Gene expression profiling of NB4 cells following knockdown of nucleostemin using DNA microarrays

Sun,Xiaoli; Jia,Yu; Wei,Yuanyu; Liu,Shuai; Yue,Baohong

doi:10.3892/mmr.2016.5213

Gene expression profiling of NB4 cells following knockdown of nucleostemin using DNA microarrays

Authors:
- Xiaoli Sun
- Yu Jia
- Yuanyu Wei
- Shuai Liu
- Baohong Yue
View Affiliations

Affiliations: Department of Laboratory, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan 450052, P.R. China
Published online on: May 5, 2016 https://doi.org/10.3892/mmr.2016.5213
Pages: 175-183
Copyright: © Sun 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

Nucleostemin (NS) is mainly expressed in stem and tumor cells, and is necessary for the maintenance of their self-renewal and proliferation. Originally, NS was thought to exert its effects through inhibiting p53, while recent studies have revealed that NS is also able to function independently of p53. The present study performed a gene expression profiling analysis of p53‑mutant NB4 leukeima cells following knockdown of NS in order to elucidate the p53‑independent NS pathway. NS expression was silenced using lentivirus‑mediated RNA interference technology, and gene expression profiling of NB4 cells was performed by DNA microarray analysis. A total of 1,953 genes were identified to be differentially expressed (fold change ≥2 or ≤0.5) following knockdown of NS expression. Furthermore, reverse‑transcription quantitative polymerase chain reaction analysis was used to detect the expression of certain candidate genes, and the results were in agreement with the micaroarray data. Pathway analysis indicated that aberrant genes were enhanced in endoplasmic, c‑Jun N‑terminal kinase and mineral absorption pathways. The present study shed light on the mechanisms of the p54‑independent NS pathway in NB4 cells and provided a foundation for the discovery of promising targets for the treatment of p53-mutant leukemia.

