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Nudt21 regulates the alternative polyadenylation of Pak1 and is predictive in the prognosis of glioblastoma patients

Oncogene volume 38, pages 4154–4168 (2019)Cite this article

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Abstract

Alternative polyadenylation (APA) has emerged as a prevalent feature associated with cancer development and progression. The advantage of APA to tumor progression is to induce oncogenes through 3′-UTR shortening, and to inactivate tumor suppressor genes via the re-routing of microRNA competition. We previously identified the Mammalian Cleavage Factor I-25 (CFIm25) (encoded by Nudt21 gene) as a master APA regulator whose expression levels directly impact the tumorigenicity of glioblastoma (GBM) in vitro and in vivo. Despite its importance, the role of Nudt21 in GBM development is not known and the genes subject to Nudt21 APA regulation that contribute to GBM progression have not been identified. Here, we find that Nudt21 is reduced in low grade glioma (LGG) and all four subtypes of high grade glioma (GBM). Reduced expression of Nudt21 associates with worse survival in TCGA LGG cohorts and two TCGA GBM cohorts. Moreover, although CFIm25 was initially identified as biochemically associated with both CFIm59 and CFIm68, we observed three CFIm distinct subcomplexes exist and CFIm59 protein level is dependent on Nudt21 expression in GBM cells, but CFIm68 is not, and that only CFIm59 predicts prognosis of GBM patients similar to Nudt21. Through the use of Poly(A)-Click-Seq to characterize APA, we define the mRNAs subject to 3′-UTR shortening upon Nudt21 depletion in GBM cells and observed enrichment in genes important in the Ras signaling pathway, including Pak1. Remarkably, we find that Pak1 expression is regulated by Nudt21 through its 3′-UTR APA, and the combination of Pak1 and Nudt21 expression generates an even stronger prognostic indicator of GBM survival versus either value used alone. Collectively, our data uncover Nudt21 and its downstream target Pak1 as a potential “combination biomarker” for predicting prognosis of GBM patients.

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References

  1. Wen PY, Kesari S. Malignant gliomas in adults. N Engl J Med. 2008;359:492–507.

    Article  CAS  Google Scholar 

  2. Chen J, Li Y, Yu TS, McKay RM, Burns DK, Kernie SG, et al. A restricted cell population propagates glioblastoma growth after chemotherapy. Nature. 2012;488:522–6.

    Article  CAS  Google Scholar 

  3. Wang R, Chadalavada K, Wilshire J, Kowalik U, Hovinga KE, Geber A, et al. Glioblastoma stem-like cells give rise to tumour endothelium. Nature. 2010;468:829–33.

    Article  CAS  Google Scholar 

  4. Kang TW, Choi SW, Yang SR, Shin TH, Kim HS, Yu KR, et al. Growth arrest and forced differentiation of human primary glioblastoma multiforme by a novel small molecule. Sci Rep. 2014;4:5546.

    Article  CAS  Google Scholar 

  5. Cancer Genome Atlas Research N. Comprehensive genomic characterization defines human glioblastoma genes and core pathways. Nature. 2008;455:1061–8.

    Article  Google Scholar 

  6. Brennan CW, Verhaak RG, McKenna A, Campos B, Noushmehr H, Salama SR, et al. The somatic genomic landscape of glioblastoma. Cell. 2013;155:462–77.

    Article  CAS  Google Scholar 

  7. Diederichs S, Bartsch L, Berkmann JC, Frose K, Heitmann J, Hoppe C, et al. The dark matter of the cancer genome: aberrations in regulatory elements, untranslated regions, splice sites, non-coding RNA and synonymous mutations. EMBO Mol Med. 2016;8:442–57.

    Article  CAS  Google Scholar 

  8. Tian B, Manley JL. Alternative polyadenylation of mRNA precursors. Nat Rev Mol Cell Biol. 2017;18:18–30.

    Article  CAS  Google Scholar 

  9. Masamha CP, Xia Z, Yang J, Albrecht TR, Li M, Shyu AB, et al. CFIm25 links alternative polyadenylation to glioblastoma tumour suppression. Nature. 2014;510:412–6.

