02767nas a2200301   4500008004600000245012100046210006900167300001400236490000700250520166600257653001201923653002801935653004901963653004302012653005602055653004102111653004802152653002902200653004502229653003802274653002202312100001302334700001102347700001602358700002102374700002002395856005002415       Submitted                   eng d     00aTissue specificity of a baculovirus-expressed, basement membrane-degrading protease in larvae of Heliothis virescens0 aTissue specificity of a baculovirusexpressed basement membranede  a431 - 4430 v393 aScathL is a cathepsin L-like cysteine protease from the flesh fly, Sarcophaga peregrina, which digests components of the basement membrane during insect metamorphosis. A recombinant baculovirus (AcMLF9.ScathL) expressing ScathL kills larvae of the tobacco budworm Heliothis virescens significantly faster than the wild type virus and triggers melanization and tissue fragmentation shortly before death. The tissue fragmentation was assumed to be a direct consequence of basement membrane degradation by ScathL. The goal of this study was to investigate the tissue specificity of ScathL when expressed by AcMLF9.ScathL using light, transmission and scanning electron microscopy. Baculovirus expression of ScathL resulted in damage to the basement membrane overlying the midgut, fat body and muscle fibers in larvae infected with AcMLF9.ScathL, but not in larvae infected with the control virus AcMLF9.ScathL.C146A or wild type virus AcMNPV C6. Injection of recombinant ScathL and high levels of baculovirus-mediated expression of ScathL resulted in complete loss of the gut. Extensive damage to the basement membrane mediated by ScathL likely resulted in loss of viability of the underlying tissue and subsequent death of the insect. These results confirm the conclusion of an earlier study (Philip, J.M.D., Fitches, E., Harrison, R.L., Bonning, B.C., Gatehouse, J.A., 2007. Characterisation of functional and insecticidal properties of a recombinant cathepsin L-like proteinase from flesh fly (Sarcophaga peregrina), which plays a role in differentiation of imaginal discs. Insect Biochem. Mol. Biol. 37, 589-600) of the remarkable specificity of this protease.10aAnimals10aBaculoviridae/*genetics10aBasement Membrane/*metabolism/ultrastructure10adevelopment/*metabolism/ultrastructure10aDigestive System/cytology/metabolism/ultrastructure10aEndopeptidases/*genetics/*metabolism10aFat Body/cytology/metabolism/ultrastructure10aInsecticides/*metabolism10aLarva/cytology/metabolism/ultrastructure10aLepidoptera/cytology/*growth &amp10aOrgan Specificity1 aTang, H.1 aLi, H.1 aLei, S., M.1 aHarrison, R., L.1 aBonning, B., C.  uhttps://sarcophagidae.myspecies.info/node/471