Figure 7

Figure 7

Caveolae Facilitate YAP/TAZ-Mediated Flow Response

(A) Confluent HEK293A cells were kept in chambers with different flow speeds for 18 hr. YAP/TAZ KO cells were kept at high flow only. Western blot of cell lysates separated on a PhosTag gel and probed for YAP is shown. Note the dephosphorylation (downshift of YAP) and therefore predicted activation of YAP upon flow.

(B) Western blots of cell lysates as in (A). Note the increase in CYR61 protein upon flow in WT cells.

(C) Confocal images of cells grown without (−) or with flow at 2.1 × 10⁻⁵ Dyn/cm² (+). Cells labeled for YAP (green) and CAV1 (red) are shown. Scale bar represents 20 μm.

(D) Dot plot representing quantification of nuclear-to-cytoplasmic ratio of YAP/TAZ localization from images as in (C). Each dot represents one cell. Means ± SEM.

(E) qPCR analysis of HEK293A WT cells kept at either 0 or 2.1 × 10⁻⁵ Dyn/cm². CTGF and CYR61 mRNA were increased upon flow. Means ± SD.

(F) Western blot of cell lysates from either control or shCAV1 cells kept at either 0 (− flow) or at 2.1 × 10⁻⁵ Dyn/cm² (+ flow). Note induction of CYR61 upon flow in WT cells (related to Figures S5F, S5G, and S6F).

(G) Quantification of CYR61 levels from western blots as in (F), normalized to HSP90 levels. n = 3; means ± SD.

(H) Relative fold induction of target gene expression in shCAV1 upon flow compared to control cells. Means ± SD.

(I) Dot plots of YAP localization from images of cells at steady state. Each dot represents one cell. Means ± SEM.

(J) Dot plots of normalized values of flow-induced (2.1 × 10⁻⁵ Dyn/cm²) YAP nuclear localization. Each dot represents one cell. Means ± SEM.

(K) Relative fold induction of target gene expression in two separate CAV1 KO clones upon flow was compared to control cells. Means ± SD. Flow-mediated induction of CTGF and CYR61 is CAV1 dependent.

Further related to this figure are Figures S6G, S6H, and S7A–S7C.