PerspectiveInfectious Disease

Inhibiting Ebola virus and SARS-CoV-2 entry

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Science  09 Oct 2020:
Vol. 370, Issue 6513, pp. 167-168
DOI: 10.1126/science.abe2977

The mechanisms by which cells defend against many viruses remain largely unknown. Defining these mechanisms is important not only for understanding viral pathogenesis but also for informing the development of antiviral therapeutics. The concerted efforts of antiviral factors within cells are central to host cell defense. Without these factors, the cell remains defenseless against potentially harmful pathogens. Understanding how the cell defends itself is particularly important for viruses that have the potential to affect global health, such as Ebola virus (EBOV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). On page 241 of this issue, Bruchez et al. (1) developed a transposon screening approach in a human osteosarcoma cell line to identify a mechanism by which CD74, previously only associated with antigen presentation, directly inhibits EBOV and SARS-CoV-2 entry into host cells.

Ebola virus cell entry

Normal cellular entry (left) of Ebola virus (EBOV) involves binding to cells expressing DC-SIGN (dendritic cell–specific ICAM-3–grabbing non-integrin 1) and TIM1 (T cell immunoglobulin mucin receptor 1), macropinocytosis, and cathepsin-mediated cleavage of the viral glycoproteins. Together with NPC1 (Niemann-Pick C1), glycoprotein cleavage allows fusion with endosomal membranes and genome release into the cytoplasm. However, CIITA (class II major histocompatibility complex transactivator) up-regulates the CD74 p41 isoform, which inhibits cathepsins and prevents genome release into the cytoplasm (right).

GRAPHIC: KELLIE HOLOSKI/SCIENCE

Viruses must gain entry into the host cell to replicate. In the case of EBOV, an enveloped virus, virions are internalized by macropinocytosis . Once virions reach endosomes, host cathepsin proteases cleave viral glycoproteins. The glycoproteins then fuse with the lysosomal membrane, which is followed by release of the viral genome into the host cell cytoplasm, where viral replication can occur (2). Thus, cathepsin-mediated cleavage is a critical step in the entry of many enveloped viruses, including EBOV, into the host cell.

Similar to EBOV, coronaviruses, including SARS-CoV-2, are enveloped viruses that undergo a series of entry steps culminating in genome release. Coronavirus entry also requires delivery of incoming viral particles to host lysosomes, where the coronavirus spike protein is cleaved by cathepsins to facilitate fusion between virus and host membranes (3, 4). However, in contrast to EBOV, SARS-CoV-2 also requires the activity of transmembrane serine protease 2 (TMPRSS2) to prime the viral spike protein (5). Thus, despite their differences in size and shape, EBOV and SARS-CoV-2 rely on similar proteolytic processes to gain entry into a target cell.

Bruchez et al. used a transposon screen in which transposable elements were inserted in front of or within genes. This approach allowed for tandem gene activation and inactivation in a single screen. To identify host factors involved in EBOV infection, the authors infected these cells with EBOV and identified two main “hits,” including Niemann-Pick C1 (NPC1), an intracellular EBOV receptor that is required for entry, thus validating the approach (6). NPC1 is a cholesterol transporter in the lysosome and is essential for EBOV fusion of the glycoproteins with the lysosomal membrane and subsequent genome release.

Additionally, the authors found that activation of the transcription factor major histocompatibility complex (MHC) class II transactivator (CIITA) inhibited EBOV infection. CIITA is a nucleotide-binding oligomerization domain–like receptor (NLR). Typically, NLRs detect pathogen-associated molecular patterns (PAMPs) within the cell and trigger an intracellular antimicrobial signaling cascade leading to nuclear factor-κB (NF-κB) nuclear translocation and expression of various proinflammatory cytokines. Unlike most NLRs, CIITA acts mainly as a transcription factor to promote the expression of other genes, including serving as the master regulator of MHC gene expression. MHC presents peptides from either intracellular (MHC class I) or extracellular (MHC class II) proteins to adaptive immune cells. CIITA induces the expression of MHC class II genes to initiate antigen presentation. The authors determined that expression of CIITA was specifically associated with inhibition of cell entry by EBOV, thus defining the step of the viral life cycle that CIITA inhibits (see the figure).

Bruchez et al. identified CD74 as the CIITA-controlled host factor responsible for inhibiting EBOV entry. CD74, often called the invariant chain or Ii, is enriched in immune cell populations and associates with MHC class II. It localizes to endoplasmic reticulum (ER) membranes and facilitates MHC class II export from the ER to vesicles that fuse with the late endosome, resulting in trafficking to the cell surface (7). CD74 also blocks the peptide-binding groove so that MHC molecules do not bind peptides prior to trafficking. Thus, without CD74, MHC class II molecules are not properly processed, and antigen presentation becomes impaired.

Bruchez et al. show that the thyroglobulin domain of CD74 is required for its antiviral activity. This domain inhibits cathepsins (8). CD74 has four isoforms but only two of them, p41 and p43, have the thyroglobulin domain. The authors show that the p41 isoform is responsible for the antiviral activity of CD74 against EBOV entry and inhibits SARS-CoV-2 fusion, suggesting a broad antiviral activity of CD74 against many cathepsin-dependent viruses. These findings highlight the often shared strategies of distinct viruses that are co-opted from host cells to promote cell entry.

These findings suggest that molecules involved in antigen presentation could also possess direct antiviral activity and that other factors with defined functions may possess additional roles in antiviral immunity. CIITA activates antiviral factors that inhibit a broad range of viruses, such as human T cell leukemia virus type 2 (HTLV-2) (9), although the steps of the viral life cycle that it targets differ from those for EBOV and SARS-CoV-2. During HTLV-2 infection, CIITA acts more directly and inhibits the viral transactivator protein (TAX2), which promotes transcription of the viral genome and thus directly inhibits HTLV-2 replication (9). Some viruses have evolved mechanisms to inhibit this restriction. For example, Epstein-Barr virus (EBV), an oncogenic DNA virus, encodes Zta, a protein that directly inhibits CIITA and results in down-regulation of MHC class II molecules. This potentially allows EBV to escape recognition from the immune system (10).

The identification of host factors that could be targeted therapeutically to limit the replication of broad families of viruses may be an effective approach to combat viral-mediated disease. However, the therapeutic benefits of viral entry inhibitors are likely most effective prior to the onset of symptoms and the development of disease, given that by these stages, viral particles have already gained entry into the cell and begun to efficiently replicate.

References and Notes

Acknowledgments: A.I.W. is supported by NIH T32-AI060525 and NIH F31-AI149866, and C.B.C. is supported by NIH AI081759, AI150151, AI145828, and AI145296.
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