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THY-1 Cell Surface Antigen (CD90) Has an Important Role in the Initial Stage of Human Cytomegalovirus Infection.

Li Q, Wilkie AR, Weller M, Liu X, Cohen JI - PLoS Pathog. (2015)

Bottom Line: THY-1 interacted with HCMV gB and gH and may form a complex important for entry.However, since gB and gH have previously been shown to interact, it is uncertain if THY-1 directly binds to both of these proteins.THY-1 may function through a complex setting, that would include viral gB and gH, and other cellular factors, thus links virus entry with signaling in host cells that ultimately leads to virus infection.

View Article: PubMed Central - PubMed

Affiliation: Medical Virology Section, Laboratory of Infectious Diseases, National Institutes of Health, Bethesda, Maryland, United States of America.

ABSTRACT
Human cytomegalovirus (HCMV) infects about 50% of the US population, is the leading infectious cause of birth defects, and is considered the most important infectious agent in transplant recipients. The virus infects many cell types in vivo and in vitro. While previous studies have identified several cellular proteins that may function at early steps of infection in a cell type dependent manner, the mechanism of virus entry is still poorly understood. Using a computational biology approach, correlating gene expression with virus infectivity in 54 cell lines, we identified THY-1 as a putative host determinant for HCMV infection in these cells. With a series of loss-of-function, gain-of-function and protein-protein interaction analyses, we found that THY-1 mediates HCMV infection at the entry step and is important for infection that occurs at a low m.o.i. THY-1 antibody that bound to the cell surface blocked HCMV during the initial 60 minutes of infection in a dose-dependent manner. Down-regulation of THY-1 with siRNA impaired infectivity occurred during the initial 60 minutes of inoculation. Both THY-1 antibody and siRNA inhibited HCMV-induced activation of the PI3-K/Akt pathway required for entry. Soluble THY-1 protein blocked HCMV infection during, but not after, virus internalization. Expression of exogenous THY-1 enhanced entry in cells expressing low levels of the protein. THY-1 interacted with HCMV gB and gH and may form a complex important for entry. However, since gB and gH have previously been shown to interact, it is uncertain if THY-1 directly binds to both of these proteins. Prior observations that THY-1 (a) interacts with αVβ3 integrin and recruits paxillin (implicated in HCMV entry), (b) regulates leukocyte extravasation (critical for HCMV viremia), and (c) is expressed on many cells targeted for HCMV infection including epithelial and endothelial cells, fibroblast, and CD34+/CD38- stem cells, all support a role for THY-1 as an HCMV entry mediator in a cell type dependent manner. THY-1 may function through a complex setting, that would include viral gB and gH, and other cellular factors, thus links virus entry with signaling in host cells that ultimately leads to virus infection.

No MeSH data available.


Related in: MedlinePlus

Expression of THY-1 positively correlates with HCMV infectivity for both fibroblast and epithelial/endothelial tropic stains of virus in 54 human cell lines.Cells were infected with both fibroblast tropic HCMV (Towne-GFP) for 2 days and epithelial/endothelial tropic HCMV (TB40E-GFP) for 3 days. We aimed to achieve an infection rate at approximately 10–20% for positive controls without acid inactivation (m.o.i 0.5 for control MRC-5 cells and 0.1–0.5 for ARPE-19 cells). The percentage of infectivity was derived from the mean of three independent infections minus the background florescence contributed by mock-infected cells and normalized against control cell lines (MRC-5 for Towne-GFP, ARPE-19 for TB40E-GFP). THY-1 and PDGFR-α expression levels were determined by multiple microarray platforms (http://dtp.nci.nih.gov/mtweb) and these are baseline levels of RNA in cells not infected with virus. HCMV infectivity positively correlated with THY-1 expression at a level similar to that of PDGFR-α. Cell lines are indicated in S1 Table.
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ppat.1004999.g001: Expression of THY-1 positively correlates with HCMV infectivity for both fibroblast and epithelial/endothelial tropic stains of virus in 54 human cell lines.Cells were infected with both fibroblast tropic HCMV (Towne-GFP) for 2 days and epithelial/endothelial tropic HCMV (TB40E-GFP) for 3 days. We aimed to achieve an infection rate at approximately 10–20% for positive controls without acid inactivation (m.o.i 0.5 for control MRC-5 cells and 0.1–0.5 for ARPE-19 cells). The percentage of infectivity was derived from the mean of three independent infections minus the background florescence contributed by mock-infected cells and normalized against control cell lines (MRC-5 for Towne-GFP, ARPE-19 for TB40E-GFP). THY-1 and PDGFR-α expression levels were determined by multiple microarray platforms (http://dtp.nci.nih.gov/mtweb) and these are baseline levels of RNA in cells not infected with virus. HCMV infectivity positively correlated with THY-1 expression at a level similar to that of PDGFR-α. Cell lines are indicated in S1 Table.

Mentions: The highest rated membrane associated protein whose expression correlated positively with virus susceptibility was PDGFR-α, which has been shown to function in HCMV entry [14,15]. Transfection of MRC-5 cells with PDGFR-α specific siRNAs reduced HCMV infection (S1A Fig). THY-1 was implicated as the next highest scoring membrane associated protein whose expression correlated positively with HCMV infectivity. Infectivity of both epithelial/endothelial and fibroblast tropic HCMV strains showed a positive correlation with THY-1 expression at a level similar to or higher than that of PDGFR-α (Fig 1). The correlation of THY-1 expression was statistically significant for both Towne-GFP (P = 0.0002, Pearson Correlation Coefficient 0.46) and TB40E-GFP HCMV (P = 0.0004, Pearson Correlation Coefficient 0.44). Likewise, expression of PDGFR-α correlated with infection for Towne-GFP (p<0.00001, Pearson Correlation Coefficient 0.53) and TB40E-GFP HCMV (p = 0.016, Pearson Correlation Coefficient 0.29). Similar correlations for THY-1 and PDGFR-α expression with infectivity were also observed for epithelial/endothelial tropic strain BADrUl131-GFP HCMV.


THY-1 Cell Surface Antigen (CD90) Has an Important Role in the Initial Stage of Human Cytomegalovirus Infection.

Li Q, Wilkie AR, Weller M, Liu X, Cohen JI - PLoS Pathog. (2015)

Expression of THY-1 positively correlates with HCMV infectivity for both fibroblast and epithelial/endothelial tropic stains of virus in 54 human cell lines.Cells were infected with both fibroblast tropic HCMV (Towne-GFP) for 2 days and epithelial/endothelial tropic HCMV (TB40E-GFP) for 3 days. We aimed to achieve an infection rate at approximately 10–20% for positive controls without acid inactivation (m.o.i 0.5 for control MRC-5 cells and 0.1–0.5 for ARPE-19 cells). The percentage of infectivity was derived from the mean of three independent infections minus the background florescence contributed by mock-infected cells and normalized against control cell lines (MRC-5 for Towne-GFP, ARPE-19 for TB40E-GFP). THY-1 and PDGFR-α expression levels were determined by multiple microarray platforms (http://dtp.nci.nih.gov/mtweb) and these are baseline levels of RNA in cells not infected with virus. HCMV infectivity positively correlated with THY-1 expression at a level similar to that of PDGFR-α. Cell lines are indicated in S1 Table.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4492587&req=5

ppat.1004999.g001: Expression of THY-1 positively correlates with HCMV infectivity for both fibroblast and epithelial/endothelial tropic stains of virus in 54 human cell lines.Cells were infected with both fibroblast tropic HCMV (Towne-GFP) for 2 days and epithelial/endothelial tropic HCMV (TB40E-GFP) for 3 days. We aimed to achieve an infection rate at approximately 10–20% for positive controls without acid inactivation (m.o.i 0.5 for control MRC-5 cells and 0.1–0.5 for ARPE-19 cells). The percentage of infectivity was derived from the mean of three independent infections minus the background florescence contributed by mock-infected cells and normalized against control cell lines (MRC-5 for Towne-GFP, ARPE-19 for TB40E-GFP). THY-1 and PDGFR-α expression levels were determined by multiple microarray platforms (http://dtp.nci.nih.gov/mtweb) and these are baseline levels of RNA in cells not infected with virus. HCMV infectivity positively correlated with THY-1 expression at a level similar to that of PDGFR-α. Cell lines are indicated in S1 Table.
Mentions: The highest rated membrane associated protein whose expression correlated positively with virus susceptibility was PDGFR-α, which has been shown to function in HCMV entry [14,15]. Transfection of MRC-5 cells with PDGFR-α specific siRNAs reduced HCMV infection (S1A Fig). THY-1 was implicated as the next highest scoring membrane associated protein whose expression correlated positively with HCMV infectivity. Infectivity of both epithelial/endothelial and fibroblast tropic HCMV strains showed a positive correlation with THY-1 expression at a level similar to or higher than that of PDGFR-α (Fig 1). The correlation of THY-1 expression was statistically significant for both Towne-GFP (P = 0.0002, Pearson Correlation Coefficient 0.46) and TB40E-GFP HCMV (P = 0.0004, Pearson Correlation Coefficient 0.44). Likewise, expression of PDGFR-α correlated with infection for Towne-GFP (p<0.00001, Pearson Correlation Coefficient 0.53) and TB40E-GFP HCMV (p = 0.016, Pearson Correlation Coefficient 0.29). Similar correlations for THY-1 and PDGFR-α expression with infectivity were also observed for epithelial/endothelial tropic strain BADrUl131-GFP HCMV.

Bottom Line: THY-1 interacted with HCMV gB and gH and may form a complex important for entry.However, since gB and gH have previously been shown to interact, it is uncertain if THY-1 directly binds to both of these proteins.THY-1 may function through a complex setting, that would include viral gB and gH, and other cellular factors, thus links virus entry with signaling in host cells that ultimately leads to virus infection.

View Article: PubMed Central - PubMed

Affiliation: Medical Virology Section, Laboratory of Infectious Diseases, National Institutes of Health, Bethesda, Maryland, United States of America.

ABSTRACT
Human cytomegalovirus (HCMV) infects about 50% of the US population, is the leading infectious cause of birth defects, and is considered the most important infectious agent in transplant recipients. The virus infects many cell types in vivo and in vitro. While previous studies have identified several cellular proteins that may function at early steps of infection in a cell type dependent manner, the mechanism of virus entry is still poorly understood. Using a computational biology approach, correlating gene expression with virus infectivity in 54 cell lines, we identified THY-1 as a putative host determinant for HCMV infection in these cells. With a series of loss-of-function, gain-of-function and protein-protein interaction analyses, we found that THY-1 mediates HCMV infection at the entry step and is important for infection that occurs at a low m.o.i. THY-1 antibody that bound to the cell surface blocked HCMV during the initial 60 minutes of infection in a dose-dependent manner. Down-regulation of THY-1 with siRNA impaired infectivity occurred during the initial 60 minutes of inoculation. Both THY-1 antibody and siRNA inhibited HCMV-induced activation of the PI3-K/Akt pathway required for entry. Soluble THY-1 protein blocked HCMV infection during, but not after, virus internalization. Expression of exogenous THY-1 enhanced entry in cells expressing low levels of the protein. THY-1 interacted with HCMV gB and gH and may form a complex important for entry. However, since gB and gH have previously been shown to interact, it is uncertain if THY-1 directly binds to both of these proteins. Prior observations that THY-1 (a) interacts with αVβ3 integrin and recruits paxillin (implicated in HCMV entry), (b) regulates leukocyte extravasation (critical for HCMV viremia), and (c) is expressed on many cells targeted for HCMV infection including epithelial and endothelial cells, fibroblast, and CD34+/CD38- stem cells, all support a role for THY-1 as an HCMV entry mediator in a cell type dependent manner. THY-1 may function through a complex setting, that would include viral gB and gH, and other cellular factors, thus links virus entry with signaling in host cells that ultimately leads to virus infection.

No MeSH data available.


Related in: MedlinePlus