FIG 4.

Bdp1 region II interacts with the Brf1 N terminus and the C128 N terminus. (A) (Top) A schematic of Brf1 indicates the N-terminal zinc ribbon, linker, and cyclin fold repeat domains as well as the C-terminal conserved sequence blocks I, II, and III. (Bottom) Western blot analysis of coimmunoprecipitation with V5 epitope-tagged Brf1 N-terminal fragments from aa 1 to 90 and aa 1 to 284 and a Bdp1 region II peptide fragment containing aa 253 to 325. Anti-V5 antibody-agarose beads were used to precipitate the V5-tagged Brf1 fragments. As indicated, either the WT Bdp1 region II peptide or the mutant peptide with triple point mutations was applied in the coimmunoprecipitation assay. (B) (Top) A schematic of the C128 N-terminal region includes the external 1, protrusion, lobe, and fork domains. (Bottom) Western blot analysis of coimmunoprecipitation with Bdp1 region II and C128 N-terminal (Nt) peptide fragments. Anti-Flag antibody-agarose beads were applied to precipitate the Flag-tagged Bdp1 peptide fragment. As indicated, either the WT Bdp1 region II fragment or the Bdp1 region II fragment with a quadruple point mutation was used. Coimmunoprecipitated polypeptides were probed with anti-Flag and anti-His antibodies, revealing the Bdp1 region and the C128 N terminus, respectively.