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. 2016 May 27;11(5):e0156274.
doi: 10.1371/journal.pone.0156274. eCollection 2016.

The lncRNA PVT1 Contributes to the Cervical Cancer Phenotype and Associates with Poor Patient Prognosis

Marissa Iden  1 , Samantha Fye  1 , Keguo Li  1 , Tamjid Chowdhury  1 , Ramani Ramchandran  1   2 , Janet S Rader  1
Affiliations

The lncRNA PVT1 Contributes to the Cervical Cancer Phenotype and Associates with Poor Patient Prognosis

Marissa Iden et al. PLoS One. .

Abstract

The plasmacytoma variant translocation 1 gene (PVT1) is an lncRNA that has been designated as an oncogene due to its contribution to the phenotype of multiple cancers. Although the mechanism by which PVT1 influences disease processes has been studied in multiple cancer types, its role in cervical tumorigenesis remains unknown. Thus, the present study was designed to investigate the role of PVT1 in cervical cancer in vitro and in vivo. PVT1 expression was measured by quantitative PCR (qPCR) in 121 invasive cervical carcinoma (ICC) samples, 30 normal cervix samples, and cervical cell lines. Functional assays were carried out using both siRNA and LNA-mediated knockdown to examine PVT1's effects on cervical cancer cell proliferation, migration and invasion, apoptosis, and cisplatin resistance. Our results demonstrate that PVT1 expression is significantly increased in ICC tissue versus normal cervix and that higher expression of PVT1 correlates with poorer overall survival. In cervical cancer cell lines, PVT1 knockdown resulted in significantly decreased cell proliferation, migration and invasion, while apoptosis and cisplatin cytotoxicity were significantly increased in these cells. Finally, we show that PVT1 expression is augmented in response to hypoxia and immune response stimulation and that this lncRNA associates with the multifunctional and stress-responsive protein, Nucleolin. Collectively, our results provide strong evidence for an oncogenic role of PVT1 in cervical cancer and lend insight into potential mechanisms by which PVT1 overexpression helps drive cervical carcinogenesis.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Fig 1
Fig 1. PVT1 and local miRNA expression in cervical cancer tissue.
(A) PVT1 expression was significantly higher in primary cervical tumors (n = 127) compared to adjacent normal cervical tissue (n = 30; p<0.001). Expression of miRNAs 1204 (B; p<0.05) and 1206 (C; p<0.001), but not other local miRNAs, was also significantly higher in cervical tumor tissue (n = 38) compared to adjacent normal tissue (n = 18). (D) Kaplan-Meier plots revealed an association of higher tumor PVT1 levels with significantly poorer survival times compared to lower PVT1 levels (p = 0.03).
Fig 2
Fig 2. PVT1 expression in cervical cancer cells.
(A) PVT1 expression in commercially available cervical cell lines. Lowest PVT1 expression was observed in HPV 16 E6/E7-transformed cells derived from normal ectocervix (E6/E7-Ecto), while SiHa cervical cancer cells displayed the highest PVT1 expression compared to 2 other cervical cancer-derived lines (HeLa and DoTc2). (B) Expression of PVT1 in SiHa cells was further increased upon 48h treatment with INF-α (10 μM) or the hypoxia mimetic cobalt chloride (CoCl2; 150 μM). Representative images of RNA FISH experiments in SiHa cells transfected with either control (C) or PVT1 (D) LNAs. Control LNA-transfected cells exhibited punctate signals for PVT1 (red) in both the nucleus (stained with DAPI in blue) and the cytoplasm. Both nuclear and cytoplasmic PVT1 staining was absent in PVT1 LNA-transfected cells. Phase image (bottom left panel) depicts cell morphology. Scale bar (white) = 10 μm. *p<0.05, **p<0.01.
Fig 3
Fig 3. PVT1 promotes cervical cancer cell proliferation, migration and invasion.
(A) Transfection of SiHa cervical cancer cells with siRNAs targeting PVT1 (siPVT1) resulted in an approximate 70% knockdown in PVT1 lncRNA expression as compared to cells transfected with a scrambled control siRNA (siCONT). (B) SiHa cells transfected with siPVT1 exhibited a significant decrease in proliferation compared to siCONT cells. Transfected SiHa cells were also assessed for changes in (C) migration and (D) invasion 6 h or 48 h following introduction of chemoattractant (FBS), respectively. siPVT1 cells showed a significant decrease in both cell migration and invasion compared to siCONT cells. Quantitative results are graphed on the left, while representative images are on the right. ***p<0.001, ****p<0.0001
Fig 4
Fig 4. PVT1 inhibits cervical cancer cell death and cisplatin sensitivity.
siPVT1 cells exhibited a significant increase in cell death (A) and apoptosis (B) compared to siCONT cells. (C) Proliferation of siPVT1 SiHa cells was significantly decreased in response to multiple doses of cisplatin. (D) Dose-response curve for control and PVT1 LNA-transfected SiHa cells following 4 h treatment with cisplatin. The IC50 value of the PVT1 knockdown cells was significantly decreased compared to control knockdown cells (p<0.0001), suggesting a role for this lncRNA in cisplatin resistance. **p<0.01, ***p<0.001, ****p<0.0001
Fig 5
Fig 5. PVT1 interacts with nucleolin in SiHa cells.
(A) SiHa cells stained for PVT1 (green) and nucleolin (red), and DAPI. Boxed cells (top panel) are at higher magnification (bottom panel). White arrows show PVT1-nucleolin colocalization. Scale bars = 20μM. (B) Western blot for Nucleolin of SiHa protein captured by PVT1 RNA affinity chromatography. (C) PVT1 RNA was immunoprecipitated with anti-Nucleolin, but not the negative control rabbit IgG (Cont). **p<0.01
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References

    1. Prensner JR, Chinnaiyan AM (2011) The emergence of lncRNAs in cancer biology. Cancer Discov 1: 391–407. 10.1158/2159-8290.CD-11-0209 - DOI - PMC - PubMed
    1. TCGA Research Network.
    1. Cory S, Graham M, Webb E, Corcoran L, Adams JM (1985) Variant (6;15) translocations in murine plasmacytomas involve a chromosome 15 locus at least 72 kb from the c-myc oncogene. EMBO J 4: 675–681. - PMC - PubMed
    1. Webb E, Adams JM, Cory S (1984) Variant (6; 15) translocation in a murine plasmacytoma occurs near an immunoglobulin kappa gene but far from the myc oncogene. Nature 312: 777–779. - PubMed
    1. Graham M, Adams JM (1986) Chromosome 8 breakpoint far 3' of the c-myc oncogene in a Burkitt's lymphoma 2;8 variant translocation is equivalent to the murine pvt-1 locus. EMBO J 5: 2845–2851. - PMC - PubMed

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Grants and funding

This work was supported by the Women's Health Research Program at the Medical College of Wisconsin (http://obgyn.mcw.edu/research/whrp/). The funders had no role in any part of preparation of this manuscript.