TRIpartite Motif (TRIM) proteins belong to a large family of proteins, many of which are inducible by type I interferon (IFN) and serve to suppress virus infection through direct interaction with viral proteins and disruption of viral replication. TRIM5α is an important cellular inhibitor of retroviruses, able to suppress virus replication in a highly host species- and virus species-specific fashion. TRIM5α functions by binding to incoming viral capsids to disrupt uncoating and reverse transcription of the viral RNA. However, TRIM5α is thought to function exclusively in the context of retrovirus infection. We previously showed that the mouse paralog of human TRIM5α, called TRIM30D, restricts infection of specific flaviviruses belonging to the tick-borne encephalitis virus (TBEV) serogroup, but not dengue virus (DENV) or West Nile virus (WNV). This finding prompted us to determine whether human TRIM5α could also function to inhibit flavivirus replication. We found that both human and Rhesus macaque TRIM5α potently inhibited replication of TBEV and closely related viruses, but not DENV, WNV or Zika virus. However, TRIM5α did not suppress events associated with virus entry as is observed for HIV-1, and instead recognized viral nonstructural proteins involved in RNA replication. The seminar will present these surprising findings as they compare and contrast between flaviviruses and HIV-1, and discuss the implications of TRIM5α-mediated restriction for flavivirus evolution and emergence in humans.