Cytotoxicity of Bacillus thuringiensis Cry1Ab toxin depends on specific binding of the toxin to the cadherin receptor BT-R1 expressed in insect cells

Abstract

The specific role of cadherin receptors in cytotoxicity involving Cry toxins of Bacillus thuringiensis and their interactions with cell membrane has not been defined. To elucidate the involvement of toxin-membrane and toxinreceptor interactions in cytotoxicity, we established a cellbased system utilizing High Five insect cells stably expressing BT-R 1 , the cadherin receptor for Cry1Ab toxin. Cry1Ab toxin is incorporated into cell membrane in both oligomeric and monomeric form. Monomeric toxin binds specifically to BT-R 1 whereas incorporation of oligomeric toxin is nonspecific and lipid dependent. Toxin oligomers in the cell membrane do not produce lytic pores and do not kill insect cells. Rather, cell death correlates with binding of the Cry1Ab toxin monomer to BT-R 1 , which apparently activates a Mg 2 þ -dependent cellular signaling pathway.

A mechanism of cell death involving an adenylyl cyclase/PKA signaling pathway is induced by the Cry1Ab toxin of Bacillus thuringiensis

“…Cultured High Five (H5) cells, which originated from ovarian cells of the cabbage looper, Trichoplusia ni, were not affected by the Cry1Ab toxin because they do not express a receptor for the toxin. However, heterologous expression of the cadherin receptor BT-R 1 in transfected H5 cells (S5) rendered them susceptible to the toxin (14). In the present work, we examine the morphological changes associated with toxin-treated S5 cells.…”

“…For example, both the number of Cry toxin binding receptors and the affinity of toxin to receptor were similar in the brush border membrane vesicles isolated from resistant and susceptible European corn borer larvae (15). Removing Mg 2ϩ by EDTA completely blocks Cry1Ab toxininduced cell death of cabbage looper cells but does not interfere with toxin-receptor binding (14). Evidently, the interaction of toxin with the receptor is prerequisite, but not sufficient, to induce cell death.…”

“…However, studies of mutated Cry toxin proteins have shown that neither the toxin oligomer complex nor commensurate changes in membrane vesicle permeability correlate directly with toxicity (17)(18)(19). Interestingly, Cry toxin oligomers also are incorporated into the cell membrane of nonsusceptible cabbage looper cells and are carried by the cells for several generations with no adverse effect (14). Apparently, toxin action is much more complicated than simply osmotic lysis.…”

“…The specificity of Cry toxins depends on individual cell surface receptors, which represent a family of cadherins expressed in the midgut epithelium of various insects (9)(10)(11)(12)(13). Cytotoxicity and cell death are the direct result of univalent binding of a Cry toxin monomer to its respective cadherin receptor (14). Impeding the toxin-receptor interaction by receptor modification has been linked to the development of resistance to Cry toxins (12).…”

“…Apparently, toxin action is much more complicated than simply osmotic lysis. Recently, we discovered that the Cry1Ab toxin produced by Bt kills insect cells by activating a Mg 2ϩ -dependent cytotoxic event upon binding of the toxin to its receptor BT-R 1 (14). Here, we report that such binding provokes cell death in insect cells by activating a previously undescribed signaling pathway involving stimulation of the stimulatory G protein ␣-subunit (G ␣s ) and adenylyl cyclase (AC), increased cyclic adenosine monophosphate (cAMP) levels, and activation of protein kinase A (PKA).…”