Figure 7.

A More Severe Cohesin LOF Induces Actin Ring Formation

(A–G) GFP:moe positively marked WT (A), SMC3KD (B), smc3^A(C), SA1KD (D), SA2KD (E), SA1KD; SA2KD (F), and NipBKD (G) clones. Actin rich rings (yellow arrows) were observed in smc3^A, SA1, and SA2 simultaneous KD and NipBKD clones.

(H) Quantification of the number of actin rings per square millimeter of clonal tissue. Eight animals were analyzed for each genotype.

(I–O) GFP:moe positively marked clones ((I) smc3^A; (J) smc3^A + RhoV14; (K) smc3^A + RhoN; (M) smc3^A + Sqh EE; (N) smc3^A + SqhKD). Dominant-negative Rho (RhoN) and SqhKD inhibit actin ring formation in smc3^A clones; phosphomimetic Sqh (SqhEE) increases the number of clones with actin rings. Quantified in (L) and (O) showing the number of actin rings or delaminated clones per square millimeter of clonal tissue. Each dot represents one animal. smc3^A + RhoV14 resulted in very small unicellular clones (J) or no clones at all and could not be quantified.

(P and Q) Genes involved in apical constriction were either knocked down or overexpressed in GFP:moe positively marked clones, either on their own (P) or within smc3^A clones (Q). Quantification shows the number of actin rings or delaminated clones per square millimeter of clonal tissue. Each dot represents 1 animal.

(R and S) GFP:moe-labeled smc3^A (R) and SA1 + SA2KD (S) clones stained for the active form of the Dpp signaling effector, phosphorylated Mad (pMad).

(T) Quantification of mean fluorescence intensity from the nuclei of cells within clones, with and without actin rings, compared with WT tissue within the same animal. 35 nuclei from 7 animals were measured. Each dot represents one animal. Scale bars, 10 μm. Error bars = ± SEM. Statistical analysis: Student's t test. p > 0.05 was considered not significant, ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001.

See also Figure S7.