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review, we have collected detailed data from more than 150 102 experiments including all to date published and many unpublished experiments.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.

0 5In Batesian mimicry, animals avoid predation by resembling distasteful models. In the swallowtail butterfly Papilio polytes, only mimetic-form females resemble the unpalatable butterfly Pachliopta aristolochiae. A recent report showed that a single gene, doublesex (dsx), controls this mimicry 1 ; however, the detailed molecular mechanisms remain unclear. Here we determined two whole-genome sequences of P. polytes and a related species, Papilio xuthus, identifying a single ~130-kb autosomal inversion, including dsx, between mimetic (H-type) and non-mimetic (h-type) chromosomes in P. polytes. This inversion is associated with the mimicry-related locus H, as identified by linkage mapping. Knockdown experiments demonstrated that female-specific dsx isoforms expressed from the inverted H allele (dsx(H)) induce mimetic coloration patterns and simultaneously repress non-mimetic patterns. In contrast, dsx(h) does not alter mimetic patterns. We propose that dsx(H) switches the coloration of predetermined wing patterns and that female-limited polymorphism is tightly maintained by chromosomal inversion.

Telomerase is a specialized DNA polymerase that extends the 3' ends of eukaryotic linear chromosomes, a process required for genomic stability and cell viability. Here we present the crystal structure of the active Tribolium castaneum telomerase catalytic subunit, TERT, bound to an RNA-DNA hairpin designed to resemble the putative RNA-templating region and telomeric DNA. The RNA-DNA hybrid adopts a helical structure, docked in the interior cavity of the TERT ring. Contacts between the RNA template and motifs 2 and B' position the solvent-accessible RNA bases close to the enzyme active site for nucleotide binding and selectivity. Nucleic acid binding induces rigid TERT conformational changes to form a tight catalytic complex. Overall, TERT-RNA template and TERT-telomeric DNA associations are remarkably similar to those observed for retroviral reverse transcriptases, suggesting common mechanistic aspects of DNA replication between the two families of enzymes.

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A pentanucleotide repetitive sequence, (TTAGG)., has been isolated from a silkworm genomic library, using cross-hybridization with a (TTNGGG)5 sequence, which is conserved among most eukaryotic telomeres. Both fluorescent in situ hybridization and Bal 31 exonuclease experiments revealed major clusters of (TTAGG) A telomere is defined as a region of a chromosomal end that is required for complete replication, meiotic pairing, and stability of the chromosome (5, 36). Most eukaryotic telomeres are composed of simple repeated sequences of a Gand C-rich complementary strand with a general structure of (T or A)m(G)n. The G-rich strand is oriented 5' to 3' toward the chromosomal end and is synthesized by an RNA-dependent telomerase activity (13-15, 22, 35). At present, telomeric DNA has been identified in many widely divergent organisms, including yeasts, protozoa, slime molds, flagellates, nematodes, plants, and vertebrates (6, 23).Telomeres for most invertebrates and insects, however, have yet to be isolated and characterized, although some telomeric DNA fragments have been isolated from species of two dipteran genera, Drosophila and Chironomus (3,29,30,32,34). Such dipteran telomeres seem to have complex and exceptional structures in which components can be distinguished with a simple repeat shown in most other eukaryotic organisms. However, the question still remains as to whether the dipteran insects carry a tandem array at the extreme ends of chromosomes. To examine evolutionarily conserved features of the telomeric terminal array in insects, we have identified the telomere sequence in the silkworm Bombyx mori, which is a representative of the lepidopteran order of insects.It is noteworthy that some insects, such as hemipteran and lepidopteran orders, including the silkworm, are believed to have a diffuse type of centromere (24,25). In this case, chromosomes with full-length kinetochores are not sensitive to breakage, induced either spontaneously or by X-ray irradiation, so that chromosome fragments are maintained through cell division (10). Therefore, chromosome fragmentation and de novo telomere formation at the breakage site may provide opportunities to make new chromosomes. Furthermore, translocation of the chromosomal fragment to * Corresponding author.other chromosomes can possibly alter chromosome organization. To study the mechanisms of karyotype evolution, it is of interest to determine the structure and distribution of telomeres in this insect.In this study, we have isolated a five-base repetitive sequence from a silkworm genomic library, based on crosshybridization with a telomeric sequence (TTNGGG) which is highly conserved among a wide variety of eukaryotic organisms. The results of fluorescent in situ hybridization and Bal 31 exonuclease digestion experiments suggest that this (TTAGG)n sequence is located at the extreme terminal region of all Bombyx chromosomes. In addition, we have demonstrated that (TTAGG)n, slightly different from (TTAGGG)n which is conserved among vertebrates and some invertebra...