    Article  CAS  Google Scholar 

  10. Singh P, Alley TL, Wright SM, Kamdar S, Schott W, Wilpan RY, et al. Global changes in processing of mRNA 3’ untranslated regions characterize clinically distinct cancer subtypes. Cancer Res. 2009;69:9422–30.

    Article  CAS  Google Scholar 

  11. Gruber AR, Martin G, Muller P, Schmidt A, Gruber AJ, Gumienny R, et al. Global 3’ UTR shortening has a limited effect on protein abundance in proliferating T cells. Nat Commun. 2014;5:5465.

    Article  CAS  Google Scholar 

  12. Elkon R, Drost J, van Haaften G, Jenal M, Schrier M, Oude Vrielink JA, et al. E2F mediates enhanced alternative polyadenylation in proliferation. Genome Biol. 2012;13:R59.

    Article  CAS  Google Scholar 

  13. Mayr C, Bartel DP. Widespread shortening of 3’UTRs by alternative cleavage and polyadenylation activates oncogenes in cancer cells. Cell. 2009;138:673–84.

    Article  CAS  Google Scholar 

  14. Muller S, Rycak L, Afonso-Grunz F, Winter P, Zawada AM, Damrath E, et al. APADB: a database for alternative polyadenylation and microRNA regulation events. Database. 2014;2014. https://doi.org/10.1093/database/bau076.

    Article  Google Scholar 

  15. Sun M, Ding J, Li D, Yang G, Cheng Z, Zhu Q. NUDT21 regulates 3’-UTR length and microRNA-mediated gene silencing in hepatocellular carcinoma. Cancer Lett. 2017;410:158–68.

    Article  CAS  Google Scholar 

  16. Masamha CP, Xia Z, Peart N, Collum S, Li W, Wagner EJ, et al. CFIm25 regulates glutaminase alternative terminal exon definition to modulate miR-23 function. RNA. 2016;22:830–8.

    Article  CAS  Google Scholar 

  17. Akman BH, Can T, Erson-Bensan AE. Estrogen-induced upregulation and 3’-UTR shortening of CDC6. Nucleic Acids Res. 2012;40:10679–88.

    Article  CAS  Google Scholar 

  18. Masamha CP, Wagner EJ. The contribution of alternative polyadenylation to the cancer phenotype. Carcinogenesis. 2018;39:2–10.

    Article  CAS  Google Scholar 

  19. Hardy JG, Norbury CJ. Cleavage factor Im (CFIm) as a regulator of alternative polyadenylation. Biochem Soc Trans. 2016;44:1051–7.

    Article  CAS  Google Scholar 

  20. Ozsolak F, Kapranov P, Foissac S, Kim SW, Fishilevich E, Monaghan AP, et al. Comprehensive polyadenylation site maps in yeast and human reveal pervasive alternative polyadenylation. Cell. 2010;143:1018–29.

    Article  CAS  Google Scholar 

  21. Fu Y, Sun Y, Li Y, Li J, Rao X, Chen C, et al. Differential genome-wide profiling of tandem 3’ UTRs among human breast cancer and normal cells by high-throughput sequencing. Genome Res. 2011;21:741–7.

    Article  CAS  Google Scholar 

  22. Shao J, Zhang J, Zhang Z, Jiang H, Lou X, Huang B, et al. Alternative polyadenylation in glioblastoma multiforme and changes in predicted RNA binding protein profiles. OMICS. 2013;17:136–49.

    Article  CAS  Google Scholar 

  23. Xia Z, Donehower LA, Cooper TA, Neilson JR, Wheeler DA, Wagner EJ, et al. Dynamic analyses of alternative polyadenylation from RNA-seq reveal a 3’-UTR landscape across seven tumour types. Nat Commun. 2014;5:5274.

    Article  CAS  Google Scholar 

  24. Park HJ, Ji P, Kim S, Xia Z, Rodriguez B, Li L, et al. 3’ UTR shortening represses tumor-suppressor genes in trans by disrupting ceRNA crosstalk. Nat Genet. 2018;50:783–89.

    Article  CAS  Google Scholar 

  25. Gruber AR, Martin G, Keller W, Zavolan M. Cleavage factor Im is a key regulator of 3’ UTR length. RNA Biol. 2012;9:1405–12.

    Article  CAS  Google Scholar 

  26. Zhu Y, Wang X, Forouzmand E, Jeong J, Qiao F, Sowd GA, et al. Molecular mechanisms for CFIm-mediated regulation of mRNA alternative polyadenylation. Mol Cell. 2018;69:62–74e4.

    Article  CAS  Google Scholar 

  27. Brown KM, Gilmartin GM. A mechanism for the regulation of pre-mRNA 3’ processing by human cleavage factor Im. Mol Cell. 2003;12:1467–76.

    Article  CAS  Google Scholar 

  28. Ruegsegger U, Beyer K, Keller W. Purification and characterization of human cleavage factor Im involved in the 3’ end processing of messenger RNA precursors. J Biol Chem. 1996;271:6107–13.

    Article  CAS  Google Scholar 

  29. Yang Q, Gilmartin GM, Doublie S. Structural basis of UGUA recognition by the Nudix protein CFI(m)25 and implications for a regulatory role in mRNA 3’ processing. Proc Natl Acad Sci USA. 2010;107:10062–7.

    Article  CAS  Google Scholar 

  30. Yang Q, Coseno M, Gilmartin GM, Doublie S. Crystal structure of a human cleavage factor CFI(m)25/CFI(m)68/RNA complex provides an insight into poly(A) site recognition and RNA looping. Structure. 2011;19:368–77.

    Article  CAS  Google Scholar 

  31. Brumbaugh J, Di Stefano B, Wang X, Borkent M, Forouzmand E, Clowers KJ, et al. Nudt21 controls cell fate by connecting alternative polyadenylation to chromatin signaling. Cell. 2017;172:629–31.

    Article  Google Scholar 

  32. Routh A, Ji P, Jaworski E, Xia Z, Li W, Wagner EJ. Poly(A)-ClickSeq: click-chemistry for next-generation 3-end sequencing without RNA enrichment or fragmentation. Nucleic Acids Res. 2017;45:e112.

    Article  CAS  Google Scholar 

  33. Han T, Kim JK. Driving glioblastoma growth by alternative polyadenylation. Cell Res. 2014;24:1023–4.

    Article  CAS  Google Scholar 

  34. Routh A, Head SR, Ordoukhanian P, Johnson JE. ClickSeq: fragmentation-free next-generation sequencing via click ligation of adaptors to stochastically terminated 3’-azido cDNAs. J Mol Biol. 2015;427:2610–6.

    Article  CAS  Google Scholar 

  35. Kumar R, Gururaj AE, Barnes CJ. p21-activated kinases in cancer. Nat Rev Cancer. 2006;6:459–71.

    Article  CAS  Google Scholar 

  36. Bagheri-Yarmand R, Mandal M, Taludker AH, Wang RA, Vadlamudi RK, Kung HJ, et al. Etk/Bmx tyrosine kinase activates Pak1 and regulates tumorigenicity of breast cancer cells. J Biol Chem. 2001;276:29403–9.

    Article  CAS  Google Scholar 

  37. Holm C, Rayala S, Jirstrom K, Stal O, Kumar R, Landberg G. Association between Pak1 expression and subcellular localization and tamoxifen resistance in breast cancer patients. J Natl Cancer Inst. 2006;98:671–80.

    Article  CAS  Google Scholar 

  38. Tian B, Hu J, Zhang H, Lutz CS. A large-scale analysis of mRNA polyadenylation of human and mouse genes. Nucleic Acids Res. 2005;33:201–12.

