP amounts were decreased in newly-emerged adults, presumably because the peptide is released at the time of ecdysis. Given the cytoplasmic localization of the lark RNA-binding protein within CCAP neurons, and the known role of CCAP in the control of ecdysis, we suggest that changes in lark abundance may regulate the translation of a factor important for CCAP release or CCAP cell excitability. © 2000 John Wiley & Sons, Inc. J Neurobiol 45: 14–29, 2000

Photic entrainment of insect circadian rhythms can occur through either extraretinal (brain) or retinal photoreceptors, which mediate sensitivity to blue light or longer wavelengths, respectively. Although visual transduction processes are well understood in the insect retina, almost nothing is known about the extraretinal blue light photoreceptor of insects. We now have identified and characterized a candidate blue light photoreceptor gene inDrosophila(DCry) that is homologous to the cryptochrome (Cry) genes of mammals and plants. TheDCrygene is located in region 91F of the third chromosome, an interval that does not contain other genes required for circadian rhythmicity. The protein encoded byDCryis ∼50% identical to the CRY1 and CRY2 proteins recently discovered in mammalian species. As expected for an extraretinal photoreceptor mediating circadian entrainment,DCrymRNA is expressed within the adult brain and can be detected within body tissues. Indeed, tissuein situhybridization demonstrates prominent expression in cells of the lateral brain, which are close to or coincident with theDrosophilaclock neurons. Interestingly,DCrymRNA abundance oscillates in a circadian manner inDrosophilahead RNA extracts, and the temporal phasing of the rhythm is similar to that documented for the mouseCry1mRNA, which is expressed in clock tissues. Finally, we show that changes inDCrygene dosage are associated predictably with alterations of the blue light resetting response for the circadian rhythm of adult locomotor activity.