BST2/Tetherin is an interferon-induced anti-viral host protein that restricts the release of newly assembled virions of enveloped viruses at the plasma membrane (PM) of infected cells. It also signals to activate NF-κB in a manner that is dependent on the tyrosine 6 residue of its cytoplasmic tail domain (CD). The main goal of this dissertation is to understand the role of BST2 cytoplasmic tail in signaling and trafficking and how BST2 signaling is modulated in response to proteins of Ebola virus.
I first asked whether the cytoplasmic domain (CD) of BST2 directly binds to the host μ1 protein of the AP1 trafficking complex. Vpu of HIV-1 also binds to this complex and traps BST2 within internal membranes leading to enhanced virion release. I showed, using yeast two-hybrid assays, that the BST2 CD interacts with μ1 via a YxYxxV motif within BST2, in which both Ys and the V are required. Next, I asked whether both tyrosine residues are required for the activation of NF-κB by BST2. I found that tyrosine 6, but not tyrosine 8, is required for signaling likely due to its phosphorylation by Syk kinase. These results indicate that the determinants of membrane sorting and signaling by BST2 CD are overlapping but genetically separable.
To understand the relationship between virion-entrapment and activation of NF- κB by BST2, I studied the modulation of BST2-mediated NF-κB activation in response to the expression of the Ebola virus (EBOV) matrix protein (VP40) and glycoprotein (GP1,2). I asked whether the relief of virus-like particle (VLP) restriction by EBOV GP1,2 would reduce or inhibit BST2 signaling. VP40 was used as a virion model because it forms VLPs that resemble actual virions of EBOV, whose release is restricted by BST2. Under the conditions of this study, I found that GP1,2 and VP40 cooperated with BST2 to induce NF-κB activity. Using a mutant of BST2 lacking the C-terminal GPI anchor, which is defective for virion-entrapment and is not expressed at the PM, I found that BST2 signaling does not require trapping VLPs at the PM and that cooperative signaling with EBOV proteins occurs within internal membranes.