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. 2022 Jul 12;119(29):e2203257119. doi: 10.1073/pnas.2203257119

Fig. 4.

Fig. 4.

Inhibition of the Hippo pathway changes Hydra morphology. (A) Live polyp electroporated with shHyLats, 14 d after electroporation. (B) Graph shows the percentage of thick tentacle formation in polyps electroporated with shGFP (control), shHyLATS alone, shHyLATS/shHyYap, and shHyLATS/shHyYapscr. N, number of polyps; n, number of experiments; two-tailed unpaired t test. (C–D′) EdU is not detected in either control (C and C′) or thick (D and D′) tentacles. Edges of tentacles are outlined by dotted lines. (E) Hydra polyp electroporated with shHyLATS and stained with phalloidin, 12 d after electroporation. Visible tear along the body column is an artifact of fixation and is common when shortened shHyLats electroporated polyps are fixed. (F and G) Ends of normal (F) and thick (G) tentacles stained with phalloidin. Arrows point to the ectodermal muscle processes that are filled with actin fibers and oriented along the length of the tentacle. (H and I) Lateral view of the ectoderm of the body columns underneath normal tentacle (H) and thick (I) tentacle stained with phalloidin. Double-headed arrows indicated the thickness of the ectoderm. (J and K) Apical view of the normal (J) and thick (K) tentacles of MRLC-GFP polyps immunostained for GFP. (L–N) Transmission electron microscopy of cross-sections visualizing the basal compartment of ectodermal epithelial cells in shHyLats tentacles (L), shHyLats body column (M), and wild-type tentacles (N). (L and M) Muscle processes (green) exhibit abnormal elongation along the apical–basal axis of the cells and sometimes ectopic positioning distant from the mesoglea. Muscle processes remain connected by normal numbers of spot desmosome-like junctions (arrowheads), but they show a dramatic loss of their parallel alignment along the polyp’s oral–aboral body axis as shown in wild-type controls (N). (L and M) At positions where the disrupted planar array of muscle fibers had gaps, the mesoglea (yellow) folded into the cytoplasm of the ectodermal epithelial cells without losing the hemidesmosome-like junctions usually located at the basal membrane surface of epithelial cells. (N) A representative image with an ectopic muscle process running along the apical–basal axis (arrow) pointing toward the mesoglea folding. ***(P ≤ 0.001)