Prohibitin protein characterization in dengue virus vector and non-vector mosquito species

Dengue virus is an arthropod-borne virus that causes dengue fever, dengue hemorrhagic fever, and dengue shock syndrome in tropical areas of the world.  The putative receptor for this virus in mosquito species is the prohibitin protein.  Through reverse transcription PCR, DNA sequencing, and 3’ rapid amplification of cDNA ends (RACE), we have deduced the amino acid sequence of the prohibitin proteins from a likely vector mosquito species, Ochlerotatus japonicus, and a laboratory competent mosquito, Ochlerotatus triseriatus. In a sequence alignment with Aedes aegypti, we found high amino acid similarity between both Oc. triseriatus (97.9%) and Oc. japonicus (98.9%) prohibitin sequences.  Further comparisons between vector and non-vector species has yielded six mutations in the gene sequences between the species that are consistent with vector competency.  We have expressed the Ae. aegypti prohibitin proteins and tested each for binding affinity with dengue virus using both Western blot and affinity binding columns. We will then induce point mutations corresponding to the differences in amino acid sequences and test those mutated proteins for virus binding affinity.  This may lead to the identification of a key nucleic acid change between vector and non-vector prohibitins necessary for dengue virus binding within the mosquito host. Understanding how dengue interacts with its receptor may lead to methods to block mosquito infection in order to reduce virus transmission.