A recent study in the journal iScience reports that cell surface vimentin functions as a co-receptor for the entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) into host cells to facilitate infection. Thus, targeting vimentin may provide a unique strategy to prevent coronavirus disease 2019 (COVID-19).
Study: Vimentin is an important ACE2 co-receptor for SARS-COV-2 in epithelial cells. Image credit: Design_Cells / Shutterstock.com
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The emergence and rapid spread of SARS-CoV-2 has significantly affected global health and the economy. As of November 2, 2022, SARS-CoV-2 has infected more than 636 million worldwide and claimed more than 6.59 million lives.
Entry of SARS-CoV-2, which is a single-stranded enveloped ribonucleic acid (RNA) virus, into host cells is a key factor for its infectivity and disease pathogenesis. To this end, SARS-CoV-2 relies primarily on its receptor-binding domain (RBD) within the spike protein to bind to the angiotensin-converting enzyme 2 (ACE2) receptor of the cell surface for viral attachment. This subsequently allows the virus to enter endosomes and fuse with the host’s lysosomal membranes.
Although COVID-19 primarily causes respiratory symptoms, ACE2 is poorly expressed throughout the respiratory tract. This suggests that additional cofactors may compensate and enable interactions between the spike protein and the host receptor.
Vimentin is a type III intermediate filament cytoskeletal protein expressed in and on the surface of various types, including fibroblasts, endothelial cells, macrophages, melanocytes, Schwann cells, and lymphocytes. More recently, researchers have reported that vimentin appears to interact with the SARS-CoV-2 spike protein.
About the study
In the current study, the researchers used a variety of epithelial cell lines, including Vero E6, human colonic epithelial cells (Caco-2), and human alveolar basal epithelial cells (A549), to determine the ‘expression of ACE2, TMPRSS288 and vimentin. . In the next step, the researchers assessed whether the expression levels of vimentin and ACE2 in Vero E6, Caco-2 and A549 cells were related to the entry of SARS-CoV-2 into the cells.
The researchers also examined whether viral infection altered vimentin expression on the cell surface. By Western blot analysis after SARS-CoV-2 infection, cell supernatant vimentin was assessed for co-localization with SARS-CoV-2.
Vimentin was also evaluated for its ability to inhibit viral infection by pretreating cells for 30 minutes with withaferin A (WFA), which is a steroid lactone that binds and causes the aggregation of vimentin in in vitro A quantitative polymerase chain reaction (qPCR) assay was used to quantify vimentin aggregation, while immunofluorescence and confocal microscopy provided visualization of this process.
Results of the study
Epithelial cells were found to express variable amounts of vimentin and ACE2. Vero E6 cells, for example, expressed higher levels of ACE2 compared to Caco-2 and A549 cells. In contrast, both Vero E6 and Caco-2 cells expressed similar levels of TMPRSS2 that were significantly higher than the expression of this cell surface protein in A549 cells.
Vimentin was expressed at very low levels in Caco-2 cells compared to Vero E6 and A549 cells. Notably, Vero E6 cells showed a higher level of vimentin expression compared to A549 cells; however, cell surface expression of vimentin was higher in A549 cells.
After SARS-CoV-2 infection, viral uptake at Vero E6 levels was greatest between ten minutes and two hours. Although uptake of SARS-CoV-2 was similarly high in Caco-2 cells at ten minutes, these levels did not increase over the course of two hours. Uptake of SARS-CoV-2 was much lower in A549 cells.
Taken together, these findings suggest that cell surface vimentin expression may promote increased SARS-CoV-2 uptake in Vero E6 cells compared to the other epithelial cell lines. Further analysis of SARS-CoV-2 infection in Vero E6 cells revealed that vimentin expression increased on the cell surface after two and thirty minutes.
Meanwhile, there was no interaction between vimentin and ACE2 in uninfected Vero E6 cells, indicating that vimentin may function as a co-receptor for SARS-CoV-2 infection.
WFA pretreatment before SARS-CoV-2 infection in Vero E6 cells did not significantly affect viral replication in the cells. These findings demonstrate that although vimentin promotes SARS-CoV-2 cell infection, it is not essential for viral replication in cells.
However, WFA pretreatment reduced cell death in SARS-CoV-2-infected cells, suggesting that inhibition of vimentin may reduce the likelihood of virus-induced cell death. Furthermore, pretreatment with anti-vimentin reduced the expression of interleukin 6 (IL-6) by 15-fold, as well as CCL5 and CSCL10 by 10-fold, thus indicating that inhibition of vimentin can also limit the inflammatory response a SARS-CoV-2 infection.
Notably, when these experiments were repeated with the Omicron SARS-CoV-2 variant, similar results were observed. Thus, mutations within the Omicron spike protein do not appear to affect the interaction between SARS-CoV-2 and vimentin during infection.
Journal reference:
- Arrindell, J., Abou Atmeh, P., Jayet, L., et al. (2022). Vimentin is an important ACE2 co-receptor for SARS-COV-2 in epithelial cells. iciency doi:10.1016/j.isci.2022.105463