Thalidomide induces cereblon (CRBN)-dependent degradation of proteins. Human cytochrome P450s are thought to provide two monohydroxylated metabolites from thalidomide, and the metabolites are also considered to be involved in thalidomide effects. However, it remains unclear. We report that human PLZF/ZBTB16 is a target protein of CRBN with thalidomide and its derivatives, and that 5-hydroxythalidomide has high potential for degrading PLZF. Using a human transcription factor protein array produced by a wheat cell-free protein synthesis system, PLZF was found to bind to CRBN with thalidomide. PLZF is degraded by the CRL4CRBN complex with thalidomide and its derivatives. Mutagenesis analysis revealed that both 1st and 3rd zinc finger domains conserved in vertebrates are recognized for thalidomide-dependent binding and degradation by CRBN. In chicken limbs, knockdown of Plzf induced skeletal abnormalities, and Plzf was degraded after thalidomide or 5-hydroxythalidomide treatment. Our findings suggest that PLZF is a pivotal substrate involving thalidomide-induced teratogenesis.