Expression of a sugar clade gustatory receptor, BmGr6, in the oral sensory organs, midgut, and central nervous system of larvae of the silkworm Bombyx mori.

Insects taste nonvolatile chemicals through gustatory receptors (Grs) and make choices for feeding, mating, and oviposition. To date, genome projects have identified 69 Gr genes in the silkworm, Bombyx mori; however, the expression sites of these Grs remain to be explored. In this study, we used reverse transcription (RT)-PCR to investigate expression of the B.

Immunohistochemical analysis of the role of hemocytin in nodule formation in the larvae of the silkworm, Bombyx mori.

Hemocytin, a multidomain protein from Bombyx mori L. (Lepidoptera: Bombycidae), is an ortholog of von Willebrand factor and is expected to be a major mediator of hemocyte aggregation. Antiserum was generated against hemocytin, and immune staining of hemocytes, hemolymph, and nodules was performed.

The ATP-binding cassette transporter subfamily C member 2 in Bombyx mori larvae is a functional receptor for Cry toxins from Bacillus thuringiensis.

Bacillus thuringiensis is the most widely used biopesticide, and its Cry toxin genes are essential transgenes for the generation of insect-resistant transgenic crops. Recent reports have suggested that ATP-binding cassette transporter subfamily C2 (ABCC2) proteins are implicated in Cry intoxication, and that a single amino acid insertion results in high levels of resistance to Cry1 toxins. However, there is currently no available direct evidence of functional interactions between ABCC2 and Cry toxins.

Cry1Aa binding to the cadherin receptor does not require conserved amino acid sequences in the domain II loops.

Characterizing the binding mechanism of Bt (Bacillus thuringiensis) Cry toxin to the cadherin receptor is indispensable to understanding the specific insecticidal activity of this toxin. To this end, we constructed 30 loop mutants by randomly inserting four serial amino acids covering all four receptor binding loops (loops α8, 1, 2 and 3) and analysed their binding affinities for Bombyx mori cadherin receptors via Biacore. High binding affinities were confirmed for all 30 mutants containing loop sequences that differed from those of wild-type.

Response of midgut epithelial cells to Cry1Aa is toxin-dependent and depends on the interplay between toxic action and the host apoptotic response.

Cry1Aa is an insecticidal protein produced by Bacilllus thuringiensis. To elucidate the mechanisms of cell/individual death and healing in the midgut epithelium, Bombyx mori larva were given different concentrations of Cry1Aa, and sections from the midgut were examined by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. In the lethal condition, most midgut columnar cells were observed to swell and burst without being stained by TUNEL 4 h after intoxication, and the epithelial layer collapsed by 24 h.

Ligand carrier protein genes expressed in larval chemosensory organs of Bombyx mori.

Expressed sequence tags (ESTs) of the maxillary galea of the silkworm were analyzed to identify proteins involved in food selection systems. From the 1251 redundant genes of the ESTs, we identified 7 odorant-binding protein-like genes (bmObpL), 6 takeout-like genes (bmToL), and 6 chemosensory protein genes (bmCsp). Quantitative RT-PCR analysis indicated that bmObpL1, bmObpL2, bmObpL3, bmObpL5, bmToL1, bmToL3, and bmorCsp15 were predominantly expressed in the larval oral appendages, such as the maxilla, labrum, labium and antenna.

Localization of the serine protease homolog BmSPH-1 in nodules of E. coli-injected Bombyx mori larvae and functional analysis of its role in nodule melanization.

The molecular mechanisms underlying nodule formation and melanization, an important pathogen defense mechanism in insects, are poorly understood. In this study, we investigated the role of BmSPH-1, a catalytically inactive Bombyx mori serine protease homolog, in nodule melanization induced by injection of Escherichia coli cells into the B. mori larval hemocoel.

Analysis of the region for receptor binding and triggering of oligomerization on Bacillus thuringiensis Cry1Aa toxin.

The determination of the receptor-binding region of Cry toxins produced by Bacillus thuringiensis is expected to facilitate an improvement in their insecticidal ability through protein engineering. We analyzed the region on Cry1Aa molecules involved in interactions with the cadherin-like protein receptor BtR175 using cysteine-substituted mutant toxins and several synthetic peptides corresponding to the loops in domain 2. In addition, the region necessary to trigger oligomerization was analyzed using these mutant toxins.

Identification and comparative analysis of three novel C-type lectins from the silkworm with functional implications in pathogen recognition.

C-type lectins can act as pattern recognition receptors (PRRs) in innate immunity. Previously, we identified two C-type lectins from silkworm (Bombyx mori), BmLBP and BmMBP, as PRRs. In the present study, we identified three homologs of these lectins by searching the silkworm genome database.

Location of the Bombyx mori 175kDa cadherin-like protein-binding site on Bacillus thuringiensis Cry1Aa toxin.

To identify and gain a better understanding of the cadherin-like receptor-binding site on Bacillus thuringiensis Cry toxins, it is advantageous to use Cry1Aa toxin, because its 3D structure is known. Therefore, Cry1Aa toxin was used to examine the locations of cadherin-like protein-binding sites. Initial experiments examining the binding compatibility for Cry1Aa toxin of partial fragments of recombinant proteins of a 175kDa cadherin-like protein from Bombyx mori (BtR175) and another putative receptor for Cry1Aa toxin, amino peptidaseN1, from Bo.