Bacillus thuringiensis CrylAa delta-endotoxin affects the K+/amino acid symport in Bombyx mori larval midgut.
We have examined the type of inhibition exerted by an activated preparation of the Bacillus thuringiensis delta-endotoxin CrylAa on K+-dependent leucine transport into midgut brush border membrane vesicles or epithelial cells of the isolated midgut from Bombyx mori to study its possible interaction with the amino acid symporter. K+ permeability and the cation-dependent amino acid translocation into brush border membrane vesicles were evaluated by monitoring the fluorescence of the voltage-sensitive cyanine dye 3, 3'-dipropylthiadicarbocyanine iodide. The symporter ability to accept Na+ instead of K+ was exploited and the dissipation of an imposed inside-negative potential (K+ gradient in>out and valinomycin) was registered in the presence of a Na+ gradient (out>in) and of the amino acid. The fluorescence quenching dissipated more rapidly when the amino acid was present. Preincubation of brush border membrane vesicles with CrylAa caused a significant decrease of the amino acid-dependent recovery of fluorescence, whereas K+ permeability was sparely affected. In the isolated midgut, CrylAa inhibits leucine uptake as well as the transepithelial electrical potential difference. The strong inhibition exerted by the delta-endotoxin was observed also in the absence of potassium and the transepithelial electrical potential difference. The results obtained strongly suggest a direct interaction of CrylAa delta-endotoxin with the K+/amino acid symporter.
Leonardi
M G
MG
Department of Biology, University of Milan, via Celoria 26, 20133 Milano, Italy.
Parenti
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P
Casartelli
M
M
Giordana
B
B
D026905:Amino Acid Transport Systems; D000818:Animals; D001413:Bacillus thuringiensis; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000494:pharmacology*; D001427:Bacterial Toxins / Q000494:pharmacology*; D001692:Biological Transport / Q000187:drug effects; D012831:Bombyx / Q000187:drug effects*; D002352:Carrier Proteins / Q000187:drug effects / Q000378:metabolism; D004731:Endotoxins / Q000494:pharmacology*; D006460:Hemolysin Proteins; D007306:Insecticides / Q000494:pharmacology*; D007814:Larva / Q000187:drug effects; D007930:Leucine / Q000378:metabolism*; D011188:Potassium / Q000378:metabolism*; D015221:Potassium Channels / Q000187:drug effects / Q000378:metabolism; D012964:Sodium / Q000378:metabolism
D026905:Amino Acid Transport Systems; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D002352:Carrier Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; D007306:Insecticides; D015221:Potassium Channels; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D012964:Sodium; D007930:Leucine; D011188:Potassium