In general, the basic shape of Silkworm (Bombyx mori) cocoons is thought to depend upon race and not the size of the cocoon. That is, it is thought that within species, changes in the size of cocoons are proportional. The ratio of cocoon length to width is commonly used to numerically express the shape of cocoons. In the present study, we employed a Fourier coefficient in order to express the cocoon shape in detail. We found that in some Chinese silkworm races, the basic shape of the cocoon changed with size. The shapes of the geographical silkworm races have been precisely reported by Enokijima et al. (1985) through the ratio of the length to the width of the cocoon.

View full abstract

Here we describe the residue-specific incorporation of para-substituted L-phenylalanine (Phe) analogues into protein biosynthesis in Bombyx mori cultured cells, BmN, which express B. mori phenylalanyl-tRNA synthetase (BmPheRS) mutants with relaxed amino acid specificity. Aminoacylation capabilities of BmPheRS mutants (Ala450 to Gly and Thr407 to Ala or Gly mutants in the α-subunit) were first investigated in vitro and then the incorporation of Phe analogues into a reporter protein, EGFP, was investigated using BmN cells expressing one of the above mutants. We also evaluated the effects of lowering the Phe concentration in culture media, which competes with Phe analogues in aminoacylation reactions inside cells, and found that lowering Phe concentration was effective in increasing the substitution rate from Phe to its analogues in EGFP. We conducted electrophoretic and mass spectrometric analyses of synthesized EGFP to clarify the incorporation of Phe analogues. We obtained direct mass spectrometric data demonstrating the incorporation of p-bromo- and p-cyano-L-phenylalanine and indirect electrophoretic data, implying the incorporation of p-azido- and p-iodo-L-phenylalanine into protein biosynthesis in BmN cells.

View full abstract

The Bombyx mori homologue of the Broad-Complex gene, BmBR-C, is composed of 13 exons, and is transcribed from two promoters—a distal promoter, Pdist, and a proximal promoter, Pprox—separated by a distance of ~101kbp. A highly homologous sequence (referred to as the cEcRE) that contains known binding sites for a functional ecdysone receptor (the canonical EcRE) was discovered on Pdist, however, surprisingly, the cEcRE does not bind to EcR/Usp. Further analyses indicated that two regions within Pdist, EcRE-D and EcRE-P, −4,950bp and −3,480bp upstream from the distal transcription start site, respectively, are important in the responsiveness of Pdist to 20-hydroxyecdysone. However, weak or non-detectable sequence similarities were found between the canonical EcRE and the EcRE-D and EcRE-P regions. Despite the fact that the sequence similarity to the canonical EcRE was less than that observed for the cEcRE that did not bind to EcR/Usp, the results obtained using electrophoretic mobility shift assays (EMSAs) strongly suggested that EcR/Usp could bind to the EcRE-D. In this study, the middle portion of EcRE-D, EcRE-Dbs3, containing an element exhibiting weak similarity to the canonical EcRE proved to be a key sequence mediating EcR/Usp binding to BmBR-C Pdist. Moreover, conserved residues, particularly the G and C residues positioned at −4, −2 and +4, between the EcRE-Dbs3 and the canonical EcRE were essential for EcR/Usp binding and for the ecdysone-driven transcriptional activation of BmBR-C Pdist.

View full abstract