Krüppel-associated boxes are potent transcriptional repression domains
Key findings
Margolin et al. demonstrated that the KRAB domain of KOX1 functioned as a potent DNA-binding-site-dependent transcriptional repressor when fused to the GAL4 DNA-binding domain. The GAL4-KOX1-(1-161) fusion repressed a GAL-TK-CAT reporter 17-fold in NIH 3T3 cells, establishing that the KRAB domain transfers repression activity to heterologous DNA-binding contexts.
Truncation analysis defined a minimal 45-aa repression domain within the KRAB A box (residues 11–55 of KOX1). The KRAB B box was dispensable, and any deletion extending into the A box abolished repression. KRAB domains from four independent zinc finger proteins—KOX1, ZNF133, ZNF140, and ZNF141—each functioned as potent repressors when fused to GAL4.
Site-directed mutagenesis of conserved residues in the KRAB A box at DV, EEW, and MLE positions significantly inhibited transcriptional repression. The MLE sequence falls within the conserved VMLENY motif of a predicted amphipathic helix, suggesting that a protein-protein interaction surface on the KRAB A box mediates repression.