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. 2021 Nov 24:9:719993.
doi: 10.3389/fcell.2021.719993. eCollection 2021.

ADAMTS9-AS1 Constrains Breast Cancer Cell Invasion and Proliferation via Sequestering miR-301b-3p

Junqing Chen  1   2 , Ling Cheng  3 , Weibin Zou  1   2 , Rong Wang  1   2 , Xiaojia Wang  1   2 , Zhanhong Chen  1   2
Affiliations

ADAMTS9-AS1 Constrains Breast Cancer Cell Invasion and Proliferation via Sequestering miR-301b-3p

Junqing Chen et al. Front Cell Dev Biol. .

V体育安卓版 - Abstract

Objective: For determination of how ADAMTS9-AS1/miR-301b-3p/TGFBR2/JAK STAT signaling axis modulates progression of breast cancer cells. Methods: Target lncRNA was determined by differential analysis of breast cancer expression data and survival analysis. Differentially expressed miRNAs and target mRNAs that had binding sites with target lncRNA were predicted. GSEA software was used to carry out pathway enrichment analysis for mRNAs. Binding of the researched genes were tested with RNA binding protein immunoprecipitation (RIP). How miR-301b-3p bound TGFBR2 mRNA was tested by dual-luciferase method. Transwell, colony formation, EdU approaches were employed for verification of invasion and proliferation of breast cancer cells in each treatment group. Results: Markedly inactivated ADAMTS9-AS1 in breast cancer pertained to patient's prognosis. MiR-301b-3p was capable of binding TGFBR2/ADAMTS9-AS1. However, overexpression of ADAMTS9-AS1 stimulated miR-301b-3p binding ADAMTS9-AS1 and repressed miR-301b-3p binding TGFBR2 mRNA. ADAMTS9-AS1 interference enhanced cancer proliferation and invasion, facilitated levels of KI67, PCNA, MMP-9 and MMP-2, and activated the JAK STAT signaling pathway. While silencing miR-301b-3p reversed the effect of ADAMTS9-AS1 interference. In addition, TGFBR2 interference or restraining JAK STAT signaling counteracted the effect of ADAMTS9-AS1. Conclusion: ADAMTS9-AS1 could sequester miR-301b-3p to inhibit progression of breast cancer via TGFBR2/JAK STAT pathway VSports手机版. This study supplies a rationale for incremental apprehension of ADAMTS9-AS1 in breast cancer progression. .

Keywords: ADAMTS9-AS1; JAK STAT signaling pathway; TGFBR2; breast cancer; miR-301b-3p. V体育安卓版.

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"VSports在线直播" Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

FIGURE 1
FIGURE 1
ADAMTS9-AS1 is lowly expressed in breast cancer. (A) Volcano map of DElncRNAs, DEmiRNAs and DEmRNAs in breast cancer tumor group and normal group. (B) The survival curve of ADAMTS9-AS1 expression level on the prognosis of patients, with X-axis representing time (in years) and Y-axis representing survival. Red: high expression, blue: low expression. (C) The level of ADAMTS9-AS1 in breast cancer cells; *p < 0.05.
FIGURE 2
FIGURE 2
ADAMTS9-AS1 restrains cancer cell invasion and proliferation. ADAMTS9-AS1 was overexpressed in MDA-MB-231 and BT-549, while being silenced in MCF7. (A) Level of ADAMTS9-AS1 in breast cancer cells. (B) Results of in vitro colony formation method. (C) Results of EdU method (400×). (D) Cell invasion results (100×). (E) The expression of Ki67, PCNA, MMP-2 and MMP-9 after interfering with ADAMTS9-AS1 expression. (F) ADAMTS9-AS1 level in cancer cells. (G) Proliferation of cancer cells. (H) Results of EdU method (400×). (I) Transwell method is used to test cell invasion (100×). (J) The expression of Ki67, PCNA, MMP-2 and MMP-9. *p < 0.05.
FIGURE 3
FIGURE 3
ADAMTS9-AS1 competitively binds to miR-301b-3p to promote TGFBR2. (A) Survival curve of miR-301b-3p level and prognosis of patients. (B) Venn plot of predicted mRNAs and DEmRNAs; diffSig denotes differentially expressed genes screened from TCGA database. (C) Association between ADAMTS9-AS1 and 8 target mRNAs. (D) The survival curve of TGFBR2 expression level on the prognosis of patients. (E,F) Results of RIP method. (G) Result of dual-luciferase method. (H) The expression of ADAMTS9-AS1, miR-301b-3p and TGFBR2. *p < 0.05.
FIGURE 4
FIGURE 4
ADAMTS9-AS1 promotes TGFBR2 through miR-301b-3p to inhibit breast cancer cell invasion and proliferation. (A) The proliferation of MCF-7 cells. (B) EdU assay used to detect MCF-7 cell proliferation (400×). (C) Invasion of MCF-7 cells (100×). (D) Levels of Ki67, PCNA, MMP-2 and MMP-9. (E) Level of TGFBR2 in si-NC, si-TGFBR2-1 and si-TGFBR2-2 groups. The silencing sequence with the highest silencing efficiency was named as si-TGFBR2. (F) ADAMTS9-AS1 and TGFBR2 expression in MDA-MB-231 cells. (G) Proliferation of MDA-MB-231. (H) Proliferation of MDA-MB-231 (400×). (I) Cancer cell invasion (100×). (J) Levels of Ki67, PCNA, MMP-2 and MMP-9. *p < 0.05.
FIGURE 5
FIGURE 5
ADAMTS9-AS1 regulates the JAK STAT signaling pathway by targeting TGFBR2 through miR-301b-3p. (A) GSEA enrichment pathway of TGFBR2 (JAK STAT signaling pathway). (B) The expression of JAK STAT signaling-relevant proteins. (C) The expression of JAK STAT signaling-relevant proteins in si-NC + inhibitor NC, si-ADAMTS9-AS1 + inhibitor NC and si-ADAMTS9-AS1 + miR-301b-3p inhibitor. (D) Levels of JAK STAT signaling-relevant proteins in oe-NC + si-NC, oe-ADAMTS9-AS1 + si-NC and oe-ADAMTS9-AS1 + si-TGFBR2.
FIGURE 6
FIGURE 6
ADAMTS9-AS1 represses cancer invasion and proliferation by regulating JAK STAT signaling pathway. (A) ADAMTS9-AS1 expression. (B) Levels of JAK STAT signaling-relevant proteins. (C) The proliferation of breast cancer cells in vitro. (D) The proliferation of breast cancer cells (400×). (E) Cancer cell invasion (100×). (F) Levels of Ki67, PCNA, MMP-2 and MMP-9. *p < 0.05.
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