UVM Theses and Dissertations
Format:
Print
Author:
Weng, Jia
Dept./Program:
Microbiology and Molecular Genetics
Year:
2010
Degree:
PhD
Abstract:
As human immunodeficiency virus (HIV) is etiologically linked to the acquired immunodeficiency syndrome (AIDS), its life cycle is of great interest to both clinical practitioners and researchers in cell biology and immunology. In order for this virus to gain entry into host cells, the viral envelope must fuse with the target cell membrane. The membrane fusion is driven by the viral envelope glycoprotein (Env) on the virus particle. Upon its expression and trafficking to the surface of the newly infected cells, Env is also capable of inducing cell-cell fusion, leading to the formation of syncytia. While syncytia formation can be readily observed in vitro, it is rarely, if at all, seen in vivo, i.e. in infected patients. The mechanisms remain unclear, though work from different investigators suggests that both viral and cellular factors are involved in the regulation of syncytia formation. With this thesis I present and discuss results of our research which identified tetraspanins, a group of cellular membrane proteins, as regulators of virus-induced fusion processes.
In the first part of the thesis, we tested if tetraspanins, well known regulators of cellular fusion processes (such as sperm-egg fusion) that are enriched at HIV-1 exit sites, regulate syncytium formation. We found that overexpression of tetraspanins in HIV producer cells leads to reduced syncytium formation, while downregulation has the opposite effect. Further, we document that repression of Env-induced cell-cell fusion by tetraspanins depends on the presence of viral Gag, and we demonstrate that fusion repression requires the recruitment of Env by Gag to tetraspanin enriched microdomains (TEMs). However, sensitivity to fusion repression by tetraspanins varies for different viral strains, despite comparable recruitment of their Envs to TEMs. These data have been published in The Journal of Virology in 2009, and that article constitutes chapter 2 of this thesis. In the next part of this thesis I document our attempts to further characterize the underlying mechanisms for the fusion repression mediated by tetraspanins. We initiated two lines of investigations to try to elucidate A.) which part of tetraspanins is required for fusion repression, and S.) which step of the fusion process is inhibited by tetraspanins. The results and a discussion of these analyses are presented in chapter 3 as a draft of a manuscript that we intend to submit upon completion of additional studies.
Altogether, our investigations establish tetraspanins as negative regulators of HIV-1-induced cell-cell fusion, and they start delineating, genetically and temporally the requirements for this regulation.
In the first part of the thesis, we tested if tetraspanins, well known regulators of cellular fusion processes (such as sperm-egg fusion) that are enriched at HIV-1 exit sites, regulate syncytium formation. We found that overexpression of tetraspanins in HIV producer cells leads to reduced syncytium formation, while downregulation has the opposite effect. Further, we document that repression of Env-induced cell-cell fusion by tetraspanins depends on the presence of viral Gag, and we demonstrate that fusion repression requires the recruitment of Env by Gag to tetraspanin enriched microdomains (TEMs). However, sensitivity to fusion repression by tetraspanins varies for different viral strains, despite comparable recruitment of their Envs to TEMs. These data have been published in The Journal of Virology in 2009, and that article constitutes chapter 2 of this thesis. In the next part of this thesis I document our attempts to further characterize the underlying mechanisms for the fusion repression mediated by tetraspanins. We initiated two lines of investigations to try to elucidate A.) which part of tetraspanins is required for fusion repression, and S.) which step of the fusion process is inhibited by tetraspanins. The results and a discussion of these analyses are presented in chapter 3 as a draft of a manuscript that we intend to submit upon completion of additional studies.
Altogether, our investigations establish tetraspanins as negative regulators of HIV-1-induced cell-cell fusion, and they start delineating, genetically and temporally the requirements for this regulation.