p53 performs an essential role in mediating the oncogenic stimulus triggered by loss of expression of neurofibromatosis type 2 during in vitro tumor progression

Li,Xiye; Chen,Hongsai; Xue,Lu; Pang,Xiuhong; Zhang,Xiaoman; Zhu,Zhengjie; Zhu,Weidong; Wang,Zhaoyan; Wu,Hao

doi:10.3892/ol.2017.6445

p53 performs an essential role in mediating the oncogenic stimulus triggered by loss of expression of neurofibromatosis type 2 during in vitro tumor progression

Authors:
- Xiye Li
- Hongsai Chen
- Lu Xue
- Xiuhong Pang
- Xiaoman Zhang
- Zhengjie Zhu
- Weidong Zhu
- Zhaoyan Wang
- Hao Wu
View Affiliations

Affiliations: Department of Otolaryngology Head and Neck Surgery, Xinhua Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200092, P.R. China
Published online on: June 21, 2017 https://doi.org/10.3892/ol.2017.6445
Pages: 2223-2231
Copyright: © Li et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

The loss of the tumor suppressor neurofibromatosis type 2 gene, encoding merlin, has been considered to be a fundamental event during the malignant progression of various cell types. However, a consensus for the mainstream mechanism, by which merlin deficiency contributes to uncontrolled cellular proliferation, has not been reached. The present study aimed to determine whether silencing of merlin using lentivirus‑based short hairpin RNA potentiates cellular proliferation and cell cycle progression in human colon carcinoma HCT116 cell lines, expressing p53. The present results demonstrated that merlin knockdown contributed to cellular proliferation and G1/S cell cycle progression to a greater extent in HCT116 cells wide‑type for p53 (p53wt) compared with p53‑null (p53‑/‑) cells. This was supported by overexpression experiments which demonstrated a significant inhibitory effect of excess merlin on cellular proliferation only in HCT116 p53wt cells. In order to investigate the underlying mechanisms of action, the expression of p53‑involved G1/S transition genes was evaluated by western blot analysis. For HCT116 p53wt cells, merlin loss suppressed p53 expression, and therefore the dysregulation of cell cycle regulatory proteins, including p21, cyclin D1/cyclin‑dependent kinase (CDK)4 and cyclin E1/CDK2 complexes. However, merlin knockdowns had no impact on the expression of any of the aforementioned molecules in p53‑/‑ cells, indicating that lack of merlin resulted in G1/S cell cycle progression, and thereby uncontrolled cellular proliferation mainly via the regulation of p53‑mediated pathways. Taken together, it was proposed that p53 performs an essential role in mediating the oncogenic stimulus triggered by merlin loss, and p53 is a molecule that should be investigated for its potential in targeted drug therapy for merlin‑deficient malignancies.

