Cadherins belong to one of the families of animal glycoproteins responsible for cal-cium-dependent cell-cell adhesion.Recent literatures showed that the cadherin-like in midgut of several insects served as the recepto...Cadherins belong to one of the families of animal glycoproteins responsible for cal-cium-dependent cell-cell adhesion.Recent literatures showed that the cadherin-like in midgut of several insects served as the receptor of Bt toxin Cry1A and the variation of cadherin-like is re-lated to insect’s resistance to Cry1A.The full-length cDNA encoding cadherin-like of Helicoverpa armigera is cloned by degenerate PCR and RACE techniques and the gene was designated as BtR-harm,which is 5581 bp in full-length,encoding 1730 amino acid residues(BtR-harm was deposited in GenBank and the accession number is AF519180).Its predicted molecular weight and isoelectric point were 195.39 kDa and 4.23,respectively.The inferred amino acid sequence includes a signal sequence,11 cadherin repeats,a membrane-proximal region,a transmem-brane region and a cytoplasmic region.Sequence analysis indicated that the deduced protein sequence was most similar to the cadherin-like from Heliothis virescens with 84.2%identity and highly similar to three other lepidopteran cadherin from Bombyx mori,Manduca sexta and Pectinophora gossypiella,with the sequence identities of 60.3.6%,57.5%and 51.0%,respec-tively.The cDNA encoding cadherin gene was expressed successfully in E.coli and the recom-binant proteins can bind with Cry1Ac.Truncation analysis and binding experiment of BtR-harm revealed that the Cry1A binding region was a contiguous 244-amino acid sequence,which lo-cated between amino acid 1217 and 1461.Semi-quantitative RT-PCR analysis showed that BtR-harm was highly expressed in midgut of H.armigera,very low expressed in foregut and hindgut and was not expressed in other tissues.After H.armigera producing resistance to Cry1Ac,the expression quantity of BtR-harm significantly decreased in midgut of H.armigera.It is the first confirmation that BtR-harm can function as receptor of Cry1Ac in H.armigera and the binding region was located on a contiguous 244 amino acid sequence,suggesting that the de-crease of expression quantity of BtR-harm is one of th展开更多
基金This work was supported by the Nati onal Natual Science F oundation of China(Grant No.30300233)Chinese Ministry of ScienceTechnology(Grant Nos.G2000016208&2001AA212271)
文摘Cadherins belong to one of the families of animal glycoproteins responsible for cal-cium-dependent cell-cell adhesion.Recent literatures showed that the cadherin-like in midgut of several insects served as the receptor of Bt toxin Cry1A and the variation of cadherin-like is re-lated to insect’s resistance to Cry1A.The full-length cDNA encoding cadherin-like of Helicoverpa armigera is cloned by degenerate PCR and RACE techniques and the gene was designated as BtR-harm,which is 5581 bp in full-length,encoding 1730 amino acid residues(BtR-harm was deposited in GenBank and the accession number is AF519180).Its predicted molecular weight and isoelectric point were 195.39 kDa and 4.23,respectively.The inferred amino acid sequence includes a signal sequence,11 cadherin repeats,a membrane-proximal region,a transmem-brane region and a cytoplasmic region.Sequence analysis indicated that the deduced protein sequence was most similar to the cadherin-like from Heliothis virescens with 84.2%identity and highly similar to three other lepidopteran cadherin from Bombyx mori,Manduca sexta and Pectinophora gossypiella,with the sequence identities of 60.3.6%,57.5%and 51.0%,respec-tively.The cDNA encoding cadherin gene was expressed successfully in E.coli and the recom-binant proteins can bind with Cry1Ac.Truncation analysis and binding experiment of BtR-harm revealed that the Cry1A binding region was a contiguous 244-amino acid sequence,which lo-cated between amino acid 1217 and 1461.Semi-quantitative RT-PCR analysis showed that BtR-harm was highly expressed in midgut of H.armigera,very low expressed in foregut and hindgut and was not expressed in other tissues.After H.armigera producing resistance to Cry1Ac,the expression quantity of BtR-harm significantly decreased in midgut of H.armigera.It is the first confirmation that BtR-harm can function as receptor of Cry1Ac in H.armigera and the binding region was located on a contiguous 244 amino acid sequence,suggesting that the de-crease of expression quantity of BtR-harm is one of th
