Infection of cells with human immunodeficiency virus 1 (HIV-1) is controlled by restriction factors, host proteins that counteract a variety of steps in the life cycle of this lentivirus. These include SAMHD1, APOBEC3G and tetherin, which block reverse transcription, hypermutate viral DNA and prevent progeny virus release, respectively. These and other HIV-1 restriction factors are conserved and have clear orthologues in the mouse. This review summarises studies in knockout mice lacking HIV-1 restriction factors. In vivo experiments in such animals have not only validated in vitro data obtained from cultured cells, but have also revealed new findings about the biology of these proteins. Indeed, genetic ablation of HIV-1 restriction factors in the mouse has provided evidence that restriction factors control retroviruses and other viruses in vivo and has led to new insights into the mechanisms by which these proteins counteract infection. For example, in vivo experiments in knockout mice demonstrate that virus control exerted by restriction factors can shape adaptive immune responses. Moreover, the availability of animals lacking restriction factors opens the possibility to study the function of these proteins in other contexts such as autoimmunity and cancer. Further in vivo studies of more recently identified HIV-1 restriction factors in gene targeted mice are, therefore, justified.