View Figures

View References

1	Wang Z, Lu Y, Tang J, Wang H and Wu H: The phosphorylation status of merlin in sporadic vestibular Schwannomas. Mol Cell Biochem. 324:201–206. 2009. View Article : Google Scholar : PubMed/NCBI
2	Zhang Z, Wang Z, Sun L, Li X, Huang Q, Yang T and Wu H: Mutation spectrum and differential gene expression in cystic and solid vestibular schwannoma. Genet Med. 16:264–270. 2014. View Article : Google Scholar : PubMed/NCBI
3	Chen H, Zhang X, Zhang Z, Yang T, Wang Z and Wu H: The role of NF2 gene mutations and pathogenesis-related proteins in sporadic vestibular schwannomas in young individuals. Mol Cell Biochem. 392:145–152. 2014. View Article : Google Scholar : PubMed/NCBI
4	Lau YK, Murray LB, Houshmandi SS, Xu Y, Gutmann DH and Yu Q: Merlin is a potent inhibitor of glioma growth. Cancer Res. 68:5733–5742. 2008. View Article : Google Scholar : PubMed/NCBI
5	Sheikh HA, Tometsko M, Niehouse L, Aldeeb D, Swalsky P, Finkelstein S, Barnes EL and Hunt JL: Molecular genotyping of medullary thyroid carcinoma can predict tumor recurrence. Am J Surg Pathol. 28:101–106. 2004. View Article : Google Scholar : PubMed/NCBI
6	Sekido Y: Genomic abnormalities and signal transduction dysregulation in malignant mesothelioma cells. Cancer Science. 101:1–6. 2010. View Article : Google Scholar : PubMed/NCBI
7	Cačev T, Aralica G, Lončar B and Kapitanović S: Loss of NF2/Merlin expression in advanced sporadic colorectal cancer. Cell Oncol (Dordr). 37:69–77. 2014. View Article : Google Scholar : PubMed/NCBI
8	Shimizu K, Nagamachi Y, Tani M, Kimura K, Shiroishi T, Wakana S and Yokota J: Molecular cloning of a novel NF2/ERM/4.1 superfamily gene, ehm2, that is expressed in high-metastatic K1735 murine melanoma cells. Genomics. 65:113–120. 2000. View Article : Google Scholar : PubMed/NCBI
9	Sainz J, Huynh DP, Figueroa K, Ragge NK, Baser ME and Pulst SM: Mutations of the neurofibromatosis type 2 gene and lack of the gene product in vestibular schwannomas. Hum Mol Genet. 3:885–891. 1994. View Article : Google Scholar : PubMed/NCBI
10	Stamenkovic I and Yu Q: Merlin, a ‘magic’ linker between the extracellular cues and intracelular signaling pathways that regulate cell motility, proliferation, and survival. Curr Protein Pept Sci. 11:471–484. 2010. View Article : Google Scholar : PubMed/NCBI
11	Schroeder RD, Angelo LS and Kurzrock R: NF2/merlin in hereditary neurofibromatosis 2 versus cancer: Biologic mechanisms and clinical associations. Oncotarget. 5:67–77. 2014.PubMed/NCBI
12	Gutmann DH, Giordano MJ, Fishback AS and Guha A: Loss of merlin expression in sporadic meningiomas, ependymomas and schwannomas. Neurology. 49:267–270. 1997. View Article : Google Scholar : PubMed/NCBI
13	Yang C, Asthagiri AR, Iyer RR, Lu J, Xu DS, Ksendzovsky A, Brady RO, Zhuang Z and Lonser RR: Missense mutations in the NF2 gene result in the quantitative loss of merlin protein and minimally affect protein intrinsic function. Proc Natl Acad Sci USA. 108:pp. 4980–4985. 2011; View Article : Google Scholar : PubMed/NCBI
14	Ammoun S, Flaiz C, Ristic N, Schuldt J and Hanemann CO: Dissecting and targeting the growth factor-dependent and growth factor-independent extracellular signal-regulated kinase pathway in human schwannoma. Cancer Res. 68:5236–5245. 2008. View Article : Google Scholar : PubMed/NCBI
15	Curto M, Cole BK, Lallemand D, Liu CH and McClatchey AI: Contact-dependent inhibition of EGFR signaling by Nf2/Merlin. J Cell Biol. 177:893–903. 2007. View Article : Google Scholar : PubMed/NCBI
16	Flaiz C, Ammoun S, Biebl A and Hanemann CO: Altered adhesive structures and their relation to RhoGTPase activation in merlin deficient Schwannoma. Brain Pathol. 19:27–38. 2009. View Article : Google Scholar : PubMed/NCBI
