Fig. 1. MEL-26 interacts with FIGL-1 by means of the substrate-binding MATH domain. (A) The domain structure of MEL-26 and the interaction with MEI-1, POD-1 and CUL-3 are schematically indicated. The C94Y mutation lies in the MATH domain. The numbers mark the amino acids starting from the amino-terminus (N) to the carboxy-terminus (C). (B) Bacterially expressed GST-MEL-26 and, as a control, GST alone were incubated with S. cerevisiae extracts containing HA–FIGL-1. Bound proteins were eluted and immunoblotted with antibodies against HA. 5% of the supernatant (input) was loaded for comparison with 5% of the beads. (C) The interaction between wild-type and the C94Y mutant of MEL-26 fused to the Gal4 DNA-binding domain (BD) was characterized by two-hybrid assay with FIGL-1 fused to the Gal4 activation domain (AD). The expression of the β-galactosidase reporter was analyzed by filter assay. An empty plasmid (AD-empty) was included as a control.