Infectious entry by amphotropic as well as ecotropic murine leukemia viruses occurs through an endocytic pathway

Infectious entry of enveloped viruses is thought to proceed by one of two m echanisms. pH-dependent viruses enter the cells by receptor-mediated endocy tosis and are inhibited by transient treatment with agents that prevent aci dification of vesicles in the endocytic pathway, while pa-independent virus es are not inhibited by such agents and are thought to enter the cell by di rect fusion with the plasma membrane. Nearly all retroviruses, including am photropic murine leukemia virus (MuLV) and human immunodeficiency virus typ e 1, are classified as pH independent. However, ecotropic MuLV is considere d to be a pH-dependent virus. We have examined the infectious entry of ecot ropic and amphotropic MuLVs and found that they were equally inhibited by N H,CI and bafilomycin A. These agents inhibited both viruses only partially over the course of the experiments, Agents that block the acidification of endocytic vesicles also arrest vesicular trafficking. Thus, partial inhibit ion of the MuLVs could be the result of virus inactivation during arrest in this pathway. In support of this contention, we found that that the loss o f infectivity of the MuLVs during treatment of target cells with the drugs closely corresponded to the loss of activity due to spontaneous inactivatio n at 37 degreesC in the same period of time. Furthermore, the drugs had no effect on the efficiency of infection under conditions in which the duratio n of infection was held to a very short period to minimize the effects of s pontaneous inactivation. These results indicate that the infectious process es of both ecotropic and amphotropic MuLVs were arrested rather than aborte d by transient treatment of the cells with the drugs. We also found that in fectious viruses were efficiently internalized during treatment. This indic ated that the arrest occurred in an intracellular compartment and that the infectious process of both the amphotropic and ecotropic MuLVs very likely involved endocytosis. An important aspect of this study pertains to the int erpretation of experiments in which agents that block endocytic acidificati on inhibit infectivity. As we have found with the MuLVs, inhibition of infe ctivity may be secondary to the block of endocytic acidification, While thi s strongly suggests the involvement of an endocytic pathway, it does not ne cessarily indicate a requirement for an acidic compartment during the infec tious process. Likewise, a lack of inhibition during transient treatment wi th the drugs would not preclude an endocytic pathway for viruses that are s table during the course of the treatment.