Tropism of human cytomegalovirus for endothelial cells is determined by a post-entry step dependent on efficient translocation to the nucleus.
Marked interstrain differences in the endothelial cell (EC) tropism of human cytomegalovirus (HCMV) isolates have been described. This study aimed to define the step during the replicative cycle of HCMV that determines this phenotype. The infection efficiency of various HCMV strains in EC versus fibroblasts was quantified by immunodetection of immediate early (IE), early and late viral antigens. Adsorption and penetration were analysed by radiolabelled virus binding assays and competitive HCMV-DNA-PCR. The translocation of penetrated viral DNA to the nucleus of infected cells was quantified by competitive HCMV-DNA-PCR in pure nuclear fractions. The intracytoplasmic translocation of capsids that had penetrated was followed by immunostaining of virus particles on a single particle level; this was correlated with the initiation of viral gene expression by simultaneous immunostaining of viral IE antigens. The infectivity of nonendotheliotropic HCMV strains in EC was found to be 100-1000-fold lower when compared to endotheliotropic strains. The manifestation of this phenotype at the level of IE gene expression indicated the importance of initial replication events. Surprisingly, no interstrain differences were detected during virus entry. However, dramatic interstrain differences were found regarding the nuclear translocation of penetrated viral DNA. With nonendotheliotropic strains, the content of viral DNA in the cell nucleus was 100-1000-fold lower in EC when compared to endotheliotropic strains, thereby reflecting the strain differences in IE gene expression. Simultaneous staining of viral particles and viral IE antigen revealed that interstrain differences in the transport of penetrated capsids towards the nucleus of endothelial cells determine the EC tropism of HCMV.
Sinzger
C
C
Department of Medical Virology1, and Department of Molecular Pathology2, University of Tübingen, D-72076 Tübingen, Germany.
Kahl
M
M
Department of Medical Virology1, and Department of Molecular Pathology2, University of Tübingen, D-72076 Tübingen, Germany.
Laib
K
K
Department of Medical Virology1, and Department of Molecular Pathology2, University of Tübingen, D-72076 Tübingen, Germany.
Klingel
K
K
Department of Medical Virology1, and Department of Molecular Pathology2, University of Tübingen, D-72076 Tübingen, Germany.
Rieger
P
P
Department of Pathology, University of Heidelberg, D-69120 Heidelberg, Germany3.
Plachter
B
B
Institute of Virology, University of Mainz, D-55101 Mainz, Germany4.
Jahn
G
G
Department of Medical Virology1, and Department of Molecular Pathology2, University of Tübingen, D-72076 Tübingen, Germany.
D021581:Active Transport, Cell Nucleus; D000327:Adsorption; D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000956:Antigens, Viral / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure / Q000821:virology*; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure; D004279:DNA, Viral / Q000378:metabolism; D004727:Endothelium / Q000166:cytology* / Q000378:metabolism / Q000648:ultrastructure / Q000821:virology*; D005347:Fibroblasts; D017781:Genes, Immediate-Early / Q000235:genetics; D014644:Genetic Variation / Q000235:genetics; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism; D008562:Membrane Glycoproteins; D008854:Microscopy, Electron; D009928:Organ Specificity; D011401:Promoter Regions, Genetic / Q000235:genetics; D015534:Trans-Activators; D014162:Transfection; D014471:Umbilical Veins; D014759:Viral Envelope Proteins; D014764:Viral Proteins; D014779:Virus Replication
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000956:Antigens, Viral; D004279:DNA, Viral; C088645:IE1 protein, cytomegalovirus; C069186:IE2 protein, Cytomegalovirus; D017874:Immediate-Early Proteins; D008562:Membrane Glycoproteins; D015534:Trans-Activators; C077406:UL115 protein, Human herpesvirus 5; D014759:Viral Envelope Proteins; D014764:Viral Proteins; C071440:glycoprotein H, Cytomegalovirus; C499358:glycoprotein H, Human cytomegalovirus; C439068:glycoprotein O, cytomegalovirus; C044582:immediate-early proteins, cytomegalovirus