Poxviruses are a large family of DNA viruses capable of infecting and causing disease in humans. Its members include variola, the causative agent of smallpox, and monkeypox, which causes a smallpox-like disease that can be lethal. In addition to these public health issues, poxviruses are being developed and used as vaccine platforms and oncolytic agents. The prototypical poxvirus, vaccinia, is able to bind and enter, every cell type tested. Currently, an unambiguous cellular receptor has not been identified for poxviruses and represents a major gap in our knowledge of their lifecycle. The long-term goal of this project is to determine which cellular receptor(s) are capable of interacting with which of the ~30 viral proteins found on the surface of poxvirus virions so that pseudotyping and cell targeting can be achieved. Our studies will also help in understanding the vast tropism exhibited by vaccinia virus and other members of this group. The immediate goal is to develop a system to screen for cellular proteins that interact with vaccinia virus. These results will then be expanded to other members of this family (ectromelia virus and myxoma virus) to determine if they utilize the same receptors or if they have evolved to use different receptors. Our specific aims are; 1) Screen a cDNA library using yeast surface display for cellular proteins that the intracellular mature form of vaccinia virus can bind. 2) Prioritize and validate hits for vaccinia virus. Determine if ectromelia, and myxoma can bind putative receptors identified in the screen. The results obtained will identify cellular receptor(s) used by poxviruses and provide greater insight into the molecular mechanism of the broad tropism of these viruses, along with establishing a new receptor screening modality for viruses that have a broad tropism.