Mammalian cells have developed diverse strategies to restrict retroviral infection. Retroviruses have therefore evolved to counteract such restriction factors, in order to colonize their hosts. Tripartite motif-containing 5 isoform-alpha (TRIM5alpha) protein from rhesus monkey (TRIM5alpharh) restricts human immunodeficiency virus type 1 (HIV-1) infection at a postentry, preintegration stage in the viral life cycle, by recognizing the incoming capsid and promoting its premature disassembly. TRIM5alpha comprises an RBCC (RING, B-box 2 and coiled-coil motifs) domain and a B30.2(SPRY) domain. Sequences in the B30.2(SPRY) domain dictate the potency and specificity of the restriction. As TRIM5alpharh targets incoming mature HIV-1 capsid, but not precursor Gag, it was assumed that TRIM5alpharh did not affect HIV-1 production. Here we provide evidence that TRIM5alpharh, but not its human ortholog (TRIM5alphahu), blocks HIV-1 production through rapid degradation of HIV-1 Gag polyproteins. The specificity for this restriction is determined by sequences in the RBCC domain. Our observations suggest that TRIM5alpharh interacts with HIV-1 Gag during or before Gag assembly through a mechanism distinct from the well-characterized postentry restriction. This finding demonstrates a cellular factor blocking HIV-1 production by actively degrading a viral protein. Further understanding of this previously unknown restriction mechanism may reveal new targets for future anti-HIV-1 therapy.