Chikungunya virus (CHIKV) has recently re-emerged causing millions of infections in countries around the Indian Ocean. While CHIKV has a broad host cell range and productively infects a number of different cell types, macrophages have been identified as a potential viral reservoir serving to increase the duration of symptoms. To date no CHIKV interacting protein has been characterized and this study sought to identify CHIKV binding proteins expressed on target cell membranes. Two-dimensional virus overlay identified prohibitin (PHB) as a microglial cell expressed CHIKV binding protein. Co-localization, co-immunoprecipitation as well as antibody and siRNA mediated infection inhibition studies all confirmed a role for PHB in mediating internalization of CHIKV into microglial cells. PHB is the first identified CHIKV receptor protein, and this study is evidence that PHB may play a role in the internalization of multiple viruses.
Dengue is transmitted primarily by mosquitoes of the Aedes genus. Despite a number of studies, no insect dengue virus receptor protein has been clearly identified and characterized. Using a number of separation methodologies and virus overlay protein binding assays we identified a 35kDa protein that segregated with susceptibility to dengue serotype 2 (DENV-2) infection in two mosquito species and two mosquito cell lines. Mass spectroscopy identified the protein to be prohibitin, a strongly conserved and ubiquitously expressed protein in eukaryotic cells. Antibody mediated inhibition of infection and siRNA mediated knockdown of prohibitin expression significantly reduced infection levels and subsequent virus production in both Aedes aegypti and Aedes albopictus cell lines. Confocal microscopy showed a significant degree of intracellular colocalization between prohibitin and DENV-2 E protein, and coimmunoprecipitation confirmed that prohibitin interacts with dengue E. Prohibitin is the first characterized insect cell expressed dengue virus receptor protein.
Chikungunya virus (CHIKV) is a re-emerging mosquito-borne alphavirus that has recently engendered large epidemics around the world. There is no specific antiviral for treatment of patients infected with CHIKV, and development of compounds with significant anti-CHIKV activity that can be further developed to a practical therapy is urgently required. Andrographolide is derived from Andrographis paniculata, a herb traditionally used to treat a number of conditions including infections. This study sought to determine the potential of andrographolide as an inhibitor of CHIKV infection. Andrographolide showed good inhibition of CHIKV infection and reduced virus production by approximately 3log10 with a 50% effective concentration (EC50) of 77 μM without cytotoxicity. Time-of-addition and RNA transfection studies showed that andrographolide affected CHIKV replication and the activity of andrographolide was shown to be cell type independent. This study suggests that andrographolide has the potential to be developed further as an anti-CHIKV therapeutic agent.
Chikungunya virus (CHIKV), the virus responsible for the disease chikungunya fever in humans, is transmitted by Aedes mosquitoes. While significant progress has been made in understanding the process by which CHIKV enters into mammalian cells, far less progress has been made in understanding the CHIKV entry process in insect cells. This study sought to identify mosquito-cell-expressed CHIKV-binding proteins through a combination of virus overlay protein binding assays (VOPBA) and mass spectroscopy. A 50-kDa CHIKV-binding protein was identified as the ATP synthase β subunit (ATPSβ). Co-immunoprecipitation studies confirmed the interaction, and colocalization analysis showed cell-surface and intracellular co-localization between CHIKV and ATPSβ. Both antibody inhibition and siRNA-mediated downregulation experiments targeted to ATPSβ showed a significant reduction in viral entry and virus production. These results suggest that ATPSβ is a CHIKV-binding protein capable of mediating the entry of CHIKV into insect cells.
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