Relative quantification (2?CT) was used for results analysis
Relative quantification (2?CT) was used for results analysis. accelerate cell proliferation and migration, and impede cell apoptosis in NSCLC cell lines. Mechanically, SPRY4 is confirmed a direct target of miR-411-5p/3p. Furthermore, our findings showed that miR-411-5p/3p promoted lung tumor growth in vivo, decreased SPRY4 expression dramatically, and induced EGFR, AKT signaling activation, as well as epithelialCmesenchymal transition (EMT) simultaneously in tumor tissues. In addition, we showed that miR-411-5p also targeted tumor suppressor TXNIP, involved in regulating positively cell cycle progress in SPC-A1 cells rather than in H1299. Whether cell specificity of low TXNIP mRNA level in H1299 is responsible for the different response to cell cycle between H1299 and SPC-A1 would need further explorations. Collectively, these results suggest that miR-411-5p/3p are required for NSCLC development by suppressing SPRY4 and TXNIP; thus, the miR-411-SPRY4-AKT axis might act as a promising target for lung cancer therapy clinically. and their alternative splicing results in multiple transcript variants (SPRY1, SPRY2, SPRY3, and SPRY4) [12C14], which are reported to downregulate the expression of epidermal growth factor receptor (EGFR) . functions as a tumor suppressor downstream of Wnt7A/Fzd9 signaling in lung cancer, whose overexpression inhibited cell growth with upregulating the tumor suppressor p53 and p21 expression, and also suppressed cell migration and invasion along with MMP-9 activity . is activated by a target downstream of Wnt7A/Fzd9 signaling , PPAR, which has vital roles in ovarian cancer , colorectal cancer , and prostate cancer , and affects ARQ 197 (Tivantinib) cell growth, differentiation, and metastasis . In melanoma , breast cancer , and prostate cancer , SPRY4 inhibits cell migration and the cancer stem cell properties of breast carcinoma cells . Nevertheless, as an oncogene, SPRY4 promotes ovarian cancer invasion through involvement in EGFR-mediated human ovarian cancer progression . ARQ 197 (Tivantinib) Thioredoxin interaction protein (TXNIP) has pivotal roles in prostate cancer, lung cancer, and breast cancer [26C28], and has an especially prognostic effect in NSCLC . MiR-411 belongs to the 14q32.31 miRNA cluster . In the present study, we confirmed SPRY4 as a common target of miR-411-5p and miR-411-3p. Moreover, Rabbit polyclonal to ERCC5.Seven complementation groups (A-G) of xeroderma pigmentosum have been described. Thexeroderma pigmentosum group A protein, XPA, is a zinc metalloprotein which preferentially bindsto DNA damaged by ultraviolet (UV) radiation and chemical carcinogens. XPA is a DNA repairenzyme that has been shown to be required for the incision step of nucleotide excision repair. XPG(also designated ERCC5) is an endonuclease that makes the 3 incision in DNA nucleotide excisionrepair. Mammalian XPG is similar in sequence to yeast RAD2. Conserved residues in the catalyticcenter of XPG are important for nuclease activity and function in nucleotide excision repair miR-411-5p/3p could promote NSCLC cell proliferation, tumor growth, and metastasis in vitro and in vivo. These results indicated that the miR-411 could be a cancer driver in lung tumorigenesis. Results MiR-411 is upregulated in human NSCLC tissues and cell lines ARQ 197 (Tivantinib) We investigated the miR-411-5p/3p expression in human NSCLC tissue samples and cell lines. Results of quantitative reverse-transcriptase PCR (qRT-PCR) indicated that the miR-411-5p/3p expression was significantly higher in the 33 human lung tumor samples than in those of adjacent non-tumor tissues (Fig. 1a, b). It was also observed that miR-411-5p/3p were upregulated in most human NSCLC cell lines compared with the normal bronchial epithelium cell line HBE135-E6E7 (HBE, Fig. 1c, d). The results indicated that miR-411 could function as an oncogene. Open in a separate window Fig. 1 MiR-411 expression was upregulated in NSCLC. a, b Relative miR-411-5p/3p expression in NSCLC and corresponding paracancerous lung tissues (is also confirmed to be a target of miR-411-5p ARQ 197 (Tivantinib) and decreased in NSCLC cell lines and lung cancer tissue samples. (Fig. 8a, b). Next, we set out to assess the effect of repression of TXNIP in H1299 and SPC-A1 cells with pLenti-miR-411 compared with pLenti cells, and thus investigated the expression of TXNIP by western blotting, which was not surprisingly decreased in both H1299 and SPC-A1 cells. (Fig. ?(Fig.8c).8c). It was further confirmed to be a direct target of miR-411-5p by dual luciferase reporter assay (Fig. .8d, e). Open in a separate window Fig. 8 TXNIP is a direct target of miR-411-5p. a TXNIP mRNA expression in A549, SPC-A1, H1299, PC-9, and 95-D cells. HBE cell line was normal control. b TXNIP mRNA expression in NSCLC tissue samples and corresponding non-tumor tissues (is a common target of miR-411-5p and miR-411-3p. Acting as an oncogene within this regulation network, miR-411-5p/3p increased cell proliferation and migration, while decreased apoptosis in NSCLC (Fig. ?(Fig.10).10). These results suggested that as a target, is involved in the oncogenic functions of miR-411-5p/3p in NSCLC. Open in a separate window Fig. 10 A diagram of the functions of miR-411 in NSCLC In lung cancer, SPRY4 is a promising target to interdict the progress of NSCLC therapy engaged in the progression of EGFR-mediated ovarian cancer and enhances melanoma cell motility and treatment of gastrointestinal tumors [12, 31]. Moreover, SPRY4 regulates various signals negatively in angiogenesis, selectively suppressing angiogenic signals independent of Ras in the tumor microenvironment . Also, it is reported to inhibit the mitogen-activated protein kinase (MAPK) signaling pathway. The high SPRY4 expression often impedes cell growth of NSCLC and induces a reversal.