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. 2019 Mar 29;17(1):30.
doi: 10.1186/s12964-019-0344-4.

PRMT5 enhances tumorigenicity and glycolysis in pancreatic cancer via the FBW7/cMyc axis

Yi Qin  1   2   3   4   5 , Qiangsheng Hu  1   2   3   4 , Jin Xu  1   2   3   4 , Shunrong Ji  1   2   3   4 , Weixing Dai  5 , Wensheng Liu  1   2   3   4 , Wenyan Xu  1   2   3   4 , Qiqing Sun  1   2   3   4 , Zheng Zhang  1   2   3   4 , Quanxing Ni  1   2   3   4 , Bo Zhang  1   2   3   4 , Xianjun Yu  6   7   8   9 , Xiaowu Xu  10   11   12   13
Affiliations

PRMT5 enhances tumorigenicity and glycolysis in pancreatic cancer via the FBW7/cMyc axis

Yi Qin et al. Cell Commun Signal. .

Abstract

Background: The epigenetic factor protein arginine methyltransferase 5 (PRMT5) has been reported to play vital roles in a wide range of cellular processes, such as gene transcription, genomic organization, differentiation and cell cycle control. However, its role in pancreatic cancer remains unclear VSports手机版. Our study aimed to investigate the roles of PRMT5 in pancreatic cancer prognosis and progression and to explore the underlying molecular mechanism. .

Methods: Real-time PCR, immunohistochemistry and analysis of a dataset from The Cancer Genome Atlas (TCGA) were performed to study the expression of PRMT5 at the mRNA and protein levels in pancreatic cancer. Cell proliferation assays, including cell viability, colony formation ability and subcutaneous mouse model assays, were utilized to confirm the role of PRMT5 in cell proliferation and tumorigenesis. A Seahorse extracellular flux analyzer, a glucose uptake kit, a lactate level measurement kit and the measurement of 18F-FDG (fluorodeoxyglucose) uptake by PET/CT (positron emission tomography/computed tomography) imaging were used to verify the role of PRMT5 in aerobic glycolysis, which sustains cell proliferation V体育安卓版. The regulatory effect of PRMT5 on cMyc, a master regulator of oncogenesis and aerobic glycolysis, was explored by quantitative PCR and protein stability measurements. .

Results: PRMT5 expression was significantly upregulated in pancreatic cancer tissues compared with that in adjacent normal tissues. Clinically, elevated expression of PRMT5 was positively correlated with worse overall survival in pancreatic cancer patients. Silencing PRMT5 expression inhibited the proliferation of pancreatic cancer cells both in vitro and in vivo. Moreover, PRMT5 regulated aerobic glycolysis in vitro in cell lines, in vivo in pancreatic cancer patients and in a xenograft mouse model used to measure 18F-FDG uptake. We found that mechanistically, PRMT5 posttranslationally regulated cMyc stability via F-box/WD repeat-containing protein 7 (FBW7), an E3 ubiquitin ligase that controls cMyc degradation V体育ios版. Moreover, PRMT5 epigenetically regulated the expression of FBW7 in pancreatic cancer cells. .

Conclusions: The present study demonstrated that PRMT5 epigenetically silenced the expression of the tumor suppressor FBW7, leading to increased cMyc levels and the subsequent enhancement of the proliferation of and aerobic glycolysis in pancreatic cancer cells. The PRMT5/FBW7/cMyc axis could be a potential therapeutic target for the treatment of pancreatic cancer VSports最新版本. .

Keywords: Aerobic glycolysis; FBW7; PRMT5; Pancreatic cancer; cMyc V体育平台登录. .

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Conflict of interest statement

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Not applicable.

Ethics approval and consent to participate

The research protocol was reviewed and approved by the Ethics Committee of FDUSCC VSports注册入口. This study complied with the Animal Care guidelines of FDUSCC.

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All of the authors listed consented to publication.

Competing interests

The authors declare that they have no competing interests.

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Figures (V体育2025版)

Fig. 1
Fig. 1
PRMT5 expression is upregulated and indicates a worse prognosis in pancreatic cancer patients a. PRMT5 expression was upregulated in PDAC tumor samples compared with that in normal adjacent samples, as indicated by real-time PCR and paired t-test analysis. b. Immunohistochemical staining of PRMT5 in PDAC tumor samples and paired adjacent normal tissues. c. PRMT5 protein levels were increased in tumor samples, as reflected by IHC analysis. d. IHC scoring of PRMT5 expression in pancreatic cancer tissues, which were divided into the low PRMT5 expression and high PRMT5 expression subgroups. e. Patients in the FDUSCC cohort with high PRMT5 expression had shorter overall survival times than those with low PRMT5 expression. f. PRMT5 expression in the TCGA-PAAD RNA-seq dataset. Analysis results demonstrated that high PRMT5 expression predicts a shorter OS time. g. Patients with high PRMT5 expression had shorter DFS times than patients with low PRMT5 expression
Fig. 2
Fig. 2
PRMT5 regulates the proliferation and tumorigenicity of pancreatic cancer cells a. PRMT5 expression in pancreatic cancer cell lines. b. Quantitative real-time results confirmed the silencing efficiency of shRNAs targeting PRMT5 in MIA PaCa-2 and SW1990 cells. c. Immunoblotting with an anti-PRMT5 antibody further confirmed that PRMT5 was efficiently silenced in MIA PaCa-2 and SW1990 cells. d. Silencing of PRMT5 decreased the viability of MIA PaCa-2 and SW1990 cells (n = 3, p < 0.05 and p < 0.01 for the shPRMT5A and shPRMT5B groups, respectively). e-f. Silencing of PRMT5 inhibited the colony formation capacity of MIA PaCa-2 and SW1990 cells (n = 3, p < 0.01). g-h. The subcutaneous xenograft mouse model showed that knockdown of PRMT5 decreased the tumor formation capacity of SW1990 cells (n = 4, p < 0.001). i. Representative images of immunohistochemical staining for Ki-67 and PRMT5
Fig. 3
Fig. 3
PRMT5 regulates aerobic glycolysis in vitro and in vivo a. The glucose uptake assay results suggested that PRMT5 knockdown decreased the glucose intake capacity of MIA PaCa-2 and SW1990 cells. b. The lactate level assay results indicated a decrease in lactate levels when PRMT5 was silenced in MIA PaCa-2 and SW1990 cells. c-d. A Seahorse extracellular flux analyzer was used to measure the ECAR, and the results indicated that decreased PRMT5 expression resulted in increased glycolysis in and glycolytic capacity of pancreatic cancer cells. e. Representative PRMT5 staining and 18F-FDG-PET/CT images for the indicated patients. f. Patients with high PRMT5 expression exhibited high levels of SUVmax values obtained by 18F-FDG-PET/CT imaging. g. Representative micro-PET/CT image of the subcutaneous xenograft mouse model. h. PRMT5 knockdown decreased 18F-FDG uptake by subcutaneous tumors, as reflected by the SUVmax value, reinforcing the roles of PRMT5 in promoting glycolysis in vivo
Fig. 4
Fig. 4
PRMT5 regulates cMyc at the posttranslational level a. Decreased PRMT5 expression resulted in a reduction in the cMyc protein level but had little impact on the HIF1α protein level. b. Knockdown of PRMT5 had a subtle impact on cMyc mRNA levels. c. We treated PRMT5-silenced pancreatic cancer cells with the proteasome inhibitor MG132 and measured the level of cMyc. The western blot analysis results demonstrated that MG132 treatment reversed the down-regulation of cMyc caused by PRMT5 knockdown, suggesting that PRMT5 regulates cMyc protein stability. d. PRMT5-silenced MIA PaCa-2 and SW1990 cells and the corresponding control cells were treated with CHX for the indicated durations, and the protein levels of cMyc were determined by western blotting. e. PRMT5 knockdown decreased the protein stability of cMyc in MIA PaCa-2 and SW1990 cells. f. In HPDE cells, overexpression of PRMT5 increased cMyc expression at the protein level, while the dominant-negative mutant of PRMT5 (PRMT5DN) had no such effect. g-h. Overexpressing PRMT5 in HPDE cells could stabilize cMyc, while PRMT5DN had no such effect
Fig. 5
Fig. 5
PRMT5-mediated epigenetic silencing of FBW7 leads to increased cMyc levels a. In PRMT5-silenced MIA PaCa-2 and SW1990 cells, the mRNA levels of FBW7 were increased. b. PRMT5 knockdown increased FBW7 protein levels. c. PRMT5 expression was negatively correlated with FBW7 expression in the TCGA-PAAD dataset of pancreatic cancer patients. d. In HPDE cells, overexpressing PRMT5 decreased FBW7 mRNA levels, but PRMT5DN did not regulate FBW7 expression. e. Overexpressing PRMT5 in HPDE cells decreased FBW7 protein levels, but PRMT5DN exerted no impact on FBW7 protein levels. f. The results of the dual luciferase assay in HEK-293 T cells showed that although PRMT5 suppressed FBW7 promoter activity, PRMT5DN did not significantly regulate FBW7 promoter activity. g-h. The ChIP assay results demonstrated that PRMT5 occupied the promoter region enriched with CpG islands. i. PRMT5 knockdown decreased the occupancy of the heterochromatin markers H4R3me2 and H3K9me3, and the euchromatin marker H3K9ac, which reflects active transcription, was increased in MIA PaCa-2 and SW1990 cells with PRMT5 knockdown. j. Finally, we performed ChIP in HPDE cells with low PRMT5 expression. The ChIP results showed that PRMT5 increased the occupancy of heterochromatin markers such as H4MR3me2 and H3K9me3 and decreased that of the active chromatin marker H3K9ac decreased. However, the transferase-dead PRMT5DN mutant had no such effect
Fig. 6
Fig. 6
PRMT5 regulates proliferation and aerobic glycolysis via the FBW7/cMyc axis a. Overexpression of wild-type FBW7 in PRMT5-overexpressing HPDE cells attenuated the increase in the cMyc protein level, but the FBW7R465H mutant had no such effect. b. The CCK-8 assay results demonstrated that wild-type FBW7 decreased the increase in cell viability caused by PRMT5, while the FBW7R465H mutant, which lacked enzymatic activity, did not. c. FBW7 suppressed the increase in glucose uptake caused by PRMT5 in HPDE cells, while the FBW7R465H mutant did not. d. FBW7 inhibited the increase in lactate production induced by PRMT5 in HPDE cells, but the FBW7R465H mutant had little impact. e-f. The ECAR measurement results showed that FBW7 mitigated the increase in glycolysis and glycolytic capacity caused by PRMT5, but the FBW7R465H mutant did not, suggesting that PRMT5 regulates aerobic glycolysis via the FBW7/cMyc axis
Fig. 7
Fig. 7
Schematic representation of the working model In pancreatic cancer, upregulated PRMT5 could epigenetically silence the expression of the E3 ubiquitin ligase FBW7, resulting in cMyc stabilization and a subsequent increase in the proliferation of and glycolysis in pancreatic cancer cells
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