Tibetan Firefly Luciferase with Low Temperature Adaptation.

Fireflies are widespread all over the world and a numerous numbers of luciferases have been isolated and characterized. In this study, we identified and characterized the luciferase and luciferase-like genes from a Tibetan firefly collected in Shangri-La, China. The altitude of this area is more than 3300 m. We saw this Tibetan firefly flying with strong luminescence after sunset at ~10°C. We analyzed the transcriptome of Tibetan firefly using head, thorax, abdomen (without light organ), and light organ tissue by RNA sequencing. We identified one luciferase gene, which was almost identical to luciferase from fireflies Pyrocoelia species, and expressed specifically in the light organ. Interestingly, the optimal temperature of the Tibetan firefly recombinant luciferase was 10°C. The Km for D-luciferin and ATP of the recombinant luciferase was 23 and 154 µm, respectively. The optimal pH was around 7.0-7.5. The emission peak was 556 nm at pH 8.0, while it shifted to 606 nm at pH 6.0. We also found a luciferase-like gene with 43% identical amino acids to the Tibetan firefly luciferase, which was scarcely expressed in any portion of the adult body. No luciferase activity was detected for this luciferase-like protein.

Coupling between elytra of some beetles: mechanism, forces and effect of surface texture.

Lightweight materials, structures and coupling mechanisms are very important for realizing advanced flight vehicles. Here, we obtained the geometric structures and morphologies of the elytra of beetles and ascertained its coupling zone by using the histological section technique and SEM. We set up a three-dimensional motion observing system to monitor the opening and closing behaviour of elytra in beetles and to determine the motion mechanism. We constructed a force measuring system to measure the coupling forces between elytra. The results show that elytra open and close by rotating about a single axle, where the coupling forces may be as high as 160 times its own bodyweight, the elytra coupling with the tenon and mortise mechanism, surface texture and opening angle between elytra heavily influence the coupling forces. These results may provide insights into the design mechanism and structure for future vehicles of flight.

A novel serine protease inhibitor from Bungarus fasciatus venom.

By Sephadex G-50 gel filtration, cation-exchange CM-Sephadex C-25 chromatography and reversed phase high-performance liquid chromatography (HPLC), a novel serine protease inhibitor named bungaruskunin was purified and characterized from venom of Bungarus fasciatus. Its cDNA was also cloned from the cDNA library of B. fasciatus venomous glands. The predicted precursor is composed of 83 amino acid (aa) residues including a 24-aa signal peptide and a 59-aa mature bungaruskunin. Bungaruskunin showed maximal similarity (64%) with the predicted serine protease inhibitor blackelin deduced from the cDNA sequence of the red-bellied black snake Pseudechis porphyriacus. Bungaruskunin is a Kunitz protease inhibitor with a conserved Kunitz domain and could exert inhibitory activity against trypsin, chymotrypsin, and elastase. By screening the cDNA library, two new B chains of beta-bungarotoxin are also identified. The overall structures of bungaruskunin and beta-bungarotoxin B chains are similar; especially they have highly conserved signal peptide sequences. These findings strongly suggest that snake Kunitz/BPTI protease inhibitors and neurotoxic homologs may have originated from a common ancestor.

Mitochondrial genomes of two luminous beetles, Rhagophthalmus lufengensis and R. ohbai (Arthropoda, Insecta, Coleoptera).

We determined the mitochondrial DNA (mtDNA) sequences of two luminous beetles (Arthropoda, Insecta, Coleoptera), Rhagophthalmus lufengensis from Yunnan, China and Rhagophthalmus ohbai from Yaeyama Island, Japan. We identified all the 37 mtDNA genes of R. lufengensis (15,982 bp) and the 34 genes of R. ohbai (15,704 bp). R. lufengensis and R. ohbai genomes have higher A + T contents than other coleopteran genomes although the gene arrangements are similar. Interestingly, in a study of the evolutionary relationship among R. lufengensis, R. ohbai and the firefly Pyrocoelia rufa, the phylogenetic tree inferred from lrRNA genes from mitochondrial genomes indicates a biogeographic relationship among the bioluminescent insects in East Asia and the phylogenetic tree inferred from luciferase-related genes from nuclear genomes shows an appropriate relationship among coleopterans, reflecting the evolutionary origin of bioluminescence. Thus, the mtDNAs of luminescent beetles can provide an insight into their evolutionary origin and biogeography.

Genetic differentiation and cryptic speciation in natural populations of Drosophila lacertosa.

Drosophila lacertosa, an Oriental member of the robusta species group in the virilis-repleta radiation, has a wide distribution from northern India throughout China to the Far East. Phylogenetic analyses of mitochondrial ND2 gene sequences revealed two genetically significantly diverged lineages with 2.1% DNA sequence divergence. These two lineages are largely allopatric: one is mainly found in southwest China (lineage A), whereas the other ranges central and east of China to northeast Asia (lineage B). The geographic distributions of these two clades narrowly overlap near the provincial boundaries of Yunnan and Guizhou or Guangxi, probably as a consequence of population expansion and secondary contact. These two lineages were estimated to have diverged about 0.56 Mya, and the estimated time of expansion was approximately 70,000 years ago for lineage A population, and 110,000 years ago for lineage B population. The present molecular analysis, together with nearly complete reproductive isolation between those two lineages, strongly suggests that the two lineages might be incipient species.

Study on the genus Euops Schoenherr (Coleoptera: attelabidae) from China, with descriptions of a new subgenus and six new species.

The present paper deals with 19 species in four subgenera of the attelabid genus Euops from China (including Taiwan), of which six species and a subgenus are new to science. The subgenus Synaptops is newly divided into three species-groups. A key to species, with photographs and line illustrations of important features of new species is provided.