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Ephrin-Bs Drive Junctional Downregulation and Actin Stress Fiber Disassembly to Enable Wound Re-epithelialization.

Nunan R, Campbell J, Mori R, Pitulescu ME, Jiang WG, Harding KG, Adams RH, Nobes CD, Martin P - Cell Rep (2015)

Bottom Line: Much is known about how lead-edge cells migrate, but very little is known about the mechanisms that enable active participation by cells further back.Here we show that ephrin-B1 and its receptor EphB2 are both upregulated in vivo, just for the duration of repair, in the first 70 or so rows of epidermal cells, and this signal leads to downregulation of the molecular components of adherens and tight (but not desmosomal) junctions, leading to loosening between neighbors and enabling shuffle room among epidermal cells.If this signaling axis is perturbed, then disrupted healing is a consequence in mouse and man.

View Article: PubMed Central - PubMed

Affiliation: Schools of Biochemistry and Physiology & Pharmacology, University of Bristol, Bristol BS8 1TD, UK.

No MeSH data available.


Related in: MedlinePlus

Ephrin-B1/B2 May Be Required to Shut Down Actomyosin-Generated Tension within the Advancing Epidermal Sheet(A) HaCaT cells back from the wound edge after efnB1/B2 KD and subsequent treatment with the ROCK inhibitor Y27632 (650 nM) or the actomyosin inhibitor blebbistatin (10 μM). Cells were stained to reveal actin (red) and E-cadherin (green). A z stack maximum projection is shown with magnification of the apical planes from the boxed area; yellow arrowheads indicate intercellular apical stress fibers linked through adherens junctions.(B) (Left) Time-lapse images of stalled ephrin-B1/B2 KD cells at the start (red) and end (yellow) of 3-hr time windows. Red line indicates position of cell sheet at time of Y27632 addition and yellow line shows wound edge after a 3-hr time period. (Right) Bar chart illustrates migration distance traversed during the 3-hr periods before or after Y27632 addition (∗∗p < 0.01, as determined by an unpaired Student’s t test; n = 4).Scale bars, 20 μm (A; 10 μm in apical images, boxed) and 50 μm (B).
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fig6: Ephrin-B1/B2 May Be Required to Shut Down Actomyosin-Generated Tension within the Advancing Epidermal Sheet(A) HaCaT cells back from the wound edge after efnB1/B2 KD and subsequent treatment with the ROCK inhibitor Y27632 (650 nM) or the actomyosin inhibitor blebbistatin (10 μM). Cells were stained to reveal actin (red) and E-cadherin (green). A z stack maximum projection is shown with magnification of the apical planes from the boxed area; yellow arrowheads indicate intercellular apical stress fibers linked through adherens junctions.(B) (Left) Time-lapse images of stalled ephrin-B1/B2 KD cells at the start (red) and end (yellow) of 3-hr time windows. Red line indicates position of cell sheet at time of Y27632 addition and yellow line shows wound edge after a 3-hr time period. (Right) Bar chart illustrates migration distance traversed during the 3-hr periods before or after Y27632 addition (∗∗p < 0.01, as determined by an unpaired Student’s t test; n = 4).Scale bars, 20 μm (A; 10 μm in apical images, boxed) and 50 μm (B).

Mentions: From the observations described above, we were curious whether stalling of epithelial migration in ephrin-B1/B2 KD scratch wounds may, in part, be a consequence of unreleased tension within the follower cells further back from the wound edge. At 3 hr post-wounding when stalling was first occurring in ephrin-B1/B2 KD cells, we observed a very different pattern of actin stress fibers in KD versus control follower cells. They exhibited considerably more stress fibers, with many of these extending fully across the cell and linking adherens junctions between cells, whereas most follower cells in control wounds exhibited only cortical actin filaments with very few stress fibers (Figure 6A). This failure to dissolve actin stress fibers in ephrin-B KD wounds was almost completely abrogated by exposure of cells to the ROCK inhibitor Y27632 (Figure 6A), which did not alter junction staining in control cells, and we saw a similar response after treatment with blebbistatin (Figure 6B). Movie analysis of scratch wounds shows that previously stalled ephrin-B KD cells rapidly responded to Y27632 (Figure 6B; Figure S6B; Movie S3) or blebbistatin (Figure S6C) by resurgent forward migration, suggesting that actin-generated tension release (although we have not directly measured it) may be part of the mechanism whereby ephrin-B signaling enables wound re-epithelialization. However, it is noticeable that this rescue was only partial and temporary (Figure 6B) as might be expected, since it only influenced tension loss and did not activate junction dissolution.


