Fig 2: Overexpression of circHIPK3 effectively reverses miR-7-induced inhibition of CRC cells progression.HCT116 and HT29 cells transfected with miR-control, miR-7, circHIPK3, or miR-7 + circHIPK3. Then the ability of cell cloning, proliferation, migration and invasion was, respectively, assessed by colony formation assay (a), EdU assay (b), transwell migration (d), and matrigel invasion (e) assay. And cell apoptosis analysis was tested by flow cytometry with Annexin V-FITC/PI double staining (c). * vs control group, # vs mir-7 group. f qRT-PCR analysis of the expression of growth and metastasis-related miR-7 targets in HCT116 and HT29 cells with circHIPK3 knockdown or overexpression. * vs si-NC group, # vs vector group. g Western blot analysis of the protein expression of FAK, IGF1R, EGFR, and YY1 in fore-mentioned four groups in HCT116 and HT29 cells. Data were represented as means ± S.D. of at least three independent experiments. *p < 0.05, **p < 0.01; #p < 0.05, ##p < 0.01. Scare bar = 50 µm

Fig 3: Integrin ß1 levels, activation and mediated cell activity are not modulated by Sprr3, but downstream signaling activation is impaired in Sprr3 -/- fibroblasts. A, Flow cytometry analysis of integrin ß1 (total and active conformation) in wild-type and Sprr3 -/- MEFs (P1) show no differences in percentage of cell expression or mean fluorescence intensity (MFI). Statistical analysis was performed by 2-way ANOVA with Sidak multiple comparisons test, ns P > .05, n = 3. B, MEFs (P1-P5) were serum starved and plated within a collagen gel and allowed to contract the gel over 72 hours. Data are presented in percentage of total area, where no differences were observed between genotypes. Statistical analysis was performed by 2-way ANOVA with repeated measures and a Sidak multiple comparisons test, P > .05, n = 12. C, MEFs (P1-P5) were serum starved then plated in transwells to perform a Boyden's chamber assay. Membranes were coated with BSA (negative control) or fibronectin. FBS or PDGF was placed in media in the bottom well as a stimulus. No differences were detected in any condition. Statistical analysis was performed by t test where ns P > .05, n = 6. D, MEFs (P1-P5) were serum starved then plated on 10 µg/mL or 0 µg/mL (negative control) fibronectin (FN) at 15-minute intervals to determine cell adhesion. Crystal violet stained cells were quantified by absorbance, and no differences were noted between genotypes. Statistical analysis was performed by 2-way ANOVA with Sidak multiple comparisons test, ns P > .05, n = 3. E, MEFs (P1-P5) were serum starved, then, plated on 1 µg/mL fibronectin coated plates at 15-minute intervals in serum-free media. Lysates were analyzed by western blot for phosphorylated and total Akt, FAK and p38, then, normalized to GAPDH loading control. Densitometry analysis of normalized ratios is presented in the graphs. Densitometry over time is n = 1 for all timepoints. Differences between 0 and 15 minutes were performed by t test where *P < .05, **P < .01, n = 3-7

Fig 4: TRPV4 and stretch-dependent Ca2+ influx induce cytoskeletal reorganization.(a) Stretch (14%) elevates [Ca2+]hTM; the effect is suppressed by HC-06 (n = 43 cells). Averaged trace from a representative experiment. Image acquisition suspended during de-focus (breaks in trace). Inset: typical stretch response, shown at 3 sec & 18sec. Scale bar = 10 µm. (b) Stretch- [Ca2+]i response for cells plated on collagen and Pronectin (fibronectin-like) substrates (N = 5 experiments; n = =20 cells/experiment). (c) Stretch (12)-evoked [Ca2+]i are inhibited by HC-06 (1 µM) and pBPB (100 µM) (N = =3 experiments; n = =14 cells/experiment). (d–f) hTM. Stretch promotes TRPV4-dependent formation of actin stress fibers and focal adhesion complexes. Cells labeled for phalloidin-488 and vinculin-ir following stretch (6%, 2 Hz; 1 hr). Stretch-evoked SF upregulation (N = 12 experiments) is antagonized by HC-06 (N = 6) or BAPTA-AM (100 µM; N = 4). Scale bars = 10 or 50 µm, respectively. (g) Cyclic stretch (6%; 1 hr) increases FAK phosphorylation, but not overall FAK protein expression (N = 3 experiments). (h,i) GSK101 (5 nM, 1 hr) promotes formation of SFs and FAs. This effect is antagonized by HC-06 (N = 3). Scale bar = 50 µm. (j,k) Simultaneous imaging of [Ca2+]i and actin remodeling. GSK (5 nM) elevates [Ca2+]i and stimulates upregulation of mApple-actin –labeled SFs. Numbers denote the time-corresponding images. Scale bars = 10 µm.

Fig 5: ITGB4 mediates LSS-induced FAK, SRC, and NF?B activation in HUVECs. (a) The protein levels of p-FAK, FAK, p-NF?B, NF?B, p-SRC, and SRC were measured by western blot in HUVECs treated with LSS for 0-24 h. (b) The protein levels of p-FAK, FAK, p-NF?B, NF?B, p-SRC, and SRC were measured by western blot isolated from static or low shear stress conditions transfected with scr siRNA or ITGB4 siRNA. (c) The representative images and location of NF?B in HUVECs were assayed by immunofluorescence staining under static or low shear stress conditions transfected with scr siRNA or ITGB4 siRNA. Scale bars: 20 µm. Data are presented as the mean ± SEM. *P < 0.05 and **P < 0.01.