We recently identified the cytoskeletal regulatory protein moesin as a novel gene that inhibits retroviral replication prior to reverse transcription by downregulation of stable microtubule formation. Here, we provide evidence that overexpression of ezrin, another closely related ezrin-radixin-moesin (ERM) family member, also blocks replication of both murine leukemia viruses and human immunodeficiency virus type 1 (HIV-1) in Rat2 fibroblasts before reverse transcription, while knockdown of endogenous ezrin increases the susceptibility of human cells to HIV-1 infection. Together, these results suggest that ERM proteins may be important determinants of retrovirus susceptibility through negative regulation of stable microtubule networks.