Fig 2: Circ_0000511 promotes BC tumor progression in vivo. MCF-7 cell line was stably transfected with sh-NC or sh-circ_0000511. Nude mice were subcutaneously injected with MCF-7 (2×106 cells/200 µl PBS) stably expressing sh-NC or sh-circ_0000511. (A) Tumor volume was measured every week. (B) Mice were euthanized after 4 weeks of inoculation, and the tumors were resected and weighed. (C) Circ_0000511 and (D) miR-326 expression was measured in the sh-NC and sh-circ_0000511 groups by RT-qPCR. (E) RT-qPCR and (F) western blot assay were used to examine the mRNA and protein expression levels of TAZ, respectively, in tumor tissues. *P<0.05 vs. sh-NC. circ_0000511, circular RNA 0000511; BC, breast cancer; miR, microRNA; sh, short hairpin RNA; NC, negative control; TAZ, transcriptional co-activator with PDZ-binding motif.

Fig 3: miR-326-mediated influences are partly reversed by the overexpression of TAZ in BC cells. MCF-7 and MDA-MB-468 cells were transfected with miR-NC, miR-326, miR-326 + pcDNA or miR-326 + TAZ. (A) Reverse transcription-quantitative PCR and (B) western blot assay were used to examine the mRNA and protein expression levels of TAZ, respectively, in BC cells. (C) Colony formation assay was performed to measure the proliferative capacity of BC cells, and the number of colonies in each group was detected. MTT assay was used to analyze the proliferative ability of (D) MCF-7 and (E) MDA-MB-468 cells. (F) Flow cytometry was conducted to assess the apoptosis of BC cells. Protein expression levels of Bcl-2, Bax and Cleaved caspase-3 were detected in (G) MCF-7 and (H) MDA-MB-468 cells by western blot assay. Transwell assays were conducted to assess the (I) migration and (J) invasion of BC cells. *P<0.05. Bcl-2, B cell leukemia/lymphoma 2; Bax, Bcl-2 associated X apoptosis regulator; BC, breast cancer; miR, microRNA; NC, negative control; TAZ, transcriptional co-activator with PDZ-binding motif; OD, optical density.

Fig 4: Circ_0000511 elevates TAZ mRNA and protein expression via sponging miR-326 in BC cells. MCF-7 and MDA-MB-468 cells were transfected with si-circ_0000511 alone or together with anti-miR-326. (A) Reverse transcription-quantitative PCR and (B) western blot assay were used to assess the mRNA and protein abundance of TAZ in BC cells. *P<0.05. circ_0000511, circular RNA 0000511; BC, breast cancer; miR, microRNA; si, small interfering RNA; NC, negative control; TAZ, transcriptional co-activator with PDZ-binding motif.

Fig 5: PAX8 is required for TAZ protein stability. A, Western blotting analysis of YAP and TAZ from stably transfected cells. B, Relative TEAD reporter activity in stably transfected SKOV3 and A2780 cells. C, RT-PCR results performed using total RNA from the indicated cell lines. GAPDH mRNA was amplified as control. D, Co-IP results from SKOV3 and A2780 showing PAX8 coprecipitates with TAZ. E, Representative Western blot of TAZ expression at baseline or after incubation with MG132 (25 µmol/L) treatment for 8 h. GAPDH was used for normalization. F, Western blot analysis of TAZ proteins in indicated cells treated with CHX (30 µg/mL) for 0, 2, 4 or 6 h. G, CHX release profiles. Data are represented as the mean ± SEM. Student's t test: **P < .01, *P < .05, n.s., not significant