Limits...
CD26/DPP4 cell-surface expression in bat cells correlates with bat cell susceptibility to Middle East respiratory syndrome coronavirus (MERS-CoV) infection and evolution of persistent infection.

Caì Y, Yú SQ, Postnikova EN, Mazur S, Bernbaum JG, Burk R, Zhāng T, Radoshitzky SR, Müller MA, Jordan I, Bollinger L, Hensley LE, Jahrling PB, Kuhn JH - PLoS ONE (2014)

Bottom Line: Overexpression of human CD26/DPP4 receptor conferred MERS-CoV susceptibility to resistant bat cell lines.Finally, sequential passage of MERS-CoV in permissive bat cells established persistent infection with concomitant downregulation of CD26/DPP4 surface expression.Together, these results imply that bats indeed could be among the MERS-CoV host spectrum, and that cellular restriction of MERS-CoV is determined by CD26/DPP4 expression rather than by downstream restriction factors.

View Article: PubMed Central - PubMed

Affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, United States of America.

ABSTRACT
Middle East respiratory syndrome coronavirus (MERS-CoV) is a recently isolated betacoronavirus identified as the etiologic agent of a frequently fatal disease in Western Asia, Middle East respiratory syndrome. Attempts to identify the natural reservoirs of MERS-CoV have focused in part on dromedaries. Bats are also suspected to be reservoirs based on frequent detection of other betacoronaviruses in these mammals. For this study, ten distinct cell lines derived from bats of divergent species were exposed to MERS-CoV. Plaque assays, immunofluorescence assays, and transmission electron microscopy confirmed that six bat cell lines can be productively infected. We found that the susceptibility or resistance of these bat cell lines directly correlates with the presence or absence of cell surface-expressed CD26/DPP4, the functional human receptor for MERS-CoV. Human anti-CD26/DPP4 antibodies inhibited infection of susceptible bat cells in a dose-dependent manner. Overexpression of human CD26/DPP4 receptor conferred MERS-CoV susceptibility to resistant bat cell lines. Finally, sequential passage of MERS-CoV in permissive bat cells established persistent infection with concomitant downregulation of CD26/DPP4 surface expression. Together, these results imply that bats indeed could be among the MERS-CoV host spectrum, and that cellular restriction of MERS-CoV is determined by CD26/DPP4 expression rather than by downstream restriction factors.

Show MeSH

Related in: MedlinePlus

Expression of human CD26/DPP4 confers MERS-CoV susceptibility to otherwise resistant bat cells.(A) Viral yields from MERS-CoV-resistant PESU-B5L, R05T, R06E, and Tb1Lu bat cells. Cells were transfected with a plasmid expressing human CD26/DPP4 or empty control plasmid and exposed 48 h later to MERS-CoV/EMC at an MOI of 3. Supernatants were harvested at 24 h after virus exposure for quantification of virus yields by plaque assay. (B) Same experiment: representative immunofluorescence assay (IFA) images of cells stained with anti-MERS-CoV spike protein antibody (green, top) or anti-human CD26/DPP4 antibody (red, bottom). (C) Merged IFA images demonstrate colocalization of MERS-CoV spike protein and CD26/DPP4. (D). Viral yields from MERS-CoV-susceptible bat cells transfected with a plasmid expressing human CD26/DPP4 or empty control plasmid using procedures identical to resistant cells in (A) except that cells were exposed to virus 24 h after transfection. Error bars indicate the standard deviation of duplicate samples.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4237331&req=5

pone-0112060-g004: Expression of human CD26/DPP4 confers MERS-CoV susceptibility to otherwise resistant bat cells.(A) Viral yields from MERS-CoV-resistant PESU-B5L, R05T, R06E, and Tb1Lu bat cells. Cells were transfected with a plasmid expressing human CD26/DPP4 or empty control plasmid and exposed 48 h later to MERS-CoV/EMC at an MOI of 3. Supernatants were harvested at 24 h after virus exposure for quantification of virus yields by plaque assay. (B) Same experiment: representative immunofluorescence assay (IFA) images of cells stained with anti-MERS-CoV spike protein antibody (green, top) or anti-human CD26/DPP4 antibody (red, bottom). (C) Merged IFA images demonstrate colocalization of MERS-CoV spike protein and CD26/DPP4. (D). Viral yields from MERS-CoV-susceptible bat cells transfected with a plasmid expressing human CD26/DPP4 or empty control plasmid using procedures identical to resistant cells in (A) except that cells were exposed to virus 24 h after transfection. Error bars indicate the standard deviation of duplicate samples.

Mentions: To test the hypothesis whether CD26/DPP4 alone determines bat cell susceptibility to MERS-CoV infection, we transfected MERS-CoV-resistant bat cell lines (PESU-B5L, R05T, R06E, TblLu) with a human CD26/DPP4 expression plasmid and then inoculated the cells with MERS-CoV. Transient expression of human CD26/DPP4 in these bat cell lines supported MERS-CoV replication as evidenced by increased viral yields compared to those measured in the same cell line transfected with empty control plasmid (Figure 4A). These results were confirmed by IFA using the same cells stained against MERS-CoV spike protein or CD26/DPP4 (Figure 4B). The merged immunofluorescent images clearly indicate the colocalization of MERS-CoV spike protein and CD26/DPP4 (Figure 4C). Thus, the restriction of MERS-CoV infection in the resistant bat cell lines may be determined by the absence of CD26/DPP4. Overexpression of human CD26/DPP4 in already susceptible bat cell lines led to an increase in virus yield in some bat cell lines (EpoNi/22.1, HypLu/45.1, RoNi/7.1, RoNi/7.2) or to a decrease in virus yield in other susceptible bat cell lines (EiD/41.3, HypNi/1.1, Figure 4D).


CD26/DPP4 cell-surface expression in bat cells correlates with bat cell susceptibility to Middle East respiratory syndrome coronavirus (MERS-CoV) infection and evolution of persistent infection.

Caì Y, Yú SQ, Postnikova EN, Mazur S, Bernbaum JG, Burk R, Zhāng T, Radoshitzky SR, Müller MA, Jordan I, Bollinger L, Hensley LE, Jahrling PB, Kuhn JH - PLoS ONE (2014)

Expression of human CD26/DPP4 confers MERS-CoV susceptibility to otherwise resistant bat cells.(A) Viral yields from MERS-CoV-resistant PESU-B5L, R05T, R06E, and Tb1Lu bat cells. Cells were transfected with a plasmid expressing human CD26/DPP4 or empty control plasmid and exposed 48 h later to MERS-CoV/EMC at an MOI of 3. Supernatants were harvested at 24 h after virus exposure for quantification of virus yields by plaque assay. (B) Same experiment: representative immunofluorescence assay (IFA) images of cells stained with anti-MERS-CoV spike protein antibody (green, top) or anti-human CD26/DPP4 antibody (red, bottom). (C) Merged IFA images demonstrate colocalization of MERS-CoV spike protein and CD26/DPP4. (D). Viral yields from MERS-CoV-susceptible bat cells transfected with a plasmid expressing human CD26/DPP4 or empty control plasmid using procedures identical to resistant cells in (A) except that cells were exposed to virus 24 h after transfection. Error bars indicate the standard deviation of duplicate samples.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4237331&req=5

pone-0112060-g004: Expression of human CD26/DPP4 confers MERS-CoV susceptibility to otherwise resistant bat cells.(A) Viral yields from MERS-CoV-resistant PESU-B5L, R05T, R06E, and Tb1Lu bat cells. Cells were transfected with a plasmid expressing human CD26/DPP4 or empty control plasmid and exposed 48 h later to MERS-CoV/EMC at an MOI of 3. Supernatants were harvested at 24 h after virus exposure for quantification of virus yields by plaque assay. (B) Same experiment: representative immunofluorescence assay (IFA) images of cells stained with anti-MERS-CoV spike protein antibody (green, top) or anti-human CD26/DPP4 antibody (red, bottom). (C) Merged IFA images demonstrate colocalization of MERS-CoV spike protein and CD26/DPP4. (D). Viral yields from MERS-CoV-susceptible bat cells transfected with a plasmid expressing human CD26/DPP4 or empty control plasmid using procedures identical to resistant cells in (A) except that cells were exposed to virus 24 h after transfection. Error bars indicate the standard deviation of duplicate samples.
Mentions: To test the hypothesis whether CD26/DPP4 alone determines bat cell susceptibility to MERS-CoV infection, we transfected MERS-CoV-resistant bat cell lines (PESU-B5L, R05T, R06E, TblLu) with a human CD26/DPP4 expression plasmid and then inoculated the cells with MERS-CoV. Transient expression of human CD26/DPP4 in these bat cell lines supported MERS-CoV replication as evidenced by increased viral yields compared to those measured in the same cell line transfected with empty control plasmid (Figure 4A). These results were confirmed by IFA using the same cells stained against MERS-CoV spike protein or CD26/DPP4 (Figure 4B). The merged immunofluorescent images clearly indicate the colocalization of MERS-CoV spike protein and CD26/DPP4 (Figure 4C). Thus, the restriction of MERS-CoV infection in the resistant bat cell lines may be determined by the absence of CD26/DPP4. Overexpression of human CD26/DPP4 in already susceptible bat cell lines led to an increase in virus yield in some bat cell lines (EpoNi/22.1, HypLu/45.1, RoNi/7.1, RoNi/7.2) or to a decrease in virus yield in other susceptible bat cell lines (EiD/41.3, HypNi/1.1, Figure 4D).

Bottom Line: Overexpression of human CD26/DPP4 receptor conferred MERS-CoV susceptibility to resistant bat cell lines.Finally, sequential passage of MERS-CoV in permissive bat cells established persistent infection with concomitant downregulation of CD26/DPP4 surface expression.Together, these results imply that bats indeed could be among the MERS-CoV host spectrum, and that cellular restriction of MERS-CoV is determined by CD26/DPP4 expression rather than by downstream restriction factors.

View Article: PubMed Central - PubMed

Affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, United States of America.

ABSTRACT
Middle East respiratory syndrome coronavirus (MERS-CoV) is a recently isolated betacoronavirus identified as the etiologic agent of a frequently fatal disease in Western Asia, Middle East respiratory syndrome. Attempts to identify the natural reservoirs of MERS-CoV have focused in part on dromedaries. Bats are also suspected to be reservoirs based on frequent detection of other betacoronaviruses in these mammals. For this study, ten distinct cell lines derived from bats of divergent species were exposed to MERS-CoV. Plaque assays, immunofluorescence assays, and transmission electron microscopy confirmed that six bat cell lines can be productively infected. We found that the susceptibility or resistance of these bat cell lines directly correlates with the presence or absence of cell surface-expressed CD26/DPP4, the functional human receptor for MERS-CoV. Human anti-CD26/DPP4 antibodies inhibited infection of susceptible bat cells in a dose-dependent manner. Overexpression of human CD26/DPP4 receptor conferred MERS-CoV susceptibility to resistant bat cell lines. Finally, sequential passage of MERS-CoV in permissive bat cells established persistent infection with concomitant downregulation of CD26/DPP4 surface expression. Together, these results imply that bats indeed could be among the MERS-CoV host spectrum, and that cellular restriction of MERS-CoV is determined by CD26/DPP4 expression rather than by downstream restriction factors.

Show MeSH
Related in: MedlinePlus