Anopheles gambiae cadherin AgCad1 binds the Cry4Ba toxin of Bacillus thuringiensis israelensis and a fragment of AgCad1 synergizes toxicity.
A midgut cadherin AgCad1 cDNA was cloned from Anopheles gambiae larvae and analyzed for its possible role as a receptor for the Cry4Ba toxin of Bacillus thuringiensis strain israelensis. The AgCad1 cadherin encodes a putative 1735-residue protein organized into an extracellular region of 11 cadherin repeats (CR) and a membrane-proximal extracellular domain (MPED). AgCad1 mRNA was detected in midgut of larvae by polymerase chain reaction (PCR). The AgCad1 protein was localized, by immunochemistry of sectioned larvae, predominately to the microvilli in posterior midgut. The localization of Cry4Ba binding was determined by the same technique, and toxin bound microvilli in posterior midgut. The AgCad1 protein was present in brush border membrane fractions prepared from larvae, and Cry4Ba toxin bound the same-sized protein on blots of those fractions. The AgCad1 protein was expressed transiently in Drosophila melanogaster Schneider 2 (S2) cells. 125I-Cry4Ba toxin bound AgCad1 from S2 cells in a competitive manner. Cry4Ba bound to beads extracted 200 kDa AgCad1 and a 29 kDa fragment of AgCad1 from S2 cells. A peptide containing the AgCad1 region proximal to the cell (CR11-MPED) was expressed in Escherichia coli. Although Cry4Ba showed limited binding to CR11-MPED, the peptide synergized the toxicity of Cry4Ba to larvae. AgCad1 in the larval brush border is a binding protein for Cry4Ba toxin. On the basis of binding results and CR11-MPED synergism of Cry4Ba toxicity, AgCad1 is probably a Cry4Ba receptor.
Hua
Gang
G
Department of Entomology, University of Georgia, Athens, Georgia 30602-2603, USA.
Zhang
Rui
R
Abdullah
Mohd Amir F
MA
Adang
Michael J
MJ
D000595:Amino Acid Sequence; D000818:Animals; D000852:Anopheles / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D001413:Bacillus thuringiensis / Q000378:metabolism*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000378:metabolism*; D001427:Bacterial Toxins / Q000378:metabolism*; D015820:Cadherins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism* / Q000633:toxicity*; D003001:Cloning, Molecular; D004064:Digestive System / Q000378:metabolism; D004331:Drosophila melanogaster; D004731:Endotoxins / Q000378:metabolism*; D005786:Gene Expression Regulation; D006460:Hemolysin Proteins / Q000378:metabolism*; D007814:Larva / Q000187:drug effects; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000378:metabolism* / Q000633:toxicity*; D011485:Protein Binding
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D015820:Cadherins; D004731:Endotoxins; D006460:Hemolysin Proteins; D010446:Peptide Fragments; C030956:insecticidal crystal protein, Bacillus Thuringiensis