The development of resistance to insecticides has become a classic exemplar of evolution occurring within human time scales. In this study we demonstrate how resistance to DDT in the major African malaria vector Anopheles gambiae is a result of both target-site resistance mechanisms that have introgressed between incipient species (the M- and S-molecular forms) and allelic variants in a DDT-detoxifying enzyme. Sequencing of the detoxification enzyme, Gste2, from DDT resistant and susceptible strains of An. gambiae, revealed a non-synonymous polymorphism (I114T), proximal to the DDT binding domain, which segregated with strain phenotype. Recombinant protein expression and DDT metabolism analysis revealed that the proteins from the susceptible strain lost activity at higher DDT concentrations, characteristic of substrate inhibition. The effect of I114T on GSTE2 protein structure was explored through X-ray crystallography. The amino acid exchange in the DDT-resistant strain introduced a hydroxyl group nearby the hydrophobic DDT-binding region. The exchange does not result in structural alterations but is predicted to facilitate local dynamics and enzyme activity. Expression of both wild-type and 114T alleles the allele in Drosophila conferred an increase in DDT tolerance. The 114T mutation was significantly associated with DDT resistance in wild caught M-form populations and acts in concert with target-site mutations in the voltage gated sodium channel (Vgsc-1575Y and Vgsc-1014F) to confer extreme levels of DDT resistance in wild caught An. gambiae.

No abstract

No abstract

No abstract

The superinvasive phenotype exhibited by paclitaxel-selected variants of an in vitro invasive clonal population of the human cancer cell line, MDA-MB-435S were examined using DIGE (Fluorescence 2-D Difference Gel Electrophoresis) and mass spectrometry. Isolation of membrane proteins from the MDA-MB-435S-F/Taxol-10p4p and parental populations was performed by temperature-dependent phase partitioning using the detergent Triton X-114. Subsequent DIGE-generated data analysed using Decyder software showed many differentially-expressed proteins in the membrane fraction. 16 proteins showing statistically significant upregulation in the superinvasive cells were identified by MALDI-ToF. Proteins upregulated in the superinvasive population include Galectin-3, Cofilin, ATP synthase beta subunit, voltage-dependent anion channel 1, voltage dependent anion channel 2, ER-60 protein, MHC class II antigen DR52, Beta actin, TOMM40 protein, Enolase 1, Prohibitin, Guanine nucleotidebinding protein, Annexin II, Heat shock 70 kDa protein, Stomatin-like protein 2 and Chaperonin. Many of these proteins are associated with inhibition of apoptosis, the progression of cancer, tumourigenicity, metastasis, actin remodelling at the leading edge of cells, polarized cell growth, endocytosis, phagocytosis, cellular activation, cytokinesis, and pathogen intracellular motility. These results suggest a correlation between the increased abundance of these proteins with the superinvasive phenotype of the paclitaxel-selected MDA-MB-435S-F/Taxol-10p4p population.