| Popis: |
With over 1.8 million newly infected people each year, the worldwide HIV-1 epidemic remains an imperative challenge for public health. Recent work has demonstrated that type I interferons (IFNs) efficiently suppress HIV infection through induction of hundreds of interferon stimulated genes (ISGs). These ISGs target distinct infection stages of invading pathogens and shape innate immunity. Among these, interferon induced transmembrane proteins (IFITMs) and lymphocyte antigen 6 complex, locus E (LY6E) have been shown to differentially modulate viral infections. However, their effects on HIV are not fully understood. In my thesis work, I provided evidence in Chapter 2 showing that IFITM proteins, particularly IFITM2 and IFITM3, specifically antagonize the HIV-1 envelope glycoprotein (Env), thereby inhibiting viral infection. IFITM proteins interacted with HIV-1 Env in viral producer cells, leading to impaired Env processing and virion incorporation. Notably, the level of IFITM incorporation into HIV-1 virions did not strictly correlate with the extent of inhibition. Prolonged passage of HIV-1 in IFITM-expressing T lymphocytes led to emergence of Env mutants that overcome IFITM restriction. The ability of IFITMs to inhibit cell-to-cell infection can be extended to HIV-1 primary isolates, HIV-2 and SIVs; however, the extent of inhibition appeared to be virus-strain dependent. Overall, this study uncovers a mechanism by which IFITM proteins specifically antagonize HIV-1 Env to restrict HIV-1 infection and provides insight into the specialized role of IFITMs in HIV infection.In Chapter three of this dissertation, I examined the effects of IFITMs on HIV-1 with different co-receptor tropism. I demonstrated that overexpression of IFITMs more dramatically suppressed entry of X4 (NL4.3 and HXB2) and dual tropic (89.6) viruses compared to their effect on R5 (JRFL and AD8) viruses, regardless of cell types. However, in all cases, the inhibition potency was ranked as IFITM3>IFITM2>IFITM1. Loss of function study revealed that knockdown IFITMs with short hairpin RNAs had modest effects on IFN-mediated viral entry compared to multi-round HIV-1 replication. Overall, this work suggests that IFITM proteins do not distinguish co-receptors CCR5 and CXCR4 to inhibit HIV-1 entry and that IFN-induced IFITMs mainly act in virus-producing cell to restrict HIV-1 replication. In Chapter four of this dissertation, I showed that LY6E promotes HIV-1 infection by enhancing viral entry and gene expression. Specifically, knockdown of LY6E in human PBMCs, SupT1, and THP-1 cells diminished HIV-1 replication. Virion-cell and cell-cell fusion experiments revealed that LY6E promotes membrane fusion of the viral entry step. Interestingly, I found that the LTR-driven HIV-1 gene expression was also enhanced by LY6E, suggesting additional roles of LY6E in HIV-1 replication. HIV-1 infection induced LY6E expression in human PBMCs, concomitant with increased production of type I IFN and some classical IFN-stimulated genes (ISGs). Altogether, these results demonstrate that IFN-inducible LY6E promotes HIV-1 entry and replication, highlighting a positive regulatory role of IFN-induced proteins in HIV-1 infection. In Chapter five of this dissertation, I showed that LY6E showed a distinct phenotype in low-CD4-expresisng Jurkat cells, i.e., LY6E inhibits HIV-1 entry by downregulating CD4. LY6E was found to be co-localized with CD4 on plasma membranes and further down-modulated CD4 on cell surface; this leads to compromised viral binding and subsequent viral entry. Ectopic expression of CD4 was able to rescue the phenotype of LY6E in Jurkat cells. The CD4 dependent inhibitory phenotype of LY6E can be recapitulated in low-CD4 primary cells human Monocyte-derived macrophages (MDMs). Taken together, this study reveals that LY6E likely modulates HIV-1 infection in a cell type dependent manner, and the CD4 surface expression level determines the sensitivity of HIV to LY6E.Overall, in this thesis work, I have investigated the role of IFITMs and LY6E in HIV-1 infection and discovered some multifaceted functions of these two families of proteins in modulating the different stages of the viral life cycle. My data highlight the more complex functions of type I IFNs and ISGs in HIV infection and AIDS pathogenesis. Information obtained in this thesis provides insights into understanding of the modes of action of type I IFNs and virus-host interaction in general, and could have therapeutic implications for prevention and treatment of HIV/AIDS. |