    Article  CAS  Google Scholar 

  39. Cornett AL, Lutz CS. RHAPA: a new method to quantify alternative polyadenylation. Methods Mol Biol. 2014;1125:157–67.

    Article  CAS  Google Scholar 

  40. Denysenko T, Gennero L, Roos MA, Melcarne A, Juenemann C, Faccani G, et al. Glioblastoma cancer stem cells: heterogeneity, microenvironment and related therapeutic strategies. Cell Biochem Funct. 2010;28:343–51.

    Article  CAS  Google Scholar 

  41. Kim S, Yamamoto J, Chen Y, Aida M, Wada T, Handa H, et al. Evidence that cleavage factor Im is a heterotetrameric protein complex controlling alternative polyadenylation. Genes Cells. 2010;15:1003–13.

    Article  CAS  Google Scholar 

  42. Sowd GA, Serrao E, Wang H, Wang W, Fadel HJ, Poeschla EM, et al. A critical role for alternative polyadenylation factor CPSF6 in targeting HIV-1 integration to transcriptionally active chromatin. Proc Natl Acad Sci USA. 2016;113:E1054–63.

    Article  CAS  Google Scholar 

  43. Ji Z, Lee JY, Pan Z, Jiang B, Tian B. Progressive lengthening of 3’ untranslated regions of mRNAs by alternative polyadenylation during mouse embryonic development. Proc Natl Acad Sci USA. 2009;106:7028–33.

    Article  CAS  Google Scholar 

  44. Brumbaugh J, Di Stefano B, Wang X, Borkent M, Forouzmand E, Clowers KJ, et al. Nudt21 controls cell fate by connecting alternative polyadenylation to chromatin signaling. Cell. 2018;172:106–20e21.

    Article  CAS  Google Scholar 

  45. Creemers EE, Bawazeer A, Ugalde AP, van Deutekom HW, van der Made I, de Groot NE, et al. Genome-wide polyadenylation maps reveal dynamic mrna 3’-end formation in the failing human heart. Circ Res. 2016;118:433–8.

    Article  CAS  Google Scholar 

  46. Chen MJ, Wu DW, Wang YC, Chen CY, Lee H. PAK1 confers chemoresistance and poor outcome in non-small cell lung cancer via beta-catenin-mediated stemness. Sci Rep. 2016;6:34933.

    Article  CAS  Google Scholar 

  47. Aoki H, Yokoyama T, Fujiwara K, Tari AM, Sawaya R, Suki D, et al. Phosphorylated Pak1 level in the cytoplasm correlates with shorter survival time in patients with glioblastoma. Clin Cancer Res. 2007;13(22 Pt 1):6603–9.

    Article  CAS  Google Scholar 

  48. Wagner EJ, Garcia-Blanco MA. RNAi-mediated PTB depletion leads to enhanced exon definition. Mol Cell. 2002;10:943–9.

    Article  CAS  Google Scholar 

  49. Kim D, Langmead B, Salzberg SL. HISAT: a fast spliced aligner with low memory requirements. Nat Methods. 2015;12:357–60.

    Article  CAS  Google Scholar 

  50. Edlinger L, Berger-Becvar A, Menzl I, Hoermann G, Greiner G, Grundschober E, et al. Expansion of BCR/ABL1+cells requires PAK2 but not PAK1. Br J Haematol. 2017;179:229–41.

    Article  CAS  Google Scholar 

  51. Larson KC, Lipko M, Dabrowski M, Draper MP. Gng12 is a novel negative regulator of LPS-induced inflammation in the microglial cell line BV-2. Inflamm Res. 2010;59:15–22.

    Article  CAS  Google Scholar 

  52. Nagarajan P, Chin SS, Wang D, Liu S, Sinha S, Garrett-Sinha LA. Ets1 blocks terminal differentiation of keratinocytes and induces expression of matrix metalloproteases and innate immune mediators. J Cell Sci. 2010;123(Pt 20):3566–75.

    Article  CAS  Google Scholar 

  53. Johnson DB, Smalley KS, Sosman JA. Molecular pathways: targeting NRAS in melanoma and acute myelogenous leukemia. Clin Cancer Res. 2014;20:4186–92.

