Additional file 2 of USP36 promotes tumorigenesis and tamoxifen resistance in breast cancer by deubiquitinating and stabilizing ERα
Creators: name: Zhuang, Ting
name: Zhang, Shuqing
name: Liu, Dongyi
name: Li, Zhongbo
name: Li, Xin
name: Li, Jiaoyan
name: Yang, Penghe
name: Zhang, Chenmiao
name: Cui, Jiayao
name: Fu, Mingxi
name: Shen, Fangyu
name: Yuan, Lei
name: Zhang, Zhao
name: Su, Peng
name: Zhu, Jian
name: Yang, Huijie
Publisher: name: figshare
Year: 2024
Subjects: subject: Biochemistry
subject: Cell Biology
subject: Genetics
subject: FOS: Biological sciences
subject: Molecular Biology
subject: Pharmacology
subject: Chemical Sciences not elsewhere classified
subject: Biological Sciences not elsewhere classified
subject: Cancer
subject: Hematology
subject: Plant Biology

description: Supplementary Material 2: Supplementary Figure 1: A The protein expression of USP36 in breast cancer tissues (n=125) and normal tissues (n=18) from TCGA database ( https://www.genome.gov/ ). B: The protein expression of USP36 in ER positive breast cancer tissues (n=64) and normal tissues (n=18) from TCGA database ( https://www.genome.gov/ ). C: Immunoblot analysis showing the expression level of USP36 in MCF-7, T47D and MCF-10A cells. β-Actin was used as the internal control. D: Real-time PCR results of USP36 mRNA expression in MCF-7, T47D and MCF-10A cells exposed as indicated. Three independent experiments were conducted to obtain the results shown in Panels C-D. All the data are presented as the means ± SDs. ***P &lt; 0.001 for comparisons (Student’s t test). Supplementary Figure 2. A: Immunoblot analysis showing the expression level of USP36 in MCF-7 cells stably transduced with indicated shRNAs by lentivirus. Immunoblotting analyses were performed using the indicated antibodies. β-Actin was used as the internal control. B: The CCK-8 assays were used to detect the cell viability of MCF-7 cells stably transduced with indicated shRNAs by lentivirus at the indicated time points. C-D: Colony formation (left panel) of MCF-7 cells stably transduced with indicated shRNAs by lentivirus. E shows the quantitative analysis of the colony formation assay results. E-F: Transwell assays were used to detect migration ability of MCF-7 cells stably transduced with indicated shRNAs by lentivirus. G shows the quantitative analysis of the Transwell assays results. G-H: Wound healing assays were used to detect migration ability of MCF-7 cells stably transduced with indicated shRNAs by lentivirus. I shows the quantitative analysis. I-J: The percentage of apoptotic cells was determined by FACS analysis of MCF-7 cells stably transduced with indicated shRNAs by lentivirus. PI and Annexin V staining were performed on the cells. K shows the quantitative analysis. K: GREB1, PKIB, PS2 and PDZK1 mRNA levels were determined by real-time PCR in MCF-7 cells stably transduced with indicated shRNAs by lentivirus. Three independent experiments were conducted to obtain the results shown in Panels A-K. All the data are presented as the means ± SDs. *P &lt; 0.05; **P &lt; 0.01;***P &lt; 0.001 for comparisons (Student’s t test). Supplementary Figure 3A: USP36 protein levels were determined by Immunoblot analysis. MCF-7 cells were transiently transfected with 20 nM siControl or 20 nM siUSP36 for a total of 15 days. Protein was extracted every five days. Using Bradford protein assays, the concentration of proteins was determined. Then, cell lysates were immunoblotted with the indicated antibodies. β-Actin was used as an internal control. Three independent experiments were conducted to obtain the results.