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A pilot program was initiated to identify genes from the American oyster, Crassostrea virginica, that are potentially involved in the stress response for use as bioindicators of exposure to environmental pollutants and to toxic and infectious agents. A PCR-based method was used to construct cDNA libraries from pooled embryos and the hemocytes of a single individual. A total of 998 randomly selected clones (expressed sequence tags, ESTs) were sequenced. Approximately 40% of the ESTs are novel sequences. Several potential biomarkers identified include an antimicrobial peptide, recognition molecules (lectin receptors), proteinases and proteinase inhibitors, and a novel metallothionein. Diversity analysis shows that 363 and 286 unique genes were identified from the hemocyte and embryo libraries, respectively, indicating that full-scale EST collection is a valuable approach for the discovery of new genes of potential significance in the molluscan stress response.

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Bombyxin 11, a product of the brain of the adult silkmoth, Bombyx mori, binds to ovarian cells of three different species of lepidoptera, i.e. B. mori (silkmoth), Samia Cynthia ricini (ailanthus moth), and an ovarian cell line of Spodopteru frugiperda (Sf9) (fall armyworm). Crude Sf9 cell membrane preparations were used to show that the purported bombyxin receptor binds its ligand in a specific, saturable, and reversible manner. The dissociation constant of the bombyxin-receptor complex is 260 -C 90 pM. Quantitative binding studies and Scatchard analysis suggest that every Sf9 cell displays 20000 receptors on the surface. The cross-linked bombyxin-receptor ligand complex has an apparent molecular mass of about 300 kDa as determined by SDSIPAGE. Reduction causes the bombyxin receptor to dissociate into two subunits with molecular masses of 90 kDa and 116 kDa. The size and subunit structure of the putative bombyxin receptor on Sf9 cells show some similarities to the mammalian insulin receptor.Keywords: bombyxin ; insulin ; bombyxin receptor; structure/function relationship ; photoactivatible crosslink Bombyxin, an invertebrate developmental factor similar in structure to insulin, was isolated from the brain of the silkmoth, Bombyx mori [I, 21. Bombyxin elicits metamorphosis in brain extirpated dormant pupae of the saturniid moth, Sumia Cynthia ricini [3], by acting on the prothoracic gland to stimulate the synthesis and release of ecdysone, the steroid hormone that causes molting and metamorphosis. This effect is ascribed to two structurally unrelated prothoracicotropic hormones, the smaller of which was later named bombyxin, while the larger one (30 kDa) retained the name prothoracicotropic hormone [4, 51. The structural dissimilarity of the two hormones means that there are different receptors that mediate a similar cell response. Indeed, the prothoracicotropic glands of Manduca sexta were sensitive to the small and large prothoracicotropic hormones, as demonstrated by Bollenbacher et al. [6].Although initial observations pointed to bombyxin as a prothoracicotropic hormone 171, its biological function in B. mori remained obscure [8, 91. Bombyxin titers in the hemolymph of postembryonic B. mori showed no correlation with ecdysone during molting [ 101. Furthermore, applications of physiological concentrations of bombyxin to brain extirpated dormant B. mori were ineffective in inducing metamorphosis [3].Based on the hypothesis that a structural relationship may indicate functional similarity, the insulin-like bombyxin has been suspected to have an effect on the blood sugar of insects, for instance in the metabolism of trehalose [Ill. Bombyxin was detected in the ovaries of the silkmoth [I21 and significantly higher levels of bombyxin were found in the hemolymph of the female [lo]. This observation suggests a role for bombyxin similar to that of the mammalian ovarian hormone relaxin, another member of the insulin family. It seems possible that bombyxin is involved in the ovarian development [13] of several specie...