Structural modeling of the interaction between GJA1-20k and actin. (A) Illustration of Cx43 and GJA1-20k featuring the translation start sites for both the GJA1-20k and GJA1-11k isoforms and the sequence of the final nine amino acids (RPRPDDLEI). (B) Modeling of GJA1-20k (PDB ID: 1R5S) docked to actin. In tan: GJA1-20k ATTRACT models docked using F-actin (PDB ID: 2ZWH), in light blue: GJA1-20k models docked using G-actin (PDB ID: 1J6Z), highlighted in purple: RPRPDDLEI tail sequence of GJA1-20k, red and black text in zoomed in box: hotspot residues as defined by Robetta computational alanine scanning. Hotspots are identified by calculating the free energy change (ΔΔG > 1) after a residue is mutated to alanine. (C) Chart of hotspot residues on actin (red) and GJA1-20k (light blue) as defined by Robetta computational alanine scanning. (D) RPEL domain sequences of Cx43, MAL, MRTF-A, and Phactr1. (E) ATTRACT docking modeling of GJA1-20k (light blue and tan) compared with known RPEL domains of Phactr1 and MRTF-A. In orange: MAL-RPEL2 (PDB ID: 2V52), in pale blue: Phactr1-RPEL1 (PDB ID: 4B1W), in silver: Phactr1-RPEL2 (PDB ID: 4B1X), in yellow: Phactr1-RPEL3 (PDB 1D: 4B1Y); in light green: Phactr1-RPELN (PDB 1D: 4B1U), and in dark blue: full Phactr1 RPEL domain (PDB 1D: 4B1Z).