Structure of the connexin-43 gap junction channel reveals a closed sieve-like molecular gate

Gap junction channels (GJCs) mediate intercellular communication by connecting two neighboring cells and enabling direct exchange of ions and small molecules. Cell coupling via connexin-43 (Cx43) GJCs is important in a wide range of cellular processes in health and disease, yet the structural basis of Cx43 function and regulation has not been determined until now. Here we describe the structure of a human Cx43 GJC solved by cryo-EM and single particle analysis at 2.26 Å resolution. The pore region of Cx43 GJC features several lipid-like densities per Cx43 monomer, located close to a putative lateral access site at the monomer boundary. We found a previously undescribed closed sieve-like gate on the cytosolic side of the pore, formed by the N-terminal domain and the transmembrane helix 2 of Cx43 and stabilized by a small molecule. Structures of the Cx43 GJC and hemichannels in nanodiscs confirm the sieve-like gate arrangement. The features of the Cx43 GJC and hemichannel cryo-EM maps and the channel properties revealed by molecular dynamics simulations suggest that the captured states of Cx43 are closed. ### Competing Interest Statement The authors have declared no competing interest.