    Article  CAS  Google Scholar 

  54. Riemann K, Struwe H, Alakus H, Obermaier B, Schmitz KJ, Schmid KW, et al. Association of GNB4 intron-1 haplotypes with survival in patients with UICC stage III and IV colorectal carcinoma. Anticancer Res. 2009;29:1271–4.

    CAS  PubMed  Google Scholar 

  55. Jasonni VM, Amadori A, Santini D, Ceccarelli C, Naldi S, Flamigni C. Epidermal growth factor receptor (EGF-R) and transforming growth factor alpha (TGFA) expression in different endometrial cancers. Anticancer Res. 1995;15:1327–32.

    CAS  PubMed  Google Scholar 

  56. Zimmermannova O, Doktorova E, Stuchly J, Kanderova V, Kuzilkova D, Strnad H, et al. An activating mutation of GNB1 is associated with resistance to tyrosine kinase inhibitors in ETV6-ABL1-positive leukemia. Oncogene. 2017;36:5985–94.

    Article  CAS  Google Scholar 

  57. Li TF, Qin SH, Ruan XZ, Wang X. p120-catenin participates in the progress of gastric cancer through regulating the Rac1 and Pak1 signaling pathway. Oncol Rep. 2015;34:2357–64.

    Article  CAS  Google Scholar 

  58. Yajima I, Kumasaka MY, Yamanoshita O, Zou C, Li X, Ohgami N, et al. GNG2 inhibits invasion of human malignant melanoma cells with decreased FAK activity. Am J Cancer Res. 2014;4:182–8.

    PubMed  PubMed Central  Google Scholar 

  59. Giubellino A, Burke TR Jr., Bottaro DP. Grb2 signaling in cell motility and cancer. Expert Opin Ther Targets. 2008;12:1021–33.

    Article  CAS  Google Scholar 

  60. Walker-Daniels J, Hess AR, Hendrix MJ, Kinch MS. Differential regulation of EphA2 in normal and malignant cells. Am J Pathol. 2003;162:1037–42.

    Article  CAS  Google Scholar 

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Acknowledgements

This work was supported by the US National Institutes of Health (NIH) grants R01CA193466-01 to W.L. and E.J.W., the Cancer Prevention Research Institute of Texas grant CPRIT RP140800 to E.J.W., and NIH R03 CA223893-01 to P.J.

Author contributions

E.J.W and P.J. conceived the project, designed the experiments and Y.C. and N.E. performed the experiments and data analysis. C.W. and Z.X. performed survival analysis. N.E. and A.R. performed the PAC-Seq APA analysis. L.L. and W.L. performed TCGA GBM RNA-seq data analysis, T.C. helped with data analysis. Y.C., P.J., and E.J.W. wrote the manuscript with input from N.E., L.L., T.C., and A.R.

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, The University of Texas Medical Branch, Galveston, TX, USA

    Yuan Chu, Nathan Elrod, Andrew Routh, Eric J. Wagner & Ping Ji

  2. Endoscopy Center, Zhongshan Hospital and Endoscopy Research Institute, Fudan University, Shanghai, China

    Yuan Chu & Tao Chen

  3. Department of Molecular Microbiology and Immunology, Computational Biology Program, OHSU, Portland, OR, 97273, USA

    Chaojie Wang & Zheng Xia

  4. Daniel Duncan Cancer Center, Baylor College of Medicine, Houston, TX, USA

    Lei Li & Wei Li

  5. Sealy Centre for Structural Biology and Molecular Biophysics, University of Texas Medical Branch, Galveston, TX, USA

    Andrew Routh & Eric J. Wagner

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Chu, Y., Elrod, N., Wang, C. et al. Nudt21 regulates the alternative polyadenylation of Pak1 and is predictive in the prognosis of glioblastoma patients. Oncogene 38, 4154–4168 (2019). https://doi.org/10.1038/s41388-019-0714-9

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