Ephrin-Bs Drive Junctional Downregulation and Actin Stress Fiber Disassembly to Enable Wound Re-epithelialization.

Nunan R, Campbell J, Mori R, Pitulescu ME, Jiang WG, Harding KG, Adams RH, Nobes CD, Martin P - Cell Rep (2015)

Ephrin-B1/B2 May Be Required to Shut Down Actomyosin-Generated Tension within the Advancing Epidermal Sheet(A) HaCaT cells back from the wound edge after efnB1/B2 KD and subsequent treatment with the ROCK inhibitor Y27632 (650 nM) or the actomyosin inhibitor blebbistatin (10 μM). Cells were stained to reveal actin (red) and E-cadherin (green). A z stack maximum projection is shown with magnification of the apical planes from the boxed area; yellow arrowheads indicate intercellular apical stress fibers linked through adherens junctions.(B) (Left) Time-lapse images of stalled ephrin-B1/B2 KD cells at the start (red) and end (yellow) of 3-hr time windows. Red line indicates position of cell sheet at time of Y27632 addition and yellow line shows wound edge after a 3-hr time period. (Right) Bar chart illustrates migration distance traversed during the 3-hr periods before or after Y27632 addition (∗∗p < 0.01, as determined by an unpaired Student’s t test; n = 4).Scale bars, 20 μm (A; 10 μm in apical images, boxed) and 50 μm (B).
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fig6: Ephrin-B1/B2 May Be Required to Shut Down Actomyosin-Generated Tension within the Advancing Epidermal Sheet(A) HaCaT cells back from the wound edge after efnB1/B2 KD and subsequent treatment with the ROCK inhibitor Y27632 (650 nM) or the actomyosin inhibitor blebbistatin (10 μM). Cells were stained to reveal actin (red) and E-cadherin (green). A z stack maximum projection is shown with magnification of the apical planes from the boxed area; yellow arrowheads indicate intercellular apical stress fibers linked through adherens junctions.(B) (Left) Time-lapse images of stalled ephrin-B1/B2 KD cells at the start (red) and end (yellow) of 3-hr time windows. Red line indicates position of cell sheet at time of Y27632 addition and yellow line shows wound edge after a 3-hr time period. (Right) Bar chart illustrates migration distance traversed during the 3-hr periods before or after Y27632 addition (∗∗p < 0.01, as determined by an unpaired Student’s t test; n = 4).Scale bars, 20 μm (A; 10 μm in apical images, boxed) and 50 μm (B).
Mentions: From the observations described above, we were curious whether stalling of epithelial migration in ephrin-B1/B2 KD scratch wounds may, in part, be a consequence of unreleased tension within the follower cells further back from the wound edge. At 3 hr post-wounding when stalling was first occurring in ephrin-B1/B2 KD cells, we observed a very different pattern of actin stress fibers in KD versus control follower cells. They exhibited considerably more stress fibers, with many of these extending fully across the cell and linking adherens junctions between cells, whereas most follower cells in control wounds exhibited only cortical actin filaments with very few stress fibers (Figure 6A). This failure to dissolve actin stress fibers in ephrin-B KD wounds was almost completely abrogated by exposure of cells to the ROCK inhibitor Y27632 (Figure 6A), which did not alter junction staining in control cells, and we saw a similar response after treatment with blebbistatin (Figure 6B). Movie analysis of scratch wounds shows that previously stalled ephrin-B KD cells rapidly responded to Y27632 (Figure 6B; Figure S6B; Movie S3) or blebbistatin (Figure S6C) by resurgent forward migration, suggesting that actin-generated tension release (although we have not directly measured it) may be part of the mechanism whereby ephrin-B signaling enables wound re-epithelialization. However, it is noticeable that this rescue was only partial and temporary (Figure 6B) as might be expected, since it only influenced tension loss and did not activate junction dissolution.

Bottom Line: Much is known about how lead-edge cells migrate, but very little is known about the mechanisms that enable active participation by cells further back.Here we show that ephrin-B1 and its receptor EphB2 are both upregulated in vivo, just for the duration of repair, in the first 70 or so rows of epidermal cells, and this signal leads to downregulation of the molecular components of adherens and tight (but not desmosomal) junctions, leading to loosening between neighbors and enabling shuffle room among epidermal cells.If this signaling axis is perturbed, then disrupted healing is a consequence in mouse and man.

View Article: PubMed Central - PubMed

Affiliation: Schools of Biochemistry and Physiology & Pharmacology, University of Bristol, Bristol BS8 1TD, UK.

No MeSH data available.


Related in: MedlinePlus