17	Li W, You L, Cooper J, Schiavon G, Pepe-Caprio A, Zhou L, Ishii R, Giovannini M, Hanemann CO, Long SB, et al: Merlin/NF2 suppresses tumorigenesis by inhibiting the E3 ubiquitin ligase CRL4(DCAF1) in the nucleus. Cell. 140:477–490. 2010. View Article : Google Scholar : PubMed/NCBI
18	Robanus-Maandag E, Giovannini M, van der Valk M, Niwa-Kawakita M, Abramowski V, Antonescu C, Thomas G and Berns A: Synergy of Nf2 and p53 mutations in development of malignant tumours of neural crest origin. Oncogene. 23:6541–6547. 2004. View Article : Google Scholar : PubMed/NCBI
19	Chang Z, Guo CL, Ahronowitz I, Stemmer-Rachamimov AO, MacCollin M and Nunes FP: A role for the p53 pathway in the pathology of meningiomas with NF2 loss. J Neurooncol. 91:265–270. 2009. View Article : Google Scholar : PubMed/NCBI
20	Kim H, Kwak NJ, Lee JY, Choi BH, Lim Y, Ko YJ, Kim YH, Huh PW, Lee KH, Rha HK and Wang YP: Merlin neutralizes the inhibitory effect of Mdm2 on p53. J Biol Chem. 279:7812–7818. 2004. View Article : Google Scholar : PubMed/NCBI
21	Wu H, Chen Y, Wang ZY, Li W, Li JQ, Zhang L and Lu YJ: Involvement of p21 (waf1) in merlin deficient sporadic vestibular schwannomas. Neuroscience. 170:149–155. 2010. View Article : Google Scholar : PubMed/NCBI
22	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) Method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
23	Poulikakos PI, Xiao GH, Gallagher R, Jablonski S, Jhanwar SC and Testa JR: Re-expression of the tumor suppressor NF2/merlin inhibits invasiveness in mesothelioma cells and negatively regulates FAK. Oncogene. 25:5960–5968. 2006. View Article : Google Scholar : PubMed/NCBI
24	Schulze KM, Hanemann CO, Müller HW and Hanenberg H: Transduction of wild-type merlin into human schwannoma cells decreases schwannoma cell growth and induces apoptosis. Hum Mol Genet. 11:69–76. 2002. View Article : Google Scholar : PubMed/NCBI
25	Lü J, Zou J, Wu H and Cai L: Compensative shuttling of merlin to phosphorylation on serine 518 in vestibular schwannoma. Laryngoscope. 118:169–174. 2008. View Article : Google Scholar : PubMed/NCBI
26	Muranen T, Grönholm M, Renkema GH and Carpén O: Cell cycle-dependent nucleocytoplasmic shuttling of the neurofibromatosis 2 tumour suppressor merlin. Oncogene. 24:1150–1158. 2005. View Article : Google Scholar : PubMed/NCBI
27	Arellano M and Moreno S: Regulation of CDK/cyclin complexes during the cell cycle. Int J Biochem Cell Biol. 29:559–573. 1997. View Article : Google Scholar : PubMed/NCBI
28	Scoles DR: The merlin interacting proteins reveal multiple targets for NF2 therapy. Biochim Biophys Acta. 1785:32–54. 2008.PubMed/NCBI
29	Lallemand D, Manent J, Couvelard A, Watilliaux A, Siena M, Chareyre F, Lampin A, Niwa-Kawakita M, Kalamarides M and Giovannini M: Merlin regulates transmembrane receptor accumulation and signaling at the plasma membrane in primary mouse Schwann cells and in human schwannomas. Oncogene. 28:854–865. 2009. View Article : Google Scholar : PubMed/NCBI
30	Planque N: Nuclear trafficking of secreted factors and cell-surface receptors: New pathways to regulate cell proliferation and differentiation and involvement in cancers. Cell Commun Signal. 4:72006. View Article : Google Scholar : PubMed/NCBI
31	Ahmad Z, Brown CM, Patel AK, Ryan AF, Ongkeko R and Doherty JK: Merlin knockdown in human Schwann cells: Clues to vestibular schwannoma tumorigenesis. Otol Neurotol. 31:460–466. 2010. View Article : Google Scholar : PubMed/NCBI
32	Monoh K, Ishikawa K, Yasui N, Mineura K, Andoh H and Togawa K: p53 tumor suppressor gene in acoustic neuromas. Acta Otolaryngol Suppl. 537:11–15. 1998.PubMed/NCBI