Identification and Expression Analysis of Ras Gene in Silkworm, Bombyx mori

Abstract

Ras proteins play important roles in development especially for cell proliferation and differentiation in various organisms. However, their functions in the most insect species are still not clear. We identified three ras cDNAs from the silk worm, Bombyx mori. These sequences corresponded to three Ras of Drosophila melanogaster, but not to three mammalian Ras (H-Ras, K-Ras, N-Ras). Subsequently, the expression profiles of ras were investigated by quantitative real-time PCR using whole body of individuals from the embryonic to adult stages, and various tissues of 4th and 5th instar larvae. Each of three Bombyx ras showed different expression patterns. We also showed membrane localization of their products. These results indicate that the three Bombyx Ras are functional and have different roles.

Ras1CA overexpression in the posterior silk gland improves silk yield

“…In Bombyx, the Ras1 mutant Ras1 V12 (termed Ras1 CA ) is constitutively active [7,9]. To improve silk yield by transgenic breeding, the GAL4/UAS system was used to overexpress Ras1 CA specifically in the posterior silk gland (Figure 1 Figure 2B and 2B'; Supplementary information, Figure S3B), which was a smaller proportional change than the posterior silk gland weight (60%).…”

“…The above two PCR fragments were conjugated, digested with AscI and SacII, and inserted into pBac{3XP3-DsRed} [14] to generate Fil-GAL4. The cDNA of Bombyx Ras1 oncogene [9] was amplified using forward primer containing EcoRI and reverse primer containing NotI. Ras1 CA was generated by substituting valine for glycine at the 12 th amino acid residue of Ras1 (Ras1 V12 ), which impairs the protein's ability to hydrolyze GTP [7].…”

“…There are three Ras genes in the Bombyx genome [9,10], but their biological significance is largely unknown. Because the posterior silk gland grows dramatically during larval life and its protein synthesis capacity is extremely high at the last instar stage [2,3], we hypothesized that Ras activity might be involved in regulating fibroin production and silk yield.…”

Comparative proteomic analysis of the silkworm middle silk gland reveals the importance of ribosome biogenesis in silk protein production

“…Of the few specific proteins, the Ras protein may closely correlate with the function of the MSG-M. Of the three Bombyx Ras genes with different expression patterns [46], the activated Ras1 in the PSG of silkworms can increase cell and nuclei sizes, enrich subcellular organelles related to protein synthesis, and stimulate ribosome biogenesis, which together leads to improved silk yields [47]. In addition, highly expressed sericin 1, eIFs and ribosomal proteins were identified, confirming that the MSG-M is the most important region for sericin synthesis.…”

Bombyx mori Ras proteins BmRas1, BmRas2 and BmRas3 are neither farnesylated nor palmitoylated but are geranylgeranylated

“…Recently, cDNAs coding for three Ras proteins (BmRas1, BmRas2 and BmRas3) were cloned from the silkworm, Bombyx mori (Ogura et al. , 2009).…”