PRRSV-macrophage interaction and putative receptors

Abstract - Porcine reproductive and respiratory syndrome virus (PRRSV), a member of the Arteriviridae family, has a restricted cell tropism in its host, the pig. Mainly cells of the monocyte/macrophage (mø) lineage are susceptible, which are situated in different locations such as the tonsils, lungs, lymph nodes and spleen. The observation that several subpopulations of this cell lineage do not permit a complete replication is important. Blood monocytes, peritoneal macrophages and progenitor cells in the bone marrow are examples of refractory mø. The basis for the restricted replication of PRRSV in certain cell types can be found in the different subsequent steps which have to be gone through successfully before a new virus is produced: attachment, entry, release of the genome, transcription, translation and assembly. The inhibition of one of these steps will result in a non-permissive status of the cell. The early events of the replication cycle have already been studied extensively and will be discussed. The entry of PRRSV starts with attachment. Binding of PRRSV to alveolar macrophages happens in a dose dependent way and is completed within one hour at . The number of binding sites on the surface of a single macrophage is quite variable but exceeds 10⁴ per cell. Recently, two monoclonal antibodies were produced which are able to partially block virus attachment to the plasma membrane and to inhibit infection. They recognise a 210 kDa protein. Since a clear colocalisation exists between attached virus and aggregates of this protein on the plasma membrane, it was concluded that these monoclonal antibodies either specifically recognise the PRRSV receptor or a protein which is in close contact with it. The fact that all PRRSV-positive cells in lungs and lymphoid tissues carry this 210 kDa protein gives further evidence that monoclonal antibodies are directed against a putative receptor. Further research is currently underway to provide definitive proof that the 210 kDa protein corresponds to the PRRSV receptor. The next step in the entry-process is the endocytosis of PRRSV. This has been visualised by confocal microscopy using labelled virus. Within one hour, bound virus is taken up by the cell. Since a clear colocalisation exists between the virus and clathrin during engulfment, it can be concluded that clathrin is involved. The fact that colocalisation is no longer present once the virus is internalised provides evidence that clathrin is released from the membrane of the endosomes during the movement of the virus-containing endosome inside the cell. This finding is in agreement with the knowledge that clathrin is recycled during clathrin-dependent, receptor-mediated endocytosis processes. Although attachment and endocytosis of PRRSV is observed in almost all alveolar macrophages, viral antigens are only produced in 30% of these cells. This discrepancy suggests that one of the next steps in the replication cycle, which is the release of the viral genome in the cytoplasm or translation, is blocked in one way or another. How the viral genome gets free from its envelope and arrives in the cytoplasm for transcription and translation has not yet been clarified. It has, however, already been shown by the use of the acidotropic agents, NH ₄Cl and chloroquine, that a low pH phase is essential during this period. It is now believed, in relation to the knowledge of other viruses, which need a low pH-phase during early replication, that acidification causes conformational changes in envelope glycoproteins and/or capsid structures and triggers membrane fusion. In conclusion, it can be stated that during the early stages of PRRSV-replication, a complex cascade of events takes place and that the non-permissive status of a cell may be caused by a block at the level of one of these events.