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. 2019 Jan 28;48(2):245–260.e7. doi: 10.1016/j.devcel.2018.12.020

Figure 5.

Figure 5

Stretch-Sensitive MyoII Polarization Pathway Induces Stabilization of MyoII on Stretched Junctions

(A) rho-RNAi clones (magenta) in stretched Sqh::GFP expressing discs. (A′) Quantification of Sqh intensity on horizontal and vertical junctions in anchor (Figure S5A) and stretch states; n = 2–3. N.B. Absolute intensities are not comparable between anchor and stretch discs as different clones are used for each analysis. (A″) Quantification of Sqh polarity on anchor and stretch states in control and rho-RNAi clones.

(B) rho-RNAi clones (indicated in magenta) in anchored Dia::GFP expressing discs. (B′) Quantification of Dia::GFP mean intensity in clone regions compared to wild-type regions in anchored disc; n = 5.

(C) zip-RNAi clones (magenta) in stretched Rho::GFP expressing discs. Discs were fixed under stretch and stained. Rho::GFP boosted with anti-GFP-FITC. (C′) Quantification of Rho::GFP polarity in stretched discs expressing zip-RNAi clones and Rho::GFP; n = 4, p < 0.001.

(D) rho-RNAi clones (magenta) in stretched RokK116a::Venus expressing discs.

(D′) Quantification of RokK116a::Venus total mean intensity in rho-RNAi clones and in WT control tissue, in anchor (n = 2), and stretched (n = 7) discs.

(E) dia-RNAi clones (magenta) in stretched RokK116a::Venus expressing discs. (E′) Quantification of RokK116a::Venus polarity in anchored (Figure S5F) and stretched discs expressing dia-RNAi clones; n = 3.

(A′–E′) Error bars indicate S.E.M.; ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗∗ p < 0.0001 with t test. (A-E) Scale bars, 10 μm.

(F) Pathway schematic summarizing the molecular pathways regulating MyoII polarity. The polarization of MyoII protein levels are stretch sensitive and via Rho and Dia, but it’s phosphorylation per se is stretch insensitive and via Rho and Rok.

(G) FRAP half time of Sqh::GFP on horizontal and vertical junctions in discs expressing either Sqh::GFP, enGal4>UAS-Act5CRFP, or Sqh::GFP, enGal4>UAS-dia-RNAi, or UAS-Act5CRFP subjected to <10 min stretch. The red line indicates the median, the red diamond the mean, edges the 25th and 75th percentiles, and whiskers the most extreme data points that are not considered to be outliers. Black points represent individual junctions. ∗ p < 0.05 with Wilcoxon rank-sum test.