TXNDC17 promotes paclitaxel resistance via inducing autophagy in ovarian cancer

Paclitaxel is recommended as a first-line chemotherapeutic agent against ovarian cancer, but drug resistance becomes a major limitation of its success clinically. The key molecule or mechanism associated with paclitaxel resistance in ovarian cancer still remains unclear. Here, we showed that TXNDC17 screened from 356 differentially expressed proteins by LC-MS/MS label-free quantitative proteomics was more highly expressed in paclitaxel-resistant ovarian cancer cells and tissues, and the high expression of TXNDC17 was associated with poorer prognostic factors and exhibited shortened survival in 157 ovarian cancer patients. Moreover, paclitaxel exposure induced upregulation of TXNDC17 and BECN1 expression, increase of autophagosome formation, and autophagic flux that conferred cytoprotection for ovarian cancer cells from paclitaxel. TXNDC17 inhibition by siRNA or enforced overexpression by a pcDNA3.1(+)-TXNDC17 plasmid correspondingly decreased or increased the autophagy response and paclitaxel resistance. Additionally, the downregulation of BECN1 by siRNA attenuated the activation of autophagy and cytoprotection from paclitaxel induced by TXNDC17 overexpression in ovarian cancer cells. Thus, our findings suggest that TXNDC17, through participation of BECN1, induces autophagy and consequently results in paclitaxel resistance in ovarian cancer. TXNDC17 may be a potential predictor or target in ovarian cancer therapeutics.     

miR-146a interacting with lncRNA EPB41L4A-AS1 and lncRNA SNHG7 inhibits proliferation of bone marrow-derived mesenchymal stem cells

Emerging evidence suggests that microRNA plays a pivotal role in cell proliferation. Our previous research has certified that miR-146a attenuates osteoarthritis through the regulation of cartilage homeostasis. However, little information about the function of miR-146a in bone marrow-derived mesenchymal stem cells (BMSCs) proliferation and the underlying mechanism was available. Therefore, this study aims at investigating the role of miR-146a on the proliferation of BMSCs and the possible mechanisms involved. The function of miR-146a on BMSCs proliferation was studied through overexpression and knockdown of miR-146a or the indicated long noncoding RNAs (lncRNAs) in BMSCs and then the proliferation rate of the BMSCs were detected by Cell Counting Kit-8 assay, colony formation assay. Besides, flow cytometry was used to test the cell cycle state of BMSCs modified by overexpression or knockdown of miR-146a or lncRNA EPB41L4A-AS1 (EPB41L4A Antisense RNA 1) and small nucleolar RNA host gene 7 (SNHG7). The expression level of marker genes involved in modulating cell proliferation was evaluated by quantitative polymerase chain reaction and western blot analysis. We discovered that the knockdown of miR-146a significantly promoted BMSCs proliferation. Moreover, miR-146a could bind to and inhibit endogenous expression of EPB41L4A-AS1 and SNHG7. Further study demonstrated that overexpression of EPB41L4A-AS1 and SNHG7 significantly enhanced proliferation of BMSCs. For the first time, we certified that miR-146a suppressed BMSCs proliferation, but EPB41L4A-AS1 and SNHG7 promoted BMSCs proliferation in the present study. Mechanistically, miR-146a significantly inhibited BMSCs proliferation partly through miR-146a/EPB41L4A-AS1 SNHG7/cell proliferation signaling pathway axis.     

FER1L4/miR‐372/E2F1 works as a ceRNA system to regulate the proliferation and cell cycle of glioma cells

Long non‐coding RNAs have recently become a key regulatory factor for cancers, whereas FER1L4, a newly discovered long non‐coding RNA, has been mostly studied in gastric carcinoma and colon cancer cases. The functions and molecular mechanism of FER1L4 have been rarely reported in glioma malignant phenotypes. In this study, it was found that the expression of LncRNA FER1L4 is upregulated in high‐grade gliomas than in low‐grade cases and that a high expression of LncRNA FER1L4 predicts poor prognosis of gliomas. Meanwhile, in vitro study suggests that expression of FER1L4 with SiRNA knockdown obviously suppresses cell cycle and proliferation. It is further demonstrated by experiments that the FER1L4 knockdown suppresses growth of in vivo glioma. Besides, it is found in our study that LncRNA FER1L4 expression is positively correlated with E2F1 mRNA expression. After knockdown of FER1L4 expression, E2F1 expression is significantly down‐regulated, whereas the expression of miR‐372 is significantly up‐regulated; the up‐regulation of miR‐372 leads to significant down‐regulation of FER1L4 and E2F1 expression. In addition, it is also found that FER1L4 can be used as competitive endogenous RNA to interact or bind with miR‐371 and thereby up‐regulate E2F1, thus promoting the cycle and proliferation of glioma cells. It may be one of the molecular mechanisms in which FER1L4 plays its oncogene‐like role in gliomas.     

Role for DUSP1 (dual-specificity protein phosphatase 1) in the regulation of autophagy

Accumulating evidence suggests that mitogen-activated protein kinases (MAPKs) regulate macroautophagy/autophagy. However, the involvement of dual-specificity protein phosphatases (DUSPs), endogenous inhibitors for MAPKs, in autophagy remains to be determined. Here we report that DUSP1/MKP-1, the founding member of the DUSP family, plays a critical role in regulating autophagy. Specifically, we demonstrate that DUSP1 knockdown by shRNA in human ovarian cancer CAOV3 cells and knockout in murine embryonic fibroblasts, increases both basal and rapamycin-increased autophagic flux. Overexpression of DUSP1 had the opposite effect. Importantly, knockout of Dusp1 promoted phosphorylation of ULK1 at Ser555, and BECN1/Beclin 1 at Ser15, and the association of PIK3C3/VPS34, ATG14, BECN1 and MAPK, leading to the activation of the autophagosome-initiating class III phosphatidylinositol 3-kinase (PtdIns3K) complex. Furthermore, knockdown and pharmacological inhibitor studies indicated that DUSP1-mediated suppression of autophagy reflected inactivation of the MAPK1-MAPK3 members of the MAPK family. Knockdown of DUSP1 sensitized CAOV3 cells to rapamycin-induced antigrowth activity. Moreover, CAOV3-CR cells, a line that had acquired cisplatin resistance, exhibited an elevated DUSP1 level and were refractory to rapamycin-induced autophagy and cytostatic effects. Knockdown of DUSP1 in CAOV3-CR cells restored sensitivity to rapamycin. Collectively, this work identifies a previously unrecognized role for DUSP1 in regulating autophagy and suggests that suppression of DUSP1 may enhance the therapeutic activity of rapamycin.     

Annexin A3 depletion overcomes resistance to oxaliplatin in colorectal cancer via the MAPK signaling pathway

Colorectal cancer (CRC) is a common disease with high mortality and morbidity. Annexin A3 (ANXA3) belongs to the structurally homologous family of Ca²⁺ and phospholipid-binding proteins. This study aimed to investigate the effects and potential mechanisms of ANXA3 on oxaliplatin (Ox) resistance in CRC. We generated two human CRC cell lines (HCT116/Ox and SW480/Ox) with acquired Ox resistance and determined their resistance properties. ANXA3 expression and cell apoptosis, migration and invasion also were evaluated. We found that cell viability of HCT116/Ox and SW480/Ox was higher than that in parental cells in the presence of Ox. ANXA3 was highly expressed in HCT116/Ox and SW480/Ox cells. ANXA3 downregulation diminished cell survival, migration and invasion, while increased the apoptosis of HCT116 and SW480 with or without Ox. Moreover, depletion of ANXA3 reduced cell viability and BrdU incorporation, increased cell apoptosis and c-caspase 3 expression in HCT116/Ox with or without Ox. A transwell assay determined that knockdown of ANXA3 impeded the migration and invasion of HCT116/Ox and SW480/Ox cells. Additionally, phosphorylation of extracellular signal–regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) decreased upon ANXA3 depletion in HCT116/Ox cells, and ANXA3 silencing suppressed Ox-induced activation of ERK and JNK signaling pathway. ANXA3 downregulation reduced Ox resistance in CRC, and treatment with the ERK inhibitor PD098059 or JNK inhibitor SP600125 contributed to this process. These results indicate that silencing ANXA3 could overcome Ox resistance in CRC via the mitogen-activated protein kinase signaling pathway.     

Down-regulation of HIV-1 infection by inhibition of the MAPK signaling pathway

The human immunodeficiency virus type 1 (HIV-1) can interact with and exploit the host cellular machinery to replicate and propagate itself. Numerous studies have shown that the Mitogen-activated protein kinase (MAPK) signal pathway can positively regulate the replication of HIV-1, but exactly how each MAPK pathway affects HIV-1 infection and replication is not understood. In this study, we used the Extracellular signal-regulated kinase (ERK) pathway inhibitor, PD98059, the Jun N-terminal kinase (JNK) pathway inhibitor, SP600125, and the p38 pathway inhibitor, SB203580, to investigate the roles of these pathways in HIV-1 replication. We found that application of PD98059 results in a strong VSV-G pseudotyped HIV-1_NL4-3 luciferase reporter virus and HIV-1_NL4-3 virus inhibition activity. In addition, SB203580 and SP600125 also elicited marked VSV-G pseudotyped HIV-1_NL4-3 luciferase reporter virus inhibition activity but no HIV-1_NL4-3 virus inhibition activity. We also found that SB203580 and SP600125 can enhance the HIV-1 inhibition activity of PD98059 when cells were treated with all three MAPK pathway inhibitors in combination. Finally, we show that HIV-1 virus inhibition activity of the MAPK pathway inhibitors was the result of the negative regulation of HIV-1 LTR promoter activity.     

Overexpression of RYBP inhibits proliferation,invasion, and chemoresistance to cisplatin in anaplastic thyroid cancer cells via the EGFR pathway

Ring1 and YY1 binding protein (RYBP), a new member of the polycomb group protein family, has been reported to play an important role in various biological processes. Recently, more and more studies have demonstrated an implication of RYBP in cancer development. However, the specific role of RYBP in anaplastic thyroid cancer (ATC) remains unknown. In this study, we investigated for the first time the expression pattern and biological functions of RYBP in ATC. We showed that RYBP was lowly expressed in ATC tissues and cell lines. We also found that overexpression of RYBP inhibited ATC cell proliferation, invasion, and cisplatin resistance. Furthermore, we observed that upregulation of RYBP decreased the phosphorylation of EGFR and ERK1/2 in ATC cells. Taken together, our data indicated that RYBP might be considered as a promising therapeutic target for the treatment of ATC.     

Down-regulated LAMA4 inhibits oxidative stress-induced apoptosis of retinal ganglion cells through the MAPK signaling pathway in rats with glaucoma

Glaucoma is a neurodegenerative disorder that is generally accepted as the main cause of vision loss. In this study, we tested the hypothesis that laminin α4 (LAMA4) is implicated in glaucoma development by controlling apoptosis of retinal ganglion cells (RGCs) through the mitogen-activated protein kinase (MAPK) signaling pathway. Expression profiles and genes associated with glaucoma were searched to determine the objective gene. Intraocular pressure (IOP) rats model were established and IOP was measured. The mRNA and protein expression of LAMA4, JNK, p38 MAPK, ERK, Bcl-2, Bax, Caspase-9, and p53 was determined in concert with the treatment of H₂O₂, si-NC, or si-LAMA4 in cultured RGCs. Viability of RGCs, reactive oxygen species (ROS) and cell apoptosis was also measured. LAMA4 was selected as the study object because of its significant difference in two expression profiles. IOP of rats with glaucoma increased significantly after model establishment, and the LAMA4 protein expression in retinal tissue of rats with glaucoma was elevated. Down-regulation of LAMA4 could inhibit the mRNA and protein expression of LAMA4, JNK, p38 MAPK, ERK, Bax, Caspase-9, and p53, as well as restrain the apoptosis and ROS of RGCs, but improve Bcl-2 expression and viability of RGCs. Collectively, the obtained data supported that downregulated LAMA4 might reduce the oxidative stress-induced apoptosis of glaucoma RGCs by inhibiting the activation of the MAPK signaling pathway.     

Upregulated lncRNA-UCA1 contributes to metastasis of bile duct carcinoma through regulation of miR-122/CLIC1 and activation of the ERK/MAPK signaling pathway

In the present study, we aimed to identify specific lncRNAs and miRNAs, as well as mRNAs, involved in bile duct carcinoma (BDC) and to further explore the way in which lncRNA UCA1 regulates cell metastasis ability. Differentially expressed RNAs were screened out from the TCGA database. In in vitro experiments, qRT-PCR was used to measure lncRNA UCA1, miR-122 and CLIC1 expression. We performed a dual luciferase assay to validate the target relationships among UCA1, CLIC1 and miR-122. The cell migration ability was measured by a wound healing assay, and Transwell assays were applied to detect cell invasive ability. Western blot analysis was employed to detect the expression of related proteins in the MAPK signaling pathway. According to the bioinformatics analysis, lncRNA UCA1 and CLIC1 were both significantly upregulated in BDC, while the expression of miR-122 declined compared with the normal group. The target relationship among UCA1, CLIC1 and miR-122 was verified. UCA1 promoted BDC cell migration and invasiveness, while miR-122 inhibited their progression. CLIC1 served as the downstream target gene of miR-122 and had opposite effects. The ERK/MAPK signaling pathway was activated after upregulating UCA1. LncRNA-UCA1 promoted the metastasis of BDC cells by regulating the expression of miR-122 and its downstream gene mRNA CLIC1 and promoted the activation of the ERK/MAPK pathway, which expanded the horizons of targeted therapy of cholangiocarcinoma.     

UCH‐L1 promotes invasion of breast cancer cells through activating Akt signaling pathway

As a de‐ubiquitin enzyme, ubiquitin C‐terminal hydrolase (UCH)‐L1 has been shown to be overexpressed in several human cancers. However, the function of UCH‐L1 in invasion of breast cancers is still unclear. Here we report that the expression of UCH‐L1 is significantly higher in cancer cells with higher invasive ability. While ectopic UCH‐L1 expression failed to alter cell proliferation in MCF‐7 cells, it caused a significant upregulation of cellular invasion. Furthermore, siRNA mediated knockdown of UCH‐L1 led to suppression of invasion in UCH‐L1 overexpressing MCF‐7 cells. In order to identify molecular mechanisms underlying these observations, a novel in vitro proximity‐dependent biotin identification method was developed by fusing UCH‐L1 protein with a bacterial biotin ligase (Escherichia coli BirA R118G, BioID). Streptavidin magnetic beads pulldown assay revealed that UCH‐L1 can interact with Akt in MCF‐7 cells. Pulldown assay with His tagged recombinant UCH‐L1 protein and cell lysate from MCF‐7 cells further demonstrated that UCH‐L1 preferentially binds to Akt2 for Akt activation. Finally, we demonstrated that overexpression of UCH‐L1 led to activation of Akt as evidenced by upregulation of phosphorylated Akt. Thus, these findings demonstrated that UCH‐L1 promotes invasion of breast cancer cells and might serve as a potential therapeutic target for treatment of human patients with breast cancers.