Contractomere motility conserves junctional length. (A) Frames from time-lapse structured-illumination of venus-alpha-actinin-1 showing contractomere at the beginning (pink circles) and the end (yellow circles) of a 4-h movie. During intercellular organization, junctional length can shorten when two contractomeres move toward each other and the junctional length can extend when two contractomeres move away from each other. The distance and direction of travel for the contractomeres are shown in the upper right panel. The lower left two panels show the cells maintaining their neighbors during the 4-h movie. The lower right three panels show contractomere motility in two cells. Scale bars are 10 μm. (B) Measurement of junctional lengths of the two cells in A. Cells #1 and #2 are used to illustrate that some junctions increased but the others decreased their lengths, resulting in near zero net change in total junctional lengths. (C) Frames from time-lapse wide-field of venus-alpha-actinin-1 and synaptopodin-venus. Gliding of contractomeres to reproportion junctional lengths. In the α-actinin movie, motility of contractomeres resulted in shortening the junction between contractomeres a and b with concomitant lengthening of the junction between contractomeres b and c. In the synaptopodin movie, the motility of contractomeres resulted in lengthening of the junction between a and b with concomitant shortening of the junction between contractomeres b and c. Scale bars are 5 μm. (D) Plating of MDCK cells on collagen I at confluent density resulted in intercellular movement and neighbor exchange. The total junctional length of individual cells remained relatively constant. Scale bar is 10 μm. (E) Frames from time-lapse of occludin showing contractomere and junctional movement over 12 h. Contractomeres are circled.