Date of Award


Document Type


Degree Name

Doctor of Philosophy (PhD)


Cellular, Molecular and Biomedical Sciences

First Advisor

Markus Thali

Second Advisor

Dimitry N. Krementsov


Human Immunodeficiency Virus type 1 (HIV-1) is a retrovirus and the causative agent of Acquired Immunodeficiency Syndrome (AIDS). HIV-1 can spread through multiple modes of transmission including cell-to-cell transmission between CD4+ T cells at a transient junction known as the virological synapse (VS). The VS forms upon HIV-1 Envelope (Env) on the surface of an infected (producer) cell binding CD4 on an uninfected (target) cell. While the VS typically resolves with complete cell separation and transfer of virus particles, Env can occasionally facilitate cell-cell fusion at this site, forming a multinucleated infected cell (syncytium). Excessive syncytium formation is prevented by viral and host factors, though this subpopulation of infected cells can still comprise ~20% of all infected cells in vivo. T cell-based syncytia detected in vivo are unique from mononucleated infected cells as they contain 2-4 nuclei, can have an elongated morphology, and appear highly motile. Despite such significant presence of syncytia, little is known about how these multinucleated infected entities contribute to HIV-1 spread and pathogenesis.

During cell-to-cell transmission at the VS, viral and host factors are enriched at this site to support virus spread (reviewed in Chapter 2). This thesis focused on fusion inhibitory factors HIV-1 Gag and several host proteins, including tetraspanins, ezrin, and EWI-2. We determined that EWI-2 is recruited specifically to the producer cell side of the VS (the presynapse) where it inhibits HIV-1-induced cell-cell fusion in a dose-dependent manner (Chapter 3). Although both EWI-2 and tetraspanins are typically downregulated upon infection, both tetraspanin CD81 and EWI-2 surface levels are partially restored on HIV-1-induced CD4+ primary T cell-based syncytia compared to mononucleated infected cells.

We sought to determine whether target cells influence the surface profile upon fusion and whether the altered protein levels are maintained for the lifetime of a syncytium (Chapter 4). We demonstrated that EWI-2 surface levels on syncytia correlate with levels of the target cell population, suggesting that EWI-2 brought along by target cells at least partially restores surface expression in syncytia. Further, we determined that newly formed, “young” syncytia, have higher levels of EWI-2 than older ones, suggesting that downregulation of EWI-2 continues in syncytia. We expect that higher levels of EWI-2 on young syncytia will render them less susceptible to continued cell-cell fusion than mononucleated infected cells and may also reduce virus particle infectivity. This will be tested by analysis of a purified syncytia population to measure fusogenicity and particle infectivity relative to fusogenicity and particle infectivity of mononucleated infected cells. Those data will be included in a future manuscript.

Collectively, the work presented in this dissertation has furthered our understanding of HIV-1-induced cell-cell fusion regulation and allowed us to characterize distinct differences in protein expression between syncytia and mononucleated infected cells. These findings open the door to future investigations aimed at understanding how syncytia contribute to virus transmission and pathogenesis.



Number of Pages

212 p.

Included in

Virology Commons