The origin of annular junctions: a mechanism of gap junction internalization

Gap junctional intercellular communication is established when connexin pro teins oligomerize into connexon hemichannels, which then pair at the cell s urface with connexons from neighboring cells to form functional gap junctio n channels. Gap junction channels routinely cluster into gap junction plaqu es, which can exhibit dynamic characteristics while under the frequent proc esses of formation and removal from the cell surface. We have three lines o f evidence to suggest that one mechanism of gap junction removal occurs whe n one of two contacting cells internalizes the gap junction contribution fr om both cells. First, in coculture experiments, green fluorescent protein-t agged connexin43 (Cx43-GFP) expressed in normal rat kidney (NRK) cells can be internalized into contacting cells that do not express Cx43-GFP, and the incidences of identifying these internalized structures increase in the pr esence of lysosomal inhibitors. Secondly, time-lapse imaging of live NRK ce lls revealed that large areas of gap junction plaques containing Cx43-GFP w ere internalized as vesicular-like structures into one of two adjacent cell s. Finally, when live NRK cells that express endogenous Cx43 were microinje cted with anti-Cx43 antibodies, antibody-tagged gap junctions were visualiz ed in cells that contacted the microinjected cell within 3-6.5 hours. Toget her our results strongly suggest that one mechanism of gap junction removal from the cell surface involves a unique process in which the entire gap ju nction or a fragment of it is internalized into one of the two contacting c ells as an annular junction.