View Figures

View References

1	Tsai RY and McKay RD: A nucleolar mechanism controlling cell proliferation in stem cells and cancer cells. Genes Dev. 16:2991–3003. 2002. View Article : Google Scholar : PubMed/NCBI
2	Liu SJ, Cai ZW, Liu YJ, Dong MY, Sun LQ, Hu GF, Wei YY and Lao WD: Role of nucleostemin in growth regulation of gastric cancer, liver cancer and other malignancies. World J Gastroenterol. 10:1246–1249. 2004.PubMed/NCBI
3	Uema N, Ooshio T, Harada K, Naito M, Naka K, Hoshii T, Tadokoro Y, Ohta K, Ali MA, Katano M, et al: Abundant nucleostemin expression supports the undifferentiated properties of germ cell tumors. Am J Pathol. 183:592–603. 2013. View Article : Google Scholar : PubMed/NCBI
4	Liu RL, Xu Y, Zhang ZH, Wang M, Sun JT, Qi SY, Zhang Y and Li SZ: Expression of nucleostemin in prostate cancer tissues and its clinical significance. Natl J Androl. 14:418–422. 2008.In Chinese.
5	Zhang GY, Yin L, Li SL, Xing WY, Zhao QM, Le XP, Gao DL, Chen KS, Zhang YH and Zhang QX: Expression of nucleostemin mRNA and protein in the esophageal squamous cell carcinoma. Chin J Oncol. 30:125–128. 2008.In Chinese.
6	Guo Y, Liao YP, Zhang D, Xu LS, Li N, Guan WJ and Liu CQ: In vitro study of nucleostemin as a potential therapeutic target in human breast carcinoma SKBR-3 cells. Asian Pac J Cancer Prev. 15:2291–2295. 2014. View Article : Google Scholar : PubMed/NCBI
7	Asadi MH, Derakhshani A and Mowla SJ: Concomitant upregulation of nucleostemin and downregulation of Sox2 and Klf4 in gastric adenocarcinoma. Tumour Biol. 35:7177–7185. 2014. View Article : Google Scholar : PubMed/NCBI
8	Kobayashi T, Masutomi K, Tamura K, Moriya T, Yamasaki T, Fujiwara Y, Takahashi S, Yamamoto J and Tsuda H: Nucleostemin expression in invasive breast cancer. BMC Cancer. 14:2152014. View Article : Google Scholar : PubMed/NCBI
9	You Y, Li X, Zheng J, Wu Y, He Y, Du W, Zou P and Zhang M: Transcript level of nucleostemin in newly diagnosed acute myeloid leukemia patients. Leuk Res. 37:1636–1641. 2013. View Article : Google Scholar : PubMed/NCBI
10	Sijin L, Ziwei C, Yajun L, Meiyu D, Hongwei Z, Guofa H, Siguo L, Hong G, Zhihong Z, Xiaolei L, et al: The effect of knocking-down nucleostemin gene expression on the in vitro proliferation and in vivo tumorigenesis of HeLa cells. J Exp Clin Cancer Res. 23:529–538. 2004.PubMed/NCBI
11	Ma H and Pederson T: Depletion of the nucleolar protein nucleostemin causes G1 cell cycle arrest via the p53 pathway. Mol Biol Cell. 18:2630–2635. 2007. View Article : Google Scholar : PubMed/NCBI
12	Liu RL, Zhang ZH, Zhao WM, Wang M, Qi SY, Li J, Zhang Y, Li SZ and Xu Y: Expression of nucleostemin in prostate cancer and its effect on the proliferation of PC-3 cells. Chin Med J (Engl). 121:299–304. 2008.
13	Seyed-Gogani N, Rahmati M, Zarghami N, Asvadi-Kermani I, Hoseinpour-Feyzi MA and Moosavi MA: Nucleostemin depletion induces post-g1 arrest apoptosis in chronic myelogenous leukemia k562 cells. Adv Pharm Bull. 4:55–60. 2014.PubMed/NCBI
14	Amini S, Fathi F, Mobalegi J, Sofimajidpour H and Ghadimi T: The expressions of stem cell markers: Oct4, Nanog, Sox2, nucleostemin, Bmi, Zfx, Tcl1, Tbx3, Dppa4 and Esrrb in bladder, colon and prostate cancer and certain cancer cell lines. Anat Cell Biol. 47:1–11. 2014. View Article : Google Scholar : PubMed/NCBI
15	Beekman C, Nichane M, De Clercq S, Maetens M, Floss T, Wurst W, Bellefroid E and Marine JC: Evolutionarily conserved role of nucleostemin: Controlling proliferation of stem/progenitor cells during early vertebrate development. Mol Cell Biol. 26:9291–9301. 2006. View Article : Google Scholar : PubMed/NCBI
16	Jafarnejad SM, Mowla SJ and Matin MM: Knocking-down the expression of nucleostemin significantly decreases rate of proliferation of rat bone marrow stromal stem cells in an apparently p53-independent manner. Cell Prolif. 41:28–35. 2008. View Article : Google Scholar : PubMed/NCBI
17	Nikpour P, Mowla SJ, Jafarnejad SM, Fischer U and Schulz WA: Differential effects of Nucleostemin suppression on cell cycle arrest and apoptosis in the bladder cancer cell lines 5637 and SW1710. Cell Prolif. 42:762–769. 2009. View Article : Google Scholar : PubMed/NCBI
18	Lo D, Zhang Y, Dai MS, Sun XX, Zeng SX and Lu H: Nucleostemin stabilizes ARF by inhibiting the ubiquitin ligase ULF. Oncogene. 34:1688–1697. 2015. View Article : Google Scholar
19	Li X, Zhou J, Chen ZR and Chng WJ: P53 mutations in colorectal cancer-molecular pathogenesis and pharmacological reactivation. World J Gastroenterol. 21:84–93. 2015. View Article : Google Scholar : PubMed/NCBI
20	Sun X, Jia Y, Wei Y, Liu S and Yue B: Gene expression profiling of HL-60 cells following knockdown of nucleostemin using DNA microarrays. Oncol Rep. 32:739–747. 2014.PubMed/NCBI
21	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
22	Liu R, Zhang Z and Xu Y: Downregulation of nucleostemin causes G1 cell cycle arrest via a p53-independent pathway in prostate cancer PC-3 cells. Urol Int. 85:221–227. 2010. View Article : Google Scholar : PubMed/NCBI
23	Zwolinska AK, Heagle Whiting A, Beekman C, Sedivy JM and Marine JC: Suppression of Myc oncogenic activity by nucleostemin haploinsufficiency. Oncogene. 31:3311–3321. 2012. View Article : Google Scholar :
24	Paridaen JT, Janson E, Utami KH, Pereboom TC, Essers PB, Rooijen C, Zivkovic D and MacInnes AW: The nucleolar GTP-binding proteins Gnl2 and nucleostemin are required for retinal neurogenesis in developing zebrafish. Dev Biol. 355:286–301. 2011. View Article : Google Scholar : PubMed/NCBI
25	Stolz A and Wolf DH: Endoplasmic reticulum associated protein degradation: A chaperone assisted journey to hell. Biochim Biophys Acta. 1803:694–705. 2010. View Article : Google Scholar : PubMed/NCBI
26	Määttänen P, Gehring K, Bergeron JJ and Thomas DY: Protein quality control in the ER: The recognition of misfolded proteins. Semin Cell Dev Biol. 21:500–411. 2010. View Article : Google Scholar : PubMed/NCBI
27	Fu XL: The regulative mechanism of organisms against stress injuries. Biofactors. 40:569–585. 2014. View Article : Google Scholar : PubMed/NCBI
28	Lu W, Hagiwara D, Morishita Y, Tochiya M, Azuma Y, Suga H, Goto M, Banno R, Sugimura Y, Oyadomari S, et al: Unfolded protein response in hupothalamic cultures of wild-type and ATF6α-knockout mice. Neurosci Lett. 612:199–203. 2015. View Article : Google Scholar
29	Rao RV, Castro-Obregon S, Frankowski H, Schuler M, Stoka V, Del RG, Bredesen DE and Ellerby HM: Coupling endoplasmic reticulum stress to the cell death program. An Apaf-1-independent intrinsic pathway. J Biol Chem. 277:21836–21842. 2002. View Article : Google Scholar : PubMed/NCBI
30	Szegezdi E, Logue SE, Gorman AM and Samali A: Mediators of endoplasmic reticulum stress-induced apoptosis. EMBO Rep. 7:880–885. 2006. View Article : Google Scholar : PubMed/NCBI
31	Raman M, Chen W and Cobb MH: Differential regulation and properties of MAPKs. Oncogene. 26:3100–3112. 2007. View Article : Google Scholar : PubMed/NCBI
32	Huang ZX: Neuronal growth-inhibitory factor (metallo-thionein-3): A unique metalloprotein. FEBS J. 277:29112010. View Article : Google Scholar
33	Vašák M and Meloni G: Chemistry and biology of mammalian metallothioneins. J Biol Inorg Chem. 16:1067–1078. 2011. View Article : Google Scholar
34	Eide DJ: The oxidative stress of zinc deficiency. Metallomics. 3:1124–1129. 2011. View Article : Google Scholar : PubMed/NCBI
35	Miura T, Muraoka S and Ogiso T: Antioxidant activity of metallothionein compared with reduced glutathione. Life Sci. 60:301–309. 1997. View Article : Google Scholar
36	Ghoshal K and Jacob ST: Regulation of metallothionein gene expression. Prog Nucleic Acid Res Mol Biol. 66:357–384. 2001. View Article : Google Scholar
37	Günes C, Heuchel R, Georgiev O, Müller KH, Lichtlen P, Bluthmann H, Marino S, Aguzzi A and Schaffner W: Embryonic lethality and liver degeneration in mice lacking the metal-responsive transcriptional activator MTF-1. EMBO J. 17:2846–2854. 1998. View Article : Google Scholar : PubMed/NCBI
38	Klassen RB, Crenshaw K, Kozyraki R, Verroust PJ, Tio L, Atrian S, Allen PL and Hammond TG: Megalin mediates renal uptake of heavy metal metallothionein complexes. Am J Physiol Renal Physiol. 287:F393–F403. 2004. View Article : Google Scholar : PubMed/NCBI
39	Shi Y, Amin K, Sato BG, Samuelsson SJ, Sambucetti L, Haroon ZA, Laderoute K and Murphy BJ: The metal-responsive transcription factor-1 protein is elevated in human tumors. Cancer Biol Ther. 9:469–476. 2010. View Article : Google Scholar : PubMed/NCBI
40	Cherian MG and Apostolova MD: Nuclear localization of metallothionein during cell proliferation and differentiation. Cell Mol Biol (Noisy-le-grand). 46:347–356. 2000.
41	Ogra Y and Suzuki KT: Nuclear trafficking of metallothionein: Possible mechanisms and current knowledge. Cell Mol Biol (Noisy-le-grand). 46:357–365. 2000.
42	Knipp M: Metallothioneins and platinum (II) anti-tumor compounds. Curr Med Chem. 16:522–537. 2009. View Article : Google Scholar
43	Boulikas T and Vougiouka M: Cisplatin and platinum drugs at the molecular level (Review). Oncol Rep. 10:1663–1682. 2003.PubMed/NCBI
44	McGee HM, Woods GM, Bennett B and Chung RS: The two faces of metallothionein in carcinogenesis: Photoprotection against UVR-induced cancer and promotion of tumour survival. Photochem Photobiol Sci. 9:586–596. 2010. View Article : Google Scholar : PubMed/NCBI
45	Dutsch-Wicherek M, Sikora J and Tomaszewska R: The possible biological role of metallothionein in apoptosis. Front Biosci. 13:4029–4038. 2008. View Article : Google Scholar : PubMed/NCBI
46	Gomulkiewicz A, Podhorska-Okolow M, Szulc R, Smorag Z, Wojnar A, Zabel M and Dziegiel P: Correlation between metallothionein (MT) expression and selected prognostic factors in ductal breast cancers. Folia Histochem Cytobiol. 48:242–248. 2010. View Article : Google Scholar : PubMed/NCBI
47	Królicka A, Kobierzycki C, Puła B, Podhorska-Okołów M, Piotrowska A, Rzeszutko M, Rzeszutko W, Rabczyński J, Domosławski P, Wojtczak B, et al: Comparison of metallo-thionein (MT) and Ki-67 antigen expression in benign and malignant thyroid tumours. Anticancer Res. 30:4945–4949. 2010.
48	Kishton RJ and Rathmell JC: Novel therapeutic targets of tumor metabolism. Cancer J. 21:62–69. 2015. View Article : Google Scholar : PubMed/NCBI
49	Gottfried E, Kreutz M and Mackensen A: Tumor metabolism as modulator of immune response and tumor progression. Semin Cancer Biol. 22:335–341. 2012. View Article : Google Scholar : PubMed/NCBI