CRKII overexpression promotes the in vitro proliferation, migration and invasion potential of murine hepatocarcinoma Hca‑P cells

  • Authors:
    • Zanmei Zhou
    • Xiuyan Sun
    • Chunmei Guo
    • Ming‑Zhong Sun
    • Shuqing Liu
  • View Affiliations

  • Published online on: March 27, 2019     https://doi.org/10.3892/ol.2019.10194
  • Pages: 5169-5174
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Abstract

Lymphatic metastasis is a major mechanism of tumor metastasis. The present study aimed to investigate the association of CRKII, a member of the CRK family, with the in vitro malignant behaviors of a murine hepatocarcinoma Hca‑P cell line, with a lymph node metastatic rate of ~25%. Total mRNA was extracted from Hca‑P cells, and then the murine CRKII gene was amplified by polymerase chain reaction and cloned into the pEASY‑blunt cloning vector. Subsequently, the recombinant pcDNA3.1/V5‑HisB‑CRKII plasmid was constructed and transfected into Hca‑P cells. Western blotting indicated that the CRKII expression level in pcDNA3.1/V5‑HisB‑CRKII‑Hca‑P cells was increased by ~185%, compared with pcDNA3.1/V5‑HisB‑Hca‑P cells. The stable overexpression of CRKII enhanced the in vitro proliferation ability, as measured with a Cell Counting Kit‑8 assay, and the colony forming capacity was measured with a soft agar colony forming assay for Hca‑P cells. The in vitro migration and invasion capacities of Hca‑P cells were increased by ~179 and 156% in Hca‑P cells, respectively, following the stable upregulation of CRKII. Collectively, the recombinant pcDNA3.1/V5‑HisB‑CRKII‑Hca‑P plasmid was constructed successfully. Additionally, the CRKII expression level was positively associated with the in vitro proliferation, migration and invasion malignant properties of Hca‑P cells.

References

1 

Feller SM: Crk family adaptors-signalling complex formation and biological roles. Oncogene. 20:6348–6371. 2001. View Article : Google Scholar : PubMed/NCBI

2 

Birge RB, Kalodimos C, Inagaki F and Tanaka S: Crk and CrkL adaptor proteins: Networks for physiological and pathological signaling. Cell Commun Signal. 7:132009. View Article : Google Scholar : PubMed/NCBI

3 

Bos JL, Rehmann H and Wittinghofer A: GEFs and GAPs: Critical elements in the control of small G proteins. Cell. 129:865–877. 2007. View Article : Google Scholar : PubMed/NCBI

4 

Hasegawa H, Kiyokawa E, Tanaka S, Nagashima K, Gotoh N, Shibuya M, Kurata T and Matsuda M: DOCK180, a major CRK-binding protein, alters cell morphology upon translocation to the cell membrane. Mol Cell Biol. 16:1770–1776. 1996. View Article : Google Scholar : PubMed/NCBI

5 

Tanaka S, Morishita T, Hashimoto Y, Hattori S, Nakamura S, Shibuya M, Matuoka K, Takenawa T, Kurata T, Nagashima K, et al: C3G, a guanine nucleotide-releasing protein expressed ubiquitously, binds to the Src homology 3 domains of CRK and GRB2/ASH proteins. Proc Natl Acad Sci USA. 91:3443–3447. 1994. View Article : Google Scholar : PubMed/NCBI

6 

Miller CT, Chen G, Gharib TG, Wang H, Thomas DG, Misek DE, Giordano TJ, Yee J, Orringer MB, Hanash SM and Beer DG: Increased C-CRK proto-oncogene expression is associated with an aggressive phenotype in lung adenocarcinomas. Oncogene. 22:7950–7957. 2003. View Article : Google Scholar : PubMed/NCBI

7 

Wang L, Tabu K, Kimura T, Tsuda M, Linghu H, Tanino M, Kaneko S, Nishihara H and Tanaka S: Signaling adaptor protein Crk is indispensable for malignant feature of glioblastoma cell line KMG4. Biochem Biophys Res Commun. 362:976–981. 2007. View Article : Google Scholar : PubMed/NCBI

8 

Sriram G and Birge RB: Emerging roles for crk in human cancer. Genes Cancer. 1:1132–1139. 2010. View Article : Google Scholar : PubMed/NCBI

9 

Fathers KE, Bell ES, Rajadurai CV, Cory S, Zhao H, Mourskaia A, Zuo D, Madore J, Monast A, Mes-Masson AM, et al: Crk adaptor proteins act as key signaling integrators for breast tumorigenesis. Breast Cancer Res. 14:R742012. View Article : Google Scholar : PubMed/NCBI

10 

Rodrigues SP, Fathers KE, Chan G, Zuo D, Halwani F, Meterissian S and Park M: CrkI and CrkII function as key signaling integrators for migration and invasion of cancer cells. Mol Cancer Res. 3:183–194. 2005.PubMed/NCBI

11 

Bell ES and Park M: Models of crk adaptor proteins in cancer. Genes Cancer. 3:341–352. 2012. View Article : Google Scholar : PubMed/NCBI

12 

Guo Y, Li S, Qu J, Wang S, Dang Y, Fan J, Yu S and Zhang J: MiR-34a inhibits lymphatic metastasis potential of mouse hepatoma cells. Mol Cell Biochem. 354:275–282. 2011. View Article : Google Scholar : PubMed/NCBI

13 

Huang Y, Wang Q, Du Y, Bai L, Jin F, Zhang J, Fan S, Wang H, Song L, Gao Y, et al: Inhibition of annexin A7 gene and protein induces the apotosis and decreases the invasion, migration of the hepatocarcinoma cell line. Biomed Pharmacother. 68:819–824. 2014. View Article : Google Scholar : PubMed/NCBI

14 

Zhang YH, Wang SQ, Sun CR, Wang M, Wang B and Tang JW: Inhibition of JNK1 expression decreases migration and invasion of mouse hepatocellular carcinoma cell line in vitro. Med Oncol. 28:966–972. 2011. View Article : Google Scholar : PubMed/NCBI

15 

Wu J, Meng J, Du Y, Huang Y, Jin Y, Zhang J, Wang B, Zhang Y, Sun M and Tang J: RