Toxin-binding proteins isolated from yellow mealworm Tenebrio molitor and wax moth Galleria mellonella

Abstract

A 67-kDa protein that can specifically bind the activated Cry9A endotoxin under ligand-blotting conditions was purified from midgut epithelium apical membranes of wax moth Galleria mellonella by affinity chromatography. N-Terminal amino acid sequencing enabled identification of this protein as aminopeptidase N. In similar experiments, 66- and 58-kDa proteins specific to endotoxin Cry3A were isolated from the midgut epithelium apical membranes of Tenebrio molitor larvae. Mass spectrometry showed close similarity of the 58-kDa protein to the Tenebrio molitor α-amylase.

Investigation of Cry3Bb1 resistance and intoxication in western corn rootworm by RNA sequencing

“…The differential expression comparison that provided the most useful information was the full data set analysis of strain effects after 8 hr of exposure, as it measured differences between strains across treatments ( Table S2). The annotations of the differentially expressed transcripts in this comparison highlighted various transcript categories previously associated with Bt resistance and Bt mode of action in other insects, including an ABC transporter, an amylase and proteases (Bulushova et al, 2011;Gahan et al, 2010;Oppert et al, 1994) (Table 2). The possibility that the selected transcripts play a role in WCR resistance to Cry3Bb1 is hypothesized based on resistance mechanisms described in other insects, and their fitness costs F I G U R E 4 Expression of five transcripts of interest in third instar midgut versus the rest of the body previously evaluated by Heckel (1994) and reviewed by Gassmann, Carrière, and Tabashnik (2009).…”

“…Glycosyl hydrolases, particularly α‐amylases, also have been identified as potential Cry toxin receptors (Bulushova et al., ; Fernandez‐Luna et al., ). A midgut α‐amylase was identified as a potential receptor of Cry4Ba and Cry11Aa in the mosquito Anopheles albimanus Wiedemann (Fernandez‐Luna et al., ) , and as a functional receptor of Cry3A in the coleopteran Tenebrio molitor Linnaeus (Bulushova et al., ). However, the differentially expressed amylases identified in this study were more highly expressed in the resistant Hopkinton strain.…”

“…The increase in amylase abundance in the Hopkinton strain may indicate toxin sequestration by an alternate binding protein. The α‐amylase described as a receptor for Cry3A in T. molitor was soluble in the gut (Bulushova et al., ), which would imply no physical link between this amylase and the epithelial cells and would reinforce the possibility of its participation in resistance by avoiding membrane integration of the toxin and pore formation. The significant differential expression in our study of one of the α‐amylases, characterized in the full data set strain effect and in the Cry3Bb1‐induced difference, indicates involvement of the transcript in Cry3Bb1 exposure.…”

“…It was also described as a receptor of Cry3Aa in L. decemlineata (Ochoa‐Campuzano, Real, Martínez‐Ramírez, Bravo, & Rausell, ). A cadherin, a soluble α‐amylase and a GPI‐anchored alkaline phosphatase have been characterized as potential receptors for Cry3Aa in Tenebrio molitor Linnaeus (Bulushova et al., ; Oppert, Dowd, et al., ; Zúñiga‐Navarrete, Gómez, Peña, Bravo, & Soberón, ). Differential expression of serine peptidases also could play a role in the efficacy of the protoxin as their expression was greatly reduced during Cry3Aa intoxication in susceptible T. molitor (Oppert, Martynov, & Elpidina, ).…”

Different forms of apolipophorin III in Galleria mellonella larvae challenged with bacteria and fungi

“…Although exopeptidases have been reported in the hemolymph and fat body (in addition to the midgut) of some insect species, e.g. dipteran D. melanogaster, and lepidopterans Lacanobia oleracea, M. sexta, and Spodoptera littoralis [3,30,38], in G. mellonella such enzymes have been studied so far only in the midgut [8,9]. Comparison of the pI values (6.1, 5.9) of the main additional apoLp-III forms detected in the immune hemolymph with the pI value 6.5 of the most abundant form would suggest C-terminal limited proteolysis.…”