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- W2029126067 abstract "Ebola virus (EBOV) infects cells through entry into endosomes. It is known that endosomal cathepsins cleave the fusion protein of EBOV (GP) into its fusion-active form and that low pH increases cathespin activity. For other viruses, such as influenza, that enter endosomes, acidic pH induces the conformational changes of the fusion protein that lead to infection. The question arises: is low pH within endosomes required for EBOV infection solely because it increases the activity of cathepsins, or does low pH also induce conformational changes in GP that leads to fusion? To answer this question, we developed a cell-cell fusion system in which cathepsin activity and pH can be independently controlled. A two color video microscopy assay was used to monitor fusion: the effector cells, expressing EBOV GP, where loaded with calcein AM and the target cells where loaded with CMAC. We verified that fusion was EBOV GP mediated: a neutralizing antibody KZ52 against EBOV GP and a peptide, 3.47, that inhibits EBOV infection inhibited fusion; single and double mutations within GP that reduce infection also reduced fusion. Results of flow cytometry experiments that monitored spread of a lipid dye were in full accord with those obtained by aqueous dye spread. Quantitative comparisons for the rate of both dyes transfers strongly indicate that EBOV GP mediated pores enlarge very slowly; this was directly confirmed by patch clamp capacitance measurements. EBOV GP can be cleaved to fusion competency in the laboratory by thermolysin, without using cathepsins. Experiments in which effector cells were treated with thermolyin and an assortment of cathepsin inhibitors were used and showed that EBOV GP induced fusion does not fundamentally require acidity, but rather, it is only the cleavage of GP that requires low pH.Supported by R01 GM101539." @default.
- W2029126067 created "2016-06-24" @default.
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- W2029126067 date "2015-01-01" @default.
- W2029126067 modified "2023-10-14" @default.
- W2029126067 title "The Role of Acidic pH in Ebola Mediated Cell-Cell Fusion" @default.
- W2029126067 doi "https://doi.org/10.1016/j.bpj.2014.11.2234" @default.
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