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Curr Microbiol (2012) 65:446454
DOI 10.1007/s00284-012-0178-4
Investigation of the Cry4BProhibitin Interaction in Aedes aegypti Cells
Atichat Kuadkitkan Duncan R. Smith
Colin Berry
Received: 20 May 2012 / Accepted: 12 June 2012 / Published online: 6 July 2012 Springer Science+Business Media, LLC 2012
Abstract Bacillus thuringiensis (Bt) produces insecticidal toxins active against insects. Cry4B, one of the major insecticidal toxins produced by Bt subsp. israelensis, is highly toxic to mosquitoes in the genus Aedes: the major vectors of dengue, yellow fever, and chikungunya. Previous work has shown that Cry4B binds to several mid-gut membrane proteins in Aedes aegypti larvae including prohibitin, a protein recently identied as a receptor that also mediates entry of dengue virus into Aedes cells. This study conrms the interaction between Cry4B and prohibitin by co-immunoprecipitation analysis and demonstrates colocalization of prohibitin and Cry4B by confocal microscopy. While activated Cry4B toxin showed high larvicidal activity, it was not cytotoxic to two Aedes cell lines, allowing determination of its effect on dengue virus infectivity in the absence of Cry4B-induced cell lysis. Pre-exposure of Aedes cells to Cry4B resulted in a signicant reduction in the number of infected cells compared to untreated cells.
Introduction
Mosquitoes of the genus Aedes are important disease vectors that transmit a large number of viruses of signicance to human populations including dengue virus (DENV), yellow fever virus, and chikungunya virus. Dengue is a signicant worldwide public health problem,
with an estimated 100 million infections occurring each year. As such, control programs against dengue are multi-pronged, and include vaccine and drug development as well as vector control strategies [1]. One method to control the insect vector is the use of Bacillus thuringiensis subsp. israelensis (Bti), which produces crystal proteins (Cry toxins) that are active in the larval gut and lead to the death of the insect.
Cry toxins are produced by B. thuringiensis (Bt) strains upon sporulation and may be toxic to insects in the orders Diptera, Lepidoptera, Coleoptera, and Hymenoptera [2]. Bti produces a range of mosquitocidal proteins [3] and among these the Cry4B toxin is highly active against mosquito larvae of the genus Aedes [4, 5]. Cry toxins are produced in an inactive precursor form and are activated through proteolytic cleavage of the precursor in the mid-gut of the...