Figure 4.

The molecular architecture of Cx43 on the plasma membrane and putative orientation at the mitochondrial/connexin contact site. (A) Cx43 has four transmembrane domains (M1–M4). The amino-terminus, carboxyl-terminus and cytoplasmic loop (CL) extend into the cytoplasm, while the two extracellular loops face the extracellular space. In gap junctions, the extracellular loops (E1 and E2) of two apposing cells are in close contact. (B) The Cx43 configuration seen in A is depicted in the gap junction plaque shown between two cells (bracket). At the contact site between the mitochondria and gap junction plaque membranes two possible configurations have been suggested (dashed box in (B) and the entirety of (C,D)). Either (C) the hemichannels on the outer mitochondrial membranes oriented with the carboxyl terminus facing the cytoplasm and carboxyl terminal to carboxyl docking would be possible or (D) hemichannels or connexin monomers on the outer mitochondrial membrane have their extracellular loops oriented toward the cytoplasm, which would allow for interactions between the “extracellular loop” and the carboxyl terminus. IMM = inner mitochondrial membrane; OMM = outer mitochondrial membrane; C = carboxyl terminus; PM = plasma membrane; IMS = Intermembrane space; EC = extracellular space. Figure created with https://www.biorender.com. Given the orientation of the connexin in the plasma membrane with the extracellular loops in the extracellular space, it is unlikely that mitochondrial connexins or other possible proteins found on the outer membrane of the mitochondria could interact with the amino acids of the gap junction plaque or annular vesicle connexin extracellular loops. Instead, it has been suggested that binding may occur between the carboxyl-terminal tail segments of connexins, which extend into the cytoplasm (Figure 4C). Specifically, in this proposed model, binding would occur between the carboxyl-terminal regions of the mitochondria connexin and gap junction plaque or annular vesicle connexin molecules (Figure 4C). Support for binding at the carboxyl-terminal regions is provided by the findings that carboxyl-terminal regions can dimerize [77,78,79]. During the gap junction plaque internalization process, which results in the release of annular gap junction vesicles within the cytoplasm, the docked hemichannels remain bound at their extracellular loops [22,24,26,31,80,81]. Thus, the annular gap junction vesicle has a double membrane that is derived from the membrane of the donor cell and the membrane of the recipient cell, and the extracellular loops are located within the space between these two membranes. The carboxyl-terminal region of the connexin hemichannels is derived from the membrane of the host cell, and thus, they are located within the cytoplasm. The carboxyl-terminal region tail of annular gap junction vesicle channels would therefore potentially be available for direct or indirect interaction with the proteins (e.g., connexins) on the outer mitochondrial membrane. However, interactions between the carboxyl-terminal tail and the “external loops” (E1 and E2) (the regions of the connexin molecule that would be docked in the extracellular space, but depending on the orientation on outer mitochondrial membrane, could be projecting into the cytoplasm) (Figure 4D) cannot be ruled out and have been also suggested [9].