To Örebro University

During infection and eclipse time, Flaviviruses induce invagination of the endoplasmic reticulum (ER) membrane to form compartments, protecting their viral replication complex. The rearrangements of ER membrane require modifications in ER membrane lipid constituents or binding of proteins to bend the membrane. Indeed, it has been implicated that both KUNV and DENV NS1, NS2A, NS4A, NS4B proteins could induce membrane remodelings. However, it is not well known whether host proteins can also participate in the formation and maintenance of these compartments.In this project, we aimed to identify host proteins interacting with Kunjin, Langat, Zika replication complex. These proteins may function for ER invagination during Flavivirus infection. We used human adenocarcinoma epithelial A549 cells as a cell model, mosquito-borne Zika, Kunjin virus, and tick-borne Langat virus as virus models. After virus infections, the ER membranes from infected and non-infected cells were harvested using ultracentrifuge with a sucrose gradient. Proteins from these ERs were identified using mass spectrometry. We compared the differences between the ER proteomes to identify host candidate proteins that can cause the RC formation. To narrows the list of true candidate proteins, we attempted to enrich the RC-containing fractions by doing co-immuno precipitation. We are doing TMT-MS to identify and quantify the host proteins from Co-IP elutions. The functions of these proteins will be characterized by using molecular techniques.