Figure 8.

circSKA3-Containing Exosomes Potentiate Tumor Invasion

(A) Left: expression of circSKA3 formed larger tumors relative to the vector control. Right: tumor sizes at day 20 after injection. (B) Tumors collected from mice injected with circSKA3-transfected MCF-7 cells showed increased levels of circSKA3 expression. (C) Quantitation of circSKA3 levels (left), circSKA3 binding to Itgb1 (middle), and percentages of circSKA3 binding with Itgb1 (right). (D) Quantitation of circSKA3 levels (left), circSKA3 binding to invadopodia (middle), and percentages of circSKA3 complexed in the invadopodia (right). (E) Quantitation of invadopodium levels. (F) After injection with MB-231 cells, CD-1 nude mice were also injected with control oligo, circSKA3 siRNA. Delivery of circSKA3 siRNA increased mouse viability significantly relative to controls (log rank test, ∗∗p < 0.01). (G) Invadopodium formation was repressed in the tumor tissues when the mice were injected with circSKA3 siRNA. (H) Immunoprecipitation showed that overexpression of circSKA3 promoted Tks5 phosphorylation, which happened in the cell membrane. (I) Left: MB-231 cells were cultured on gelatin-coated dish for 24 h. The subcellular fractions and total invadopodia were isolated and lysed, and 10 μg of the samples was processed to immunoblot. Both Itgb1 and Tks5 were highly expressed in the invadopodia. The above fraction immunoprecipitation with antibody against Tks5 showed that the Tks5 was highly tyrosine phosphorylated in the invadopodia. Right: western blot showed that the lipid raft fractions expressed higher levels of Itgb1 and Tks5 compared with the non-raft fraction. Tks5 in the raft fractions displayed high levels of tyrosine phosphorylation. (J) Tumor sections were subjected to H&E staining. Tumor cells invading into muscle tissues were indicated by arrows.