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. 2021 Jul 23;11(8):380. doi: 10.1007/s13205-021-02922-5

Tumor-promoting function of PIMREG in glioma by activating the β-catenin pathway

Dekang Wang 1, Aili Hu 2, Hao Peng 1, Dongbo Li 2, Li Zhang 1,✉
PMCID: PMC8302699  PMID: 34458056

Abstract

Glioma is the most common primary brain tumor in adults with an adverse prognosis and obscure pathogenesis. PICALM interacting mitotic regulator protein (PIMREG) functions as an oncogene in multiple types of cancer, but its function in glioma remains unknown. The Gene Expression Profiling Interactive Analysis 2 (GEPIA2, http://gepia2.cancer-pku.cn/#index) showed that PIMREG expression in the glioma tissues was higher than that in normal brain tissues. Herein, cell counting kit-8 assay and flow cytometry analysis exhibited that overexpression of PIMREG significantly promoted the proliferation of glioma cells and the transition from G1 phase of the cell cycle to S phase. Wound-healing and transwell assays showed that overexpression of PIMREG markedly enhanced the migration and invasion of glioma cells. Western blot analysis revealed that overexpression of PIMREG increased the expression of cyclin D1, cyclin E, Vimentin, matrix metalloproteinase (MMP)-2, and MMP-9, but reduced the expression of E-cadherin. In addition, overexpression of PIMREG activated the β-catenin signaling pathway, as evidenced by the increased total and nuclear expression of β-catenin and the up-regulated expression of its downstream target c-myc. Furthermore, immunofluorescence staining further indicated the increased nuclear translocation of β-catenin in PIMREG-overexpressing cells. However, knockdown of PIMREG exerted opposite effects on glioma cells. Blockade of the β-catenin signaling by ICG-001 markedly impeded the promoting effects of PIMREG on glioma cell proliferation and invasion. In conclusion, PIMREG acts as a tumor promoter in glioma at least partly via activating the β-catenin signaling pathway. This study provides new insights into the molecular mechanism for glioma pathogenesis and treatment.

Keywords: PIMREG, Glioma, β-Catenin, Oncogene, Cell proliferation, Cell invasion

Introduction

Gliomas are the most common primary central nervous system tumors in adults. Despite efforts made in treatment strategies, patients may still develop tumor relapse and metastasis after surgical resection, leading to an unfavorable prognosis (Gusyatiner and Hegi 2018; Lapointe et al. 2018). The most malignant type of glioma, glioblastoma, is characterized by high invasiveness and proliferation rate (Ostrom et al. 2014; Wesseling and Capper 2018). To date, the pathogenesis of glioma remains unclear, and the tumorigenesis of glioma is considered to result from the combination of genetic and environmental factors (Chen et al. 2017; Ostrom et al. 2014). Thus, further investigations of the potential targets for the treatment of glioma are still of great significance.

PICALM interacting mitotic regulator protein (PIMREG, also known as CATS, FAM64A, and RCS1) is located on chromosome 17 band p13 and is considered as a marker of proliferation in both normal and malignant cells (Archangelo et al. 2008). PIMREG is identified as a substrate of anaphase-promoting complex/cyclosome (APC/C) and controls the metaphase-to-anaphase transition (Zhao et al. 2008). Moreover, the in vivo experiments reveal that PIMREG is a cell cycle promoter of hypoxic fetal cardiomyocytes in mice (Hashimoto et al. 2017). Recently, a series of studies have reported that PIMREG is up-regulated in tumor tissues and its up-regulation is associated with poorer overall survival in several types of cancer including osteosarcoma, pancreatic cancer, breast cancer, and cholangiocarcinoma (Jiang et al. 2019, 2020a, b; Jiao et al. 2019; Yao et al. 2019; Zhang et al. 2019). The in vitro experiments further confirm that overexpression of PIMREG markedly facilitates the aggressive development of breast cancer cells (Jiang et al. 2019) and cholangiocarcinoma cells (Jiang et al. 2020b), while knockdown of PIMREG significantly suppresses the proliferation and migration of breast cancer cells (Yao et al. 2019). Although the promoting effects of PIMREG have been reported in some types of tumor, the role of PIMREG in glioma has not been investigated yet.

The deregulation of epithelial-to-mesenchymal transition (EMT) and the β-catenin signaling pathway is reported to participate in the development of glioma (Sun et al. 2019; Wang et al. 2015, 2017; Zhang et al. 2018). The genes related to EMT process and the β-catenin signaling pathway were highly expressed in PIMREG-overexpressing breast cancer cells, indicating the activation of EMT process and the β-catenin signaling pathway induced by up-regulation of PIMREG (Zhang et al. 2019). However, whether EMT process and the β-catenin pathway are implicated in the role of PIMREG in glioma has not been verified.

In the present study, the expression of PIMREG was evaluated in the glioma tissues and cells. Furthermore, this study explored the effects and the underlying molecular mechanism of PIMREG on the proliferation, migration, and invasion of glioma cells, thus providing new insights into the pathogenesis and treatment for glioma.

Methods

Bioinformatics analysis

Gene Expression Profiling Interactive Analysis 2 (GEPIA2) online analysis tool (http://gepia2.cancer-pku.cn/#index) (Tang et al. 2019) was used for analysis of PIMREG expression level in the glioblastoma tissues (n = 163) and normal tissues (n = 207). The log2 [transcripts per million (TPM) + 1] was used to represent the expression level of PIMREG.

Cell culture and transfection

Human glioma cell lines (T98G, U251, SHG-44, and A172) were obtained from Procell (China). U251, SHG-44, and A172 were cultured in Dulbecco’s modified eagle medium (DMEM, Gibco, USA) with 10% fetal bovine serum (FBS, Tianhang, China) (Zhong et al. 2015). T98G was cultured in minimum essential medium (MEM, Gibco, USA) with 10% FBS (Shi et al. 2019). Each cell line was maintained in a 5% CO2 humidified incubator at 37 °C. PIMREG-overexpressing plasmids (OE-PIMREG), empty vector, small interfering RNA (siRNA) for PIMREG, and negative control (NC) siRNA were transfected into cells using Lipofectamine 3000 (Invitrogen, USA).

Cell counting kit-8 (CCK-8) assay

U251 and A172 cells were seeded at a density of 2 × 103 cells per well in 96-well plates (Chen et al. 2020). After cell transfection for 0, 24, 48, and 72 h or administration of β-catenin inhibitor ICG-001 (25 µM, Yuanye, China) for 24 h (Ma et al. 2005), CCK-8 reagent (Beyotime, China) was added into each well followed by a 2-h incubation at 37 °C, and the optical density (OD) values at a wavelength of 450 nm were detected using a microplate reader (BioTek, USA) to evaluate cell viability.

Flow cytometry analysis

According to the manufacturer’s protocol of cell cycle analysis kit (Beyotime, China), U251 and A172 cells were centrifuged at 1000×g for 5 min, washed twice with phosphate-buffered saline (PBS), and fixed in ice-cold 70% ethanol at 4 °C for 12 h. Afterwards, fixed cells were washed twice with cold PBS, resuspended in staining buffer, and incubated in propidium iodide (PI) and RNase A for 30 min at 37 °C in the darkroom. Finally, a flow cytometer (Acea Biosciences, USA) was used to analyze the cells in different phases of the cell cycle.

Wound-healing assay

Twenty-four hours after transfection, U251 and A172 cells were incubated in serum-free medium with mitomycin C (1 μg/ml, Sigma, USA). When cells approximately reached the confluence, the cells were scratched using a 200-μl pipette tip to create a uniform gap as previously described (Zhou et al. 2020). Cells were photographed at a fixed location at 0 and 24 h under the microscope (100× magnification), and the ratio of wound closure was taken as an indicator of cell migration.

Transwell invasion assay

Cell invasion was detected using the transwell chamber (Corning, USA) precoated with Matrigel (Corning, USA). Twenty-four hours after cell transfection or ICG-001 treatment, cells were seeded on the upper chamber (3 × 104/well) with serum-free medium, while the lower chamber was placed with medium supplemented with 10% FBS according to a previous study (Zhao et al. 2020). After incubation for 24 h, the invading cells were fixed with 4% paraformaldehyde (PFA, Aladdin, China), stained with 0.5% crystal violet (Amresco, USA), and counted under the microscope in five randomly selected fields (200× magnification).

Immunofluorescence

According to a previous study (Scharpenseel et al. 2019), cells grown on coverslips were fixed in 4% PFA (Sinopharm, China) for 15 min, washed with PBS, blocked with goat serum (Solarbio, China), and subsequently incubated with primary antibody anti-β-catenin (1:200, ABclonal, China) at 4 °C overnight. Next, cells were washed with PBS, incubated with Cy3-conjugated secondary antibody (1:200, Beyotime, China) at room temperature for 1 h and subsequently stained with 4′,6-diamidino-2-phenylindole (DAPI, Beyotime, China). Finally, the samples were observed and captured under the fluorescence microscope (400× magnification).

Reverse transcription PCR and quantitative real-time PCR (RT-qPCR)

Total mRNA was extracted using TRIpure reagent (BioTeke, China) and reversely transcribed into cDNA using Super M-MLV reverse transcriptase (BioTeke, China) as previously described (Zhao et al. 2021). Next, RT-qPCR was performed using SYBR Green and 2 × Taq PCR MasterMix (Solarbio, China) in an Exicycler 96 instrument (Bioneer, Korea) as previously described (Zhao et al. 2021). The primers were synthesized by Genscript (China) and the primer sequences were: PIMREG forward, 5′-GTGCTTTGGGTGCCGTGTC-3′; PIMREG reverse, 5′-ATCGCCGTAATGGGTGGG-3′; β-actin forward, 5′-GGCACCCAGCACAATGAA-3′, β-actin reverse, 5′-TAGAAGCATTTGCGGTGG-3′. Changes in the mRNA levels were calculated by the 2−ΔΔCt method with β-actin as the internal control.

Western blot

Total cell lysates were obtained using phenylmethanesulfonyl fluoride (PMSF) and cell lysis buffer (Beyotime, China) according to the instructions. The nuclear protein was prepared according to the manufacturer’s instructions of the nuclear protein extraction kit (Beyotime, China). The protein samples were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), subsequently transferred to polyvinylidene difluoride (PVDF) membranes (Millipore, USA). Next, the membranes were incubated overnight at 4 °C with various primary antibodies, and then incubated with horseradish peroxidase (HRP)-conjugated secondary antibodies (1:5000, Beyotime, China) at 37 °C for 45 min. Finally, protein bands were visualized using an enhanced chemiluminescence liquid (ECL, Beyotime, China) and the grey values were analyzed using Gel-Pro-Analyzer (Liuyi, China).

Primary antibodies were anti-PIMREG (1:1000, ABclonal, China), anti-Vimentin (1:1000, Affinity, China), anti-matrix metalloproteinase (MMP)-2 (1:500, Proteintech, China), anti-MMP-9 (1:1000, Proteintech, China), anti-cyclin D1 (1:500, ABclonal, China), anti-E-cadherin (1:500, ABclonal, China), anti-c-myc (1:500, ABclonal, China), anti-β-catenin (1:1000, ABclonal, China), anti-Histone H3 (1:2000, ABGENT, USA), and anti-β-actin (1:1000, Santa Cruz, USA) antibodies.

Statistical analysis

GraphPad Prism 8 software was used for statistical analyses. Data were presented as mean values ± standard deviation (SD) of three independent experiments. Unpaired t test, one-way analysis of variance (ANOVA), and two-way ANOVA were used to analyze the differences among groups. p values < 0.05 were considered statistically significant.

Results

PIMREG was up-regulated in human glioma

To investigate the role of PIMREG in glioma, the expression level of PIMREG in the glioblastoma and normal tissues was analyzed through GEPIA2 database. It was found that PIMREG expression was higher in the glioblastoma tissues than that in normal tissues (Fig. 1A), indicating that PIMREG might participate in the progression of glioma. Next, the mRNA and protein level of PIMREG in four glioma cell lines including T98G, U251, SHG-44, and A172 were detected. U251 cells with relatively low expression of PIMREG and A172 cells with relatively high expression of PIMREG were selected to perform the subsequent experiments (Fig. 1B, C).

Fig. 1.

Fig. 1

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PIMREG was up-regulated in human glioma. A PIMREG expression analysis in the glioblastoma tissues (n = 163) and normal tissues (n = 207) using GEPIA2 analyzing tool. B, C The expression level of PIMREG mRNA and protein was detected in human glioma cell lines (T98G, U251, SHG-44, and A172) by RT-qPCR and western blot, respectively. β-actin served as the internal control. *p < 0.05, **p < 0.01. PIMREG, PICALM interacting mitotic regulator protein. (n = 3)

PIMREG facilitated cell proliferation and cell cycle transition from G1 to S phase in glioma

To explore the function of PIMREG in gliomas, overexpression (Fig. 2A, B) or knockdown (Fig. 2C, D) of PIMREG was performed in human glioma cell lines. CCK-8 assay showed that PIMREG overexpression significantly promoted the proliferation of U251 cells (Fig. 2E), while PIMREG silencing dramatically suppressed the proliferation of A172 cells (Fig. 2F). Furthermore, cell cycle distribution in PIMREG-overexpressing/silencing cells was explored by flow cytometry analysis. The results demonstrated that up-regulation of PIMREG markedly reduced the percentage of cells in G1 phase and increased the percentage of cells in S phase, indicating a promoting effect on the cell cycle transition from G1 to S phase (Fig. 2G). Conversely, silencing of PIMREG significantly inhibited the cell cycle transition from G1 to S phase (Fig. 2H). Furthermore, the expression level of cell cycle markers cyclin D1 and cyclin E was increased by PIMREG overexpression (Fig. 2I) and decreased by PIMREG silencing (Fig. 2J). These results indicated that PIMREG exhibited promoting effects on glioma cell proliferation and cell cycle transition from G1 to S phase.

Fig. 2.

Fig. 2

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PIMREG promoted cell proliferation and cell cycle transition from G1 phase to S phase in glioma cells. U251 cells were transfected with PIMREG-overexpressing plasmids or empty vector. A172 cells were transfected with siRNA for PIMREG or NC siRNA. A–D The expression level of PIMREG mRNA and protein was detected by RT-qPCR and western blot 48 h after cell transfection. E, F At 0, 24, 48, and 72 h after cell transfection, cell viability was determined using CCK-8 assay. G, H Flow cytometry analysis was used to analyze cell cycle distribution. I, J Western blot was used to detect the expression levels of cell cycle markers including cyclin D1 and cyclin E. β-actin served as the loading control. *p < 0.05, **p < 0.01. PIMREG, PICALM interacting mitotic regulator protein. OE-PIMREG, PIMREG-overexpressing plasmids. siRNA, small interfering RNA. NC, negative control. (n = 3)

PIMREG promoted the migration and invasion of glioma cells

Since migration and invasion are essential in the development of glioma, the effects of PIMREG on the migration and invasion capability of glioma cells were further evaluated. Wound-healing assay showed that PIMREG overexpression significantly enhanced the migration of glioma cells (Fig. 3A), while PIMREG silencing dramatically inhibited cell migration capability (Fig. 3B). Consistently, transwell assay showed that the invasion of glioma cells was dramatically enhanced by PIMREG overexpression (Fig. 3C) and weakened by PIMREG down-regulation (Fig. 3D). Results from western blot assay further confirmed the promoting effects of PIMREG on the migration and invasion of glioma cells, as evidenced by the decreased expression level of E-cadherin and the increased expression level of MMP-2, MMP-9, and Vimentin in PIMREG-overexpressing cells (Fig. 3E), while the opposite results were observed in PIMREG-silencing cells (Fig. 3F). Taken together, these results supported that PIMREG markedly promoted the migration and invasion of glioma cells.

Fig. 3.

Fig. 3

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PIMREG promoted the migration and invasion of glioma cells. U251 cells were transfected with PIMREG-overexpressing plasmids or empty vector. A172 cells were transfected with siRNA for PIMREG or NC siRNA. A, B Cell migration was determined using wound-healing assay. Scale bare, 200 μm. C, D Cell invasion was determined using transwell assay. Scale bar, 100 μm. E, F Western blot was performed to detect the expression level of Vimentin, E-cadherin, MMP-2, and MMP-9. β-actin served as the loading control. **p < 0.01. PIMREG, PICALM interacting mitotic regulator protein. OE-PIMREG, PIMREG-overexpressing plasmids. siRNA, small interfering RNA. NC, negative control. (n = 3)

PIMREG activated the β-catenin signaling pathway in glioma cells

In PIMREG-overexpressing glioma cells, a significant increase in the total expression level of β-catenin and its downstream target c-myc and the nuclear expression level of β-catenin was observed, indicating activation of the β-catenin signaling pathway (Fig. 4A, B). However, PIMREG silencing dramatically decreased the total expression level of c-myc and β-catenin and the nuclear expression level of β-catenin (Fig. 4C, D). Similar results from immunofluorescence staining further proved the alterations in nuclear translocation of β-catenin induced by PIMREG overexpression or down-regulation (Fig. 4E, F). These results indicated that the β-catenin signaling pathway might participate in the function of PIMREG in glioma cells.

Fig. 4.

Fig. 4

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PIMREG activated the β-catenin signaling pathway in glioma cells. U251 cells were transfected with PIMREG-overexpressing plasmids or empty vector. A172 cells were transfected with siRNA for PIMREG or NC siRNA. A, C Western blot analysis was used to evaluate the total expression level of β-catenin and c-myc. β-actin served as the loading control. B, D Western blot analysis was used to evaluate the nuclear expression level of β-catenin. Histone H3 served as the loading control. E, F The intracellular location of β-catenin was determined using immunofluorescence analysis (β-catenin, red; DAPI, blue). Scale bare, 50 μm. PIMREG, PICALM interacting mitotic regulator protein. OE-PIMREG, PIMREG-overexpressing plasmids. siRNA, small interfering RNA. NC, negative control. (n = 3)

The β-catenin inhibitor ICG-001 impeded the promoting effects of PIMREG on glioma cell proliferation and invasion

To further investigate the role of the β-catenin signaling pathway in the promoting effects of PIMREG on glioma cell proliferation and invasion, 24 h after cell transfection, β-catenin inhibitor ICG-001 was used to treat glioma cells for 24 h. CCK-8 assay and western blot analysis showed that ICG-001 treatment dramatically reversed the promoting effects of PIMREG overexpression on cell proliferation (Fig. 5A) and the increased protein level of MMP-2, Vimentin, and cyclin D1 induced by PIMREG overexpression in glioma cells (Fig. 5B). Transwell assay proved that ICG-001 reversed the invasion capability of glioma cells enhanced by overexpression of PIMREG (Fig. 5C). Taken together, these results showed that the blockade of the β-catenin pathway significantly impeded the function of PIMREG in glioma cell proliferation and invasion, indicating that PIMREG promoted the proliferation and invasion of glioma cells at least partly through regulating the β-catenin signaling pathway.

Fig. 5.

Fig. 5

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The β-catenin inhibitor ICG-001 impeded the promoting effects of PIMREG on glioma cell proliferation and invasion. U251 cells were transfected with PIMREG-overexpressing plasmids and then treated with 25 µM the β-catenin inhibitor ICG-001 for 24 h at 24 h after cell transfection. A CCK-8 assay was used to detect cell viability. B Western blot was performed to evaluate the expression level of MMP-2, Vimentin, and cyclin D1. β-actin served as the loading control. C Transwell assay was used to measure the capability of cell invasion. Scale bare, 100 μm. *p < 0.05, **p < 0.01. PIMREG, PICALM interacting mitotic regulator protein. OE-PIMREG, PIMREG-overexpressing plasmids. (n = 3)

Discussion

The present study showed that PIMREG was highly expressed in the glioma tissues. The in vitro experiments exhibited that PIMREG overexpression significantly promoted the proliferation, migration, and invasion of glioma cells, increased the total and nuclear expression of β-catenin, and up-regulated the expression of its downstream target c-myc, whereas PIMREG silencing exerted the opposite effects on glioma cells. Furthermore, β-catenin inhibitor ICG-001 markedly impeded the promoting effects of PIMREG on glioma cell proliferation and invasion.

PIMREG is well recognized as a marker of proliferation in both normal and malignant cells (Archangelo et al. 2008). Over the past decade, researchers have shown an increased interest in the correlation between PIMREG and tumor development. PIMREG was found to be significantly up-regulated in multiple types of cancer and led to an unfavorable overall survival (Jiang et al. 2020a, b; Jiao et al. 2019; Yao et al. 2019). The in vitro experiments showed that overexpression of PIMREG significantly promoted the tumorigenicity of tumor cells including proliferation, invasion, and migration, while knockdown of PIMREG exhibited the opposite effects (Jiang et al. 2020a; Yao et al. 2019; Zhang et al. 2019). Moreover, the experimental metastasis mouse model of breast cancer showed that the mice-bearing PIMREG-transduced tumors displayed prominent lung metastasis and shorter survival time, confirming the enhancement of tumor aggressiveness induced by the elevated PIMREG expression (Jiang et al. 2019). PIMREG was significantly up-regulated in the glioma tissues. The present study verified the promoting effects of PIMREG on the proliferation of glioma cells for the first time.

Cell cycle relative markers have been recognized as the key drivers of malignant transformation in multiple types of tumor including glioma (Arato-Ohshima and Sawa 1999; Fraczek et al. 2008; Gautschi et al. 2007; Hui et al. 2013; Pang et al. 2020). Cyclin D1 and cyclin E are cell cycle specific proteins that interact with cyclin-dependent kinases (CDKs) and regulate multiple downstream molecules like E2F, thus playing an important role in the transition from G1 to S phase (Johnson and Walker 1999; Pang et al. 2020). Highly expressed cyclin E was reported to accelerate transition from G1 phase of the cell cycle to S phase and prolong S phase (Fraczek et al. 2008). In the present study, PIMREG up-regulation dramatically increased the percentage of cells in S phase and the expression level of cyclin D1 and cyclin E, as well as decreased the percentage of cells in G1 phase, while the silence of PIMREG showed opposite effects. Consistently, Yao et al. (2019) found that knockdown of PIMREG inhibited the proliferation of breast cancer cells by prolonging the G1-cell phase. Taken together, PIMREG might play an important role in cell cycle and therefore exhibit its promoting effects on tumor cell proliferation.

High invasiveness is one of the most important characteristics of high-grade glioma. EMT is a conserved process in which cells lose their epithelial characteristics and acquire mesenchymal properties. Selective transcriptional factors including ZEB, Snail, Slug, and the Twist family can trigger this process, leading to the down-regulated expression of proteins that are required to maintain the cell polarity and cell adhesion, such as E-cadherin, occludins, and claudins (Dongre and Weinberg 2019). These transcriptional factors could also enhance the expression of proteins associated with the mesenchymal traits, such as N-cadherin, Vimentin, β-integrin, and the extracellular matrix-degrading proteases like MMPs (Nieto et al. 2016). In addition to the important role in normal biological processes including embryogenesis and wound healing, EMT is also involved in the formation of an advantageous microenvironment for cancer progression and metastasis. In 92 glioma cases, the weak expression of E-cadherin and the high expression of N-cadherin were observed by immunohistochemical analysis and western blot analysis (Noh et al. 2017). Besides, many factors involved in the progression of glioma also influenced the expression level of the representative EMT markers (Kahlert et al. 2013; Lee et al. 2017). In the present study, overexpression of PIMREG significantly promoted the migration and invasion of glioma cells, reduced the level of E-cadherin, and up-regulated the level of Vimentin, MMP-2, and MMP-9. The similar results were also observed in PIMREG-overexpressing breast cancer cells, and knockdown of PIMREG markedly suppressed the expression of EMT transcriptional factors including Snail, Twist, and Slug (Yao et al. 2019; Zhang et al. 2019). These findings strongly indicated that PIMREG might participate in tumor progression via regulating the EMT process.

The aberrant β-catenin signaling pathway is closely related to various types of tumor (Huo et al. 2019). In normal cells, β-catenin mostly binds to E-cadherin on the cell membrane to maintain the cell adhesion, and a small part in the cytoplasm binds to the degrading complex consisting APC, GSK-3β, and Axin. The Wnt cascade signaling could protect the β-catenin from degradation, leading to the accumulation of β-catenin in the cytosol and subsequent translocation to the nucleus. Afterwards, a series of oncogenes including cyclin D1 and c-myc were up-regulated, thus facilitating the cell proliferation and tumor development (He et al. 2019). In the previous studies, the β-catenin signaling pathway-dependent manner was reported to participate in promoting the proliferation and metastasis of human glioma (Huo et al. 2019; Wang et al. 2017). Besides, several studies confirmed that the inactivation of the β-catenin signaling pathway suppressed the malignant progression of glioma (Meng et al. 2019; Xu et al. 2019a). The activation of the β-catenin signaling pathway in PIMREG-overexpressing glioma cells was verified for the first time. Furthermore, treatment of β-catenin signaling inhibitor ICG-001 (Hirakawa et al. 2019) significantly reversed the promoting effects of PIMREG on glioma cell proliferation and invasion. These results strongly supported that PIMREG participated in the progression of glioma through activating the β-catenin signaling.

In addition to the β-catenin signaling pathway, the relation of PIMREG to other signaling pathways has also been reported. For instance, through modulating the transcription activity of signal transducer and activator of transcription 3 (STAT3), PIMREG could regulate the T helper cell differentiation in colitis and inflammation-associated cancer (Xu et al. 2019b). Besides, overexpression of PIMREG was reported to promote the aggressiveness of breast cancer through constitutive activation of the nuclear factor κB (NF-κB) signaling (Jiang et al. 2019). Interestingly, the NF-κB and STAT3 signaling pathways were also involved in the progression and mesenchymal transformation of glioma (Carro et al. 2010; Edwards et al. 2011). However, the relationship between these signaling pathways and PIMREG has not been investigated in glioma. Therefore, further studies on the signaling network involved in the role of PIMREG in glioma are needed.

Conclusion

In summary, the present study revealed the promoting effects of PIMREG on the proliferation, migration, and invasion of glioma cells through regulating the β-catenin signaling pathway. This study may be helpful for exploring the pathogenesis of gliomas and new targets for glioma treatment.

Author contributions

Conceptualization: LZ; methodology: LZ and DW; formal analysis and investigation: DW, AH, and HP; writing—original draft preparation: DW; writing—review and editing: DW; supervision: DL.

Funding

No funding was received to assist with the preparation of this manuscript.

Availability of data and materials

The data analyzed in the present study are available from the corresponding author on reasonable request.

Code availability

Not applicable.

Declarations

Conflict of interest

The authors have no conflicts of interest to declare that are relevant to the content of this article.

References

  1. Arato-Ohshima T, Sawa H. Over-expression of cyclin D1 induces glioma invasion by increasing matrix metalloproteinase activity and cell motility. Int J Cancer. 1999;83:387–392. doi: 10.1002/(sici)1097-0215(19991029)83:3<387::aid-ijc15>3.0.co;2-o. [DOI] [PubMed] [Google Scholar]
  2. Archangelo LF, et al. The CALM and CALM/AF10 interactor CATS is a marker for proliferation. Mole Oncol. 2008;2:356–367. doi: 10.1016/j.molonc.2008.08.001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Carro MS, et al. The transcriptional network for mesenchymal transformation of brain tumours. Nature. 2010;463:318–325. doi: 10.1038/nature08712. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chen R, Smith-Cohn M, Cohen AL, Colman H. Glioma subclassifications and their clinical significance. Neurotherapeutics. 2017;14:284–297. doi: 10.1007/s13311-017-0519-x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chen Y, Yuan S, Ning T, Xu H, Guan B. SNHG7 facilitates glioblastoma progression by functioning as a molecular sponge for MicroRNA-449b-5p and thereby increasing MYCN expression. Technol Cancer Res Treat. 2020;19:1533033820945802. doi: 10.1177/1533033820945802. [DOI] [PMC free article] [PubMed] [Google Scholar] [Retracted]
  6. Dongre A, Weinberg RA. New insights into the mechanisms of epithelial-mesenchymal transition and implications for cancer. Nat Rev Mol Cell Biol. 2019;20:69–84. doi: 10.1038/s41580-018-0080-4. [DOI] [PubMed] [Google Scholar]
  7. Edwards LA, et al. Effect of brain- and tumor-derived connective tissue growth factor on glioma invasion. J Natl Cancer Inst. 2011;103:1162–1178. doi: 10.1093/jnci/djr224. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Fraczek M, Wozniak Z, Ramsey D, Zatonski T, Krecicki T. Clinicopathologic significance and prognostic role of cyclin E and cyclin A expression in laryngeal epithelial lesions. Acta Otolaryngol. 2008;128:329–334. doi: 10.1080/00016480701487742. [DOI] [PubMed] [Google Scholar]
  9. Gautschi O, Ratschiller D, Gugger M, Betticher DC, Heighway J. Cyclin D1 in non-small cell lung cancer: a key driver of malignant transformation. Lung Cancer. 2007;55:1–14. doi: 10.1016/j.lungcan.2006.09.024. [DOI] [PubMed] [Google Scholar]
  10. Gusyatiner O, Hegi ME. Glioma epigenetics: From subclassification to novel treatment options. Semin Cancer Biol. 2018;51:50–58. doi: 10.1016/j.semcancer.2017.11.010. [DOI] [PubMed] [Google Scholar]
  11. Hashimoto K, et al. Fam64a is a novel cell cycle promoter of hypoxic fetal cardiomyocytes in mice. Sci Rep. 2017;7:4486. doi: 10.1038/s41598-017-04823-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. He L, Zhou H, Zeng Z, Yao H, Jiang W, Qu H. Wnt/β-catenin signaling cascade: a promising target for glioma therapy. J Cell Physiol. 2019;234:2217–2228. doi: 10.1002/jcp.27186. [DOI] [PubMed] [Google Scholar]
  13. Hirakawa T, Nasu K, Miyabe S, Kouji H, Katoh A, Uemura N, Narahara H. β-catenin signaling inhibitors ICG-001 and C-82 improve fibrosis in preclinical models of endometriosis. Sci Rep. 2019;9:20056. doi: 10.1038/s41598-019-56302-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hui W, Yuntao L, Lun L, WenSheng L, ChaoFeng L, HaiYong H, Yueyang B. MicroRNA-195 inhibits the proliferation of human glioma cells by directly targeting cyclin D1 and cyclin E1. PLoS ONE. 2013;8:e54932. doi: 10.1371/journal.pone.0054932. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Huo XY, Zhang XY, Yuan F, Zhao XY, You BA. HOXB7 promotes proliferation and metastasis of glioma by regulating the Wnt/β-catenin pathway. Eur Rev Med Pharmacol Sci. 2019;23:2476–2485. doi: 10.26355/eurrev_201903_17395. [DOI] [PubMed] [Google Scholar]
  16. Jiang L, et al. Overexpression of PIMREG promotes breast cancer aggressiveness via constitutive activation of NF-κB signaling. EBioMedicine. 2019;43:188–200. doi: 10.1016/j.ebiom.2019.04.001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jiang Y, Zhou C, Gao Q, Yin ZQ, Wang J, Mu H, Yan J. FAM64A promotes osteosarcoma cell growth and metastasis and is mediated by miR-493. J Oncol. 2020;2020:2518297. doi: 10.1155/2020/2518297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Jiang ZM, Li HB, Chen SG. PIMREG, a marker of proliferation, facilitates aggressive development of cholangiocarcinoma cells partly through regulating cell cycle-related markers. Technol Cancer Res Treat. 2020;19:1533033820979681. doi: 10.1177/1533033820979681. [DOI] [PMC free article] [PubMed] [Google Scholar] [Retracted]
  19. Jiao Y, Fu Z, Li Y, Zhang W, Liu Y. Aberrant FAM64A mRNA expression is an independent predictor of poor survival in pancreatic cancer. PLoS ONE. 2019;14:e0211291. doi: 10.1371/journal.pone.0211291. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Johnson DG, Walker CL. Cyclins and cell cycle checkpoints. Annu Rev Pharmacol Toxicol. 1999;39:295–312. doi: 10.1146/annurev.pharmtox.39.1.295. [DOI] [PubMed] [Google Scholar]
  21. Kahlert UD, Nikkhah G, Maciaczyk J. Epithelial-to-mesenchymal(-like) transition as a relevant molecular event in malignant gliomas. Cancer Lett. 2013;331:131–138. doi: 10.1016/j.canlet.2012.12.010. [DOI] [PubMed] [Google Scholar]
  22. Lapointe S, Perry A, Butowski NA. Primary brain tumours in adults. Lancet. 2018;392:432–446. doi: 10.1016/s0140-6736(18)30990-5. [DOI] [PubMed] [Google Scholar]
  23. Lee W, et al. Epimedium koreanum Nakai inhibits PMA-induced cancer cell migration and invasion by modulating NF-κB/MMP-9 signaling in monomorphic malignant human glioma cells. Oncol Rep. 2017;38:3619–3631. doi: 10.3892/or.2017.6043. [DOI] [PubMed] [Google Scholar]
  24. Ma H, Nguyen C, Lee KS, Kahn M. Differential roles for the coactivators CBP and p300 on TCF/beta-catenin-mediated survivin gene expression. Oncogene. 2005;24:3619–3631. doi: 10.1038/sj.onc.1208433. [DOI] [PubMed] [Google Scholar]
  25. Meng Y, Shang FR, Zhu YL. MiR-491 functions as a tumor suppressor through Wnt3a/β-catenin signaling in the development of glioma. Eur Rev Med Pharmacol Sci. 2019;23:10899–10907. doi: 10.26355/eurrev_201912_19793. [DOI] [PubMed] [Google Scholar]
  26. Nieto MA, Huang RY, Jackson RA, Thiery JP. EMT: 2016. Cell. 2016;166:21–45. doi: 10.1016/j.cell.2016.06.028. [DOI] [PubMed] [Google Scholar]
  27. Noh MG, et al. Prognostic significance of E-cadherin and N-cadherin expression in gliomas. BMC Cancer. 2017;17:583. doi: 10.1186/s12885-017-3591-z. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Ostrom QT, et al. The epidemiology of glioma in adults: a “state of the science” review. Neuro Oncol. 2014;16:896–913. doi: 10.1093/neuonc/nou087. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Pang W, Li Y, Guo W, Shen H. Cyclin E: a potential treatment target to reverse cancer chemoresistance by regulating the cell cycle. Am J Transl Res. 2020;12:5170–5187. [PMC free article] [PubMed] [Google Scholar]
  30. Scharpenseel H, et al. EGFR and HER3 expression in circulating tumor cells and tumor tissue from non-small cell lung cancer patients. Sci Rep. 2019;9:7406. doi: 10.1038/s41598-019-43678-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Shi F, Shi Z, Zhao Y, Tian J. CircRNA hsa-circ-0014359 promotes glioma progression by regulating miR-153/PI3K signaling. Biochem Biophys Res Commun. 2019;510:614–620. doi: 10.1016/j.bbrc.2019.02.019. [DOI] [PubMed] [Google Scholar]
  32. Sun Y, Peng X, Li Y, Ma H, Li D, Shi X. The effects of histamine H1 type receptor (H1R) in regulating osteoblastic cell differentiation and mineralization. Artif Cell Nanomed B. 2019;47:1281–1287. doi: 10.1080/21691401.2019.1596924. [DOI] [PubMed] [Google Scholar]
  33. Tang Z, Kang B, Li C, Chen T, Zhang Z. GEPIA2: an enhanced web server for large-scale expression profiling and interactive analysis. Nucleic Acids Res. 2019;47:W556–W560. doi: 10.1093/nar/gkz430. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Wang Z, Zhang S, Siu TL, Huang S. Glioblastoma multiforme formation and EMT: role of FoxM1 transcription factor. Curr Pharm Des. 2015;21:1268–1271. doi: 10.2174/1381612821666141211115949. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Wang Y, et al. β-catenin-mediated YAP signaling promotes human glioma growth. J Exp Clin Cancer Res. 2017;36:136. doi: 10.1186/s13046-017-0606-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Wesseling P, Capper D. WHO 2016 classification of gliomas. Neuropathol Appl Neurobiol. 2018;44:139–150. doi: 10.1111/nan.12432. [DOI] [PubMed] [Google Scholar]
  37. Xu H, et al. Mesenchymal stem cell-derived exosomal microRNA-133b suppresses glioma progression via Wnt/β-catenin signaling pathway by targeting EZH2. Stem Cell Res Ther. 2019;10:381. doi: 10.1186/s13287-019-1446-z. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Xu ZS, et al. FAM64A positively regulates STAT3 activity to promote Th17 differentiation and colitis-associated carcinogenesis. Proc Natl Acad Sci USA. 2019;116:10447–10452. doi: 10.1073/pnas.1814336116. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Yao Z, et al. Knockdown of FAM64A suppresses proliferation and migration of breast cancer cells. Breast Cancer. 2019;26:835–845. doi: 10.1007/s12282-019-00991-2. [DOI] [PubMed] [Google Scholar]
  40. Zhang J, et al. LGR5, a novel functional glioma stem cell marker, promotes EMT by activating the Wnt/β-catenin pathway and predicts poor survival of glioma patients. J Exp Clin Cancer Res. 2018;37:225. doi: 10.1186/s13046-018-0864-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Zhang J, et al. Up-regulation of FAM64A promotes epithelial-to-mesenchymal transition and enhances stemness features in breast cancer cells. Biochem Biophys Res Commun. 2019;513:472–478. doi: 10.1016/j.bbrc.2019.03.207. [DOI] [PubMed] [Google Scholar]
  42. Zhao WM, Coppinger JA, Seki A, Cheng XL, Yates JR, 3rd, Fang G. RCS1, a substrate of APC/C, controls the metaphase to anaphase transition. Proc Natl Acad Sci USA. 2008;105:13415–13420. doi: 10.1073/pnas.0709227105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Zhao XF, Yang YS, Park YK. HOXC9 overexpression is associated with gastric cancer progression and a prognostic marker for poor survival in gastric cancer patients. Int J Clin Oncol. 2020;25:2044–2054. doi: 10.1007/s10147-020-01772-0. [DOI] [PubMed] [Google Scholar]
  44. Zhao W, Zhao J, Guo X, Feng Y, Zhang B, Tian L. LncRNA MT1JP plays a protective role in intrahepatic cholangiocarcinoma by regulating miR-18a-5p/FBP1 axis. BMC Cancer. 2021;21:142. doi: 10.1186/s12885-021-07838-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Zhong D, et al. LRG1 modulates invasion and migration of glioma cell lines through TGF-beta signaling pathway. Acta Histochem. 2015;117:551–558. doi: 10.1016/j.acthis.2015.05.001. [DOI] [PubMed] [Google Scholar]
  46. Zhou Y, An H, Wu G. MicroRNA-6071 suppresses glioblastoma progression through the inhibition of PI3K/AKT/mTOR pathway by binding to ULBP2. Onco Targets Ther. 2020;13:9429–9441. doi: 10.2147/OTT.S265791. [DOI] [PMC free article] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Data Availability Statement

The data analyzed in the present study are available from the corresponding author on reasonable request.

Not applicable.

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