The human cancer cell active toxin Cry41Aa fromBacillus thuringiensisacts like its insecticidal counterparts.
Understanding how certain protein toxins from the normally insecticidal bacteriumBacillus thuringiensis(Bt) target human cell lines has implications for both the risk assessment of products containing these toxins and potentially for cancer therapy. This understanding requires knowledge of whether the human cell active toxins work by the same mechanism as their insecticidal counterparts or by alternative ones. TheBtCry41Aa (also known as Parasporin3) toxin is structurally related to the toxins synthesised by commercially produced transgenic insect-resistant plants, with the notable exception of an additional C-terminal β-trefoil ricin domain. To better understand its mechanism of action, we developed an efficient expression system for the toxin and created mutations in regions potentially involved in the toxic mechanism. Deletion of the ricin domain did not significantly affect the activity of the toxin against the human HepG2 cell line, suggesting that this region was not responsible for the mammalian specificity of Cry41Aa. Various biochemical assays suggested that unlike some other human cell active toxins fromBtCry41Aa did not induce apoptosis, but that its mechanism of action was consistent with that of a pore-forming toxin. The toxin induced a rapid and significant decrease in metabolic activity. Adenosine triphosphate depletion, cell swelling and membrane damage were also observed. An exposed loop region believed to be involved in receptor binding of insecticidal Cry toxins was shown to be important for the activity of Cry41Aa against HepG2 cells.
School of Life Sciences, University of Sussex, Falmer, Brighton BN1 9QG, U.K.
Vidisha
V
Krishnan
School of Life Sciences, University of Sussex, Falmer, Brighton BN1 9QG, U.K.
Barbara
B
Domanska
School of Life Sciences, University of Sussex, Falmer, Brighton BN1 9QG, U.K.
Alicia
A
Elhigazi
School of Life Sciences, University of Sussex, Falmer, Brighton BN1 9QG, U.K.
Fatai
F
Afolabi
School of Life Sciences, University of Sussex, Falmer, Brighton BN1 9QG, U.K.
Michelle J
MJ
West
School of Life Sciences, University of Sussex, Falmer, Brighton BN1 9QG, U.K. n.crickmore@sussex.ac.uk.
Neil
N
Crickmore
D000255:Adenosine Triphosphate / Q000037:antagonists & inhibitors / Q000378:metabolism; D001413:Bacillus thuringiensis / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000633:toxicity*; D001427:Bacterial Toxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000633:toxicity*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000187:drug effects; D048429:Cell Size / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004731:Endotoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000633:toxicity*; D006367:HeLa Cells; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007306:Insecticides / Q000737:chemistry / Q000378:metabolism / Q000633:toxicity; D008958:Models, Molecular; D009154:Mutation; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000633:toxicity; D052899:Pore Forming Cytotoxic Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000633:toxicity*; D054730:Protein Interaction Domains and Motifs; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism / Q000633:toxicity; D040681:Structural Homology, Protein
D001426:Bacterial Proteins; D001427:Bacterial Toxins; C000620821:Cry41Aa protein, Bacillus thuringiensis; D004731:Endotoxins; D007306:Insecticides; D010446:Peptide Fragments; D052899:Pore Forming Cytotoxic Proteins; D011994:Recombinant Proteins; D000255:Adenosine Triphosphate
HepG2; parasporin; pore-forming toxin; ricin domain