Structures of the Phactr1 RPEL Domain and RPEL Motif Complexes with G-Actin Reveal the Molecular Basis for Actin Binding Cooperativity

The Phactr family of PP1-binding proteins and the myocardin-related transcription factor family of transcriptional coactivators contain regulatory domains comprising three copies of the RPEL motif, a G-actin binding element. We report the structure of a Phactr1 G-actin circle RPEL domain complex. Three G-actins surround the crank-shaped RPEL domain forming a closed helical assembly. Their spatial relationship is identical to the RPEL-actins within the pentavalent MRTF G-actin circle RPEL domain complex, suggesting that conserved cooperative interactions between actin.RPEL units organize the assembly. In the trivalent Phactr1 complex, each G-actin circle RPEL unit makes secondary contacts with its downstream actin involving distinct RPEL residues. Similar secondary contacts are seen in G-actin circle RPEL peptide crystals. Loss-of-secondary-contact mutations destabilize the Phactr1 G-actin.RPEL assembly. Furthermore, actin-mediated inhibition of Phactr1 nuclear import requires secondary contact residues in the Phactr1 N-terminal RPEL-N motif, suggesting that it involves interaction of RPEL-N with the C-terminal assembly. Secondary actin contacts by actin-bound RPEL motifs thus govern formation of multivalent actin circle RPEL assemblies.