Challenges and Future Directions

HCMV is an opportunistic pathogen that causes diverse disease manifestations in immune deficient individuals. Intrinsic to the broad range of clinical syndromes associated with HCMV infection is the ability of the virus to infect cells of distinct and divergent developmental lineages, including epithelial, endothelial, neuronal, and blood cells. From the perspective of virus entry, this either means that the virus uses very broadly distributed cell surface molecules to facilitate the entry pathway or has distinct or overalapping or compensatory mechanisms available. It is likely that certain steps in the entry pathway may be conserved. For example, the initial attachment to the ubiquitously distributed HSPG molecules may facilitate stable attachment in most cell types. In addition, the fusion machinery responsible for the final aspect of penetration is prob ably conserved. However, considering the coding capacity of HCMV and the number of potential glycoproteins, specialized ligand-recep-tor interactions may mediate entry into distinct cell types. Similarly, functional compensation mechanisms may exist making negative phe-notypes for null mutations difficult to interpret.

To establish definitively that a cellular molecule serves as an entry facilitator, one needs to express that molecule in a cell that is resistant to HCMV entry and confer the ability to enter when the molecule is present. Similarly, if a receptor is unknown, an entry-deficient cell can be used for expression cloning, as was recently done for the herpes simplex virus (HSV) entry mediator protein (33). The irony of HCMV is that while the virus has exquisite in vitro tropism for productive replication and infection, HCMV entry is quite promiscuous. The virus has the ability to bind, enter, and initiate infection in most laboratory cell lines (23); thus, we have lacked an important technical tool to identify cellular receptors. We recently found murine L cells to be highly resistant to HCMV entry, and this cell line may prove very useful in the future for HCMV receptor biology. Another approach that may prove useful is ligand affinity purification. As individual HCMV envelope glycoproteins become characterized with respect to their ligand properties, they may lend themselves to purification tools and the receptor identified by protein sequence analysis. Finally, entry analysis in biologically targeted cell types such as endothelial cells and monocytes/ macrophages may yield important insights into the conservation and/or divergence of the HCMV entry mechanism as well as lend clues to the identity of cellular receptors.

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