Cell–cell contacts: Gap junctions visualized via connexin-43​

A. Z-stack

B. Timelapse

Figures. Z-stack and timelapse movies of gap junctions. A) Z-stack of a live hiPS cell colony expressing mEGFP-tagged connexin-43. Cells were imaged in 3D on a spinning-disk confocal microscope. Movie starts at the bottom of the cells and ends at the top. Scale bar = 5 µm. B) Timelapse movie of a live hiPS cell colony expressing mEGFP-tagged connexin-43. Cells were imaged on a spinning-disk confocal microscope every 3.25 min. Image is a maximum intensity projection of 20 z-slices acquired at the top of the cells. Contrast is enhanced to show localization of connexin-43 to internal structures. Frames were histogram matched to adjust for photobleaching. Movie plays at 975x real time. Scale bar = 5 µm.
​
Observations

• Connexin-43 is a key component of gap junctions, which are channels between cells that allow for small molecules and ions to pass directly between the cytoplasm of adjacent cells. Gap junctions often cluster in larger gap junction plaques.
• In hiPS cells, connexin-43 localizes most prominently to many puncta (presumably clusters of individual channels) at apical cell-cell junctions near the very top of the cells. Some puncta are also seen near the bottom and middle of the cell although in smaller numbers.
• Connexin-43 is also localized to the plasma membrane and in internal structures that resemble the Golgi. This localization likely reflects the assembly and trafficking of connexin into gap junctions.
• ​As hiPS cells progress though mitosis, the apical puncta at cell-cell junctions remain intact, but the connexin-43 associated with internal organelles, e.g. the Golgi, becomes diffuse throughout the cell. ​