Gene silencing through RNA interference (RNAi) has revolutionized the study of gene 98 function, particularly in non-model insects. However, in Lepidoptera (moths and butterflies) 99 RNAi has many times proven to be difficult to achieve. Most of the negative results have been 100 anecdotal and the positive experiments have not been collected in such a way that they are 101 possible to analyze. In this review, we have collected detailed data from more than 150 102 experiments including all to date published and many unpublished experiments. Despite a 103 large variation in the data, trends that are found are that RNAi is particularly successful in the 104 family Saturniidae and in genes involved in immunity. On the contrary, gene expression in 105 epidermal tissues seems to be most difficult to silence. In addition, gene silencing by feeding 106 dsRNA requires high concentrations for success. Possible causes for the variability of success 107 in RNAi experiments in Lepidoptera are discussed. The review also points to a need to further 108 investigate the mechanism of RNAi in lepidopteran insects and its possible connection to the 109 innate immune response. Our general understanding of RNAi in Lepidoptera will be further 110 aided in the future as our public database at http://insectacentral.org/RNAi will continue to 111 gather information on RNAi experiments.

Although it is well established that the circadian clock regulates mammalian reproductive physiology, the molecular mechanisms by which this regulation occurs are not clear. The authors investigated the reproductive capacity of mice lacking Bmal1 (Arntl, Mop3), one of the central circadian clock genes. They found that both male and female Bmal1 knockout (KO) mice are infertile. Gross and microscopic inspection of the reproductive anatomy of both sexes suggested deficiencies in steroidogenesis. Male Bmal1 KO mice had low testosterone and high luteinizing hormone serum concentrations, suggesting a defect in testicular Leydig cells. Importantly, Leydig cells rhythmically express BMAL1 protein, suggesting peripheral control of testosterone production by this clock protein. Expression of steroidogenic genes was reduced in testes and other steroidogenic tissues of Bmal1 KO mice. In particular, expression of the steroidogenic acute regulatory protein (StAR) gene and protein, which regulates the rate-limiting step of steroidogenesis, was decreased in testes from Bmal1 KO mice. A direct effect of BMAL1 on StAR expression in Leydig cells was indicated by in vitro experiments showing enhancement of StAR transcription by BMAL1. Other hormonal defects in male Bmal1 KO mice suggest that BMAL1 also has functions in reproductive physiology outside of the testis. These results enhance understanding of how the circadian clock regulates reproduction.Keywords circadian rhythms; fertility; testosterone; testes; sperm; StAR; mice Disruption of circadian rhythms results in a variety of pathophysiologic states (Hastings et al., 2003). Reproductive physiology, in particular, is profoundly influenced by circadian rhythms (Boden and Kennaway, 2006). In various insect species, the circadian clock is necessary for proper ovulation, sperm production, and fertility (Giebultowicz et al., 1989;Beaver et al., 2002;Beaver et al., 2003;Beaver and Giebultowicz, 2004 (Lucas and Eleftheriou, 1980;Clair et al., 1985;Chappell et al., 2003;Miller et al., 2004). For example, the surge of luteinizing hormone (LH) necessary for ovulation in rodents, which occurs at the same time of day during each estrous cycle, requires a functional circadian clock (Barbacka-Surowiak et al., 2003). In addition, at the onset of puberty, a clear diurnal rhythm of gonadotropin serum levels is established in both mice and humans (Andrews and Ojeda, 1981;Jean-Faucher et al., 1986;Dunkel et al., 1992;Apter et al., 1993). It is unclear whether this diurnal rhythm continues into adulthood, but testosterone serum concentration shows daily oscillations in adult male mice and humans (Lucas and Eleftheriou, 1980;Clair et al., 1985). Although the association between circadian rhythms and testosterone is a long-established phenomenon, the molecular mechanisms by which the circadian clock regulates testosterone production are unknown.The circadian clock is based on a transcription translation feedback loop that results in the cyclic expression of genes and proteins over a 24-h p...

Light is necessary for life, but prolonged exposure to artificial light is a matter of increasing health concern. Humans are exposed to increased amounts of light in the blue spectrum produced by light-emitting diodes (LEDs), which can interfere with normal sleep cycles. The LED technologies are relatively new; therefore, the long-term effects of exposure to blue light across the lifespan are not understood. We investigated the effects of light in the model organism, Drosophila melanogaster, and determined that flies maintained in daily cycles of 12-h blue LED and 12-h darkness had significantly reduced longevity compared with flies maintained in constant darkness or in white light with blue wavelengths blocked. Exposure of adult flies to 12 h of blue light per day accelerated aging phenotypes causing damage to retinal cells, brain neurodegeneration, and impaired locomotion. We report that brain damage and locomotor impairments do not depend on the degeneration in the retina, as these phenotypes were evident under blue light in flies with genetically ablated eyes. Blue light induces expression of stress-responsive genes in old flies but not in young, suggesting that cumulative light exposure acts as a stressor during aging. We also determined that several known blue-light-sensitive proteins are not acting in pathways mediating detrimental light effects. Our study reveals the unexpected effects of blue light on fly brain and establishes Drosophila as a model in which to investigate long-term effects of blue light at the cellular and organismal level.

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