Characterization of the Early Steps of Human Parvovirus B19 Infection

Abstract

The early steps of human parvovirus B19 (B19V) infection were investigated in UT7/Epo cells. B19V and its receptor globoside (Gb4Cer) associate with lipid rafts, predominantly of the noncaveolar type. Pharmacological disruption of the lipid rafts inhibited infection when the drug was added prior to virus attachment but not after virus uptake. B19V is internalized by clathrin-dependent endocytosis and spreads rapidly throughout the endocytic pathway, reaching the lysosomal compartment within minutes, where a substantial proportion is degraded. B19V did not permeabilize the endocytic vesicles, indicating a mechanism of endosomal escape without apparent membrane damage. Bafilomycin A 1 (BafA1) and NH 4 Cl, which raise endosomal pH, blocked the infection by preventing endosomal escape, resulting in a massive accumulation of capsids in the lysosomes. In contrast, in the presence of chloroquine (CQ), the transfer of incoming viruses from late endosomes to lysosomes was prevented; the viral DNA was not degraded; and the infection was boosted. In contrast to the findings for untreated or BafA1-treated cells, the viral DNA was progressively associated with the nucleus in CQ-treated cells, reaching a plateau by 3 h postinternalization, a time coinciding with the initiation of viral transcription. At this time, more than half of the total intracellular viral DNA was associated with the nucleus; however, the capsids remained extranuclear. Our studies provide the first insight into the early steps of B19V infection and reveal mechanisms involved in virus uptake, endocytic trafficking, and nuclear penetration.

Blood Groups in Infection and Host Susceptibility

“…To enhance detection, cells can be enzymatically treated with proteases or neuraminidase (360,373). Expression levels may vary between cells due to differential GSL synthesis during the cell cycle, cell activation, and differentiation (374)(375)(376)(377)(378). Fresh (or frozen) tissue should be used for immunological analysis and isolation (369).…”

“…The virus's tropism for erythroblasts makes it particularly difficult to propagate in vitro, with most investigators using CD36 ϩ erythroid progeni-tors or erythroblastic cell lines (UT7/Epo-S1 and KU812Ep6). Successful infection requires capsid binding to noncaveolar glycolipid-enriched lipid rafts, followed by clathrin-mediated endocytosis and endosomal acidification (377). B19 infection is enhanced in the presence of chloroquine, an antimalarial which destabilizes endosomal membranes (377).…”

“…Successful infection requires capsid binding to noncaveolar glycolipid-enriched lipid rafts, followed by clathrin-mediated endocytosis and endosomal acidification (377). B19 infection is enhanced in the presence of chloroquine, an antimalarial which destabilizes endosomal membranes (377). Once released, B19 inhibits erythropoiesis by inducing cell cycle arrest necessary for viral replication, followed by apoptosis and viral release (481).…”

“…Brown et al demonstrated that Gb4 is necessary for infection by showing that erythroblasts from p individuals were resistant to B19 in vitro (483). Likewise, B19 binding and infection can be blocked by disruption of lipid rafts (377). One factor that may favor erythroid cells is their high Gb4 content relative to that in other tissues (368).…”

Parvovirus B19 Achievements and Challenges

“…A first insight into the early steps of B19V infection, the path of virions, and mechanisms involved in virus uptake, trafficking, and nuclear import has recently been investigated in UT7/Epo cells [77]. In these, B19V and its receptor Gb4Cer associate with lipid rafts, predominantly of the noncaveolar type.…”