The mode of action of Cry toxins has been described principally in lepidopt
eran insects as a multistep process. In this work we describe the mode of a
ction of a Cry toxin active in the common pine sawfly Diprion pini (Hymenop
tera, Diprionidae). considered a major forest pest in Europe. Strain PS86Q3
contains a long bipyramidal crystal composed of five major proteins. The N
-terminal sequence shows that the 155 kDa protein corresponds to Cry5B toxi
n and the other proteins belong to the Cry5A subgroup. PCR analysis indicat
es the presence of cry5Ac and cry5Ba genes, suggesting that Cry5A protein s
hould be Cry5Ac. Activation of protoxins with trypsin or with midgut conten
t from D. pini and Cephacia abietis (Hymenoptera, Pamphiliidae) (spruce web
spinning sawfly), another important hymenopteran forest pest, produced a si
ngle 75 kDa toxin that corresponded to Cry5A by N-terminal sequence and is
responsible for the insecticidal activity. Homologous competition experimen
ts with D. pini and C. abietis brush border membrane vesicles (BBMV) showed
that the binding interaction of Cry5A is specific. Membrane potential meas
urements using a fluorescent dye indicate that Cry5A toxin at nM concentrat
ion caused immediate permeability changes in the BBMV isolated from both hy
menopteran larvae. The initial response and the sustained permeability chan
ge are cationic as previously shown for Cry1 toxins. These results indicate
that the hymenopteran specific Cry5A toxin exerts toxicity by a similar me
chanism as Cry1 toxins. (C) 2001 Elsevier Science Ltd. All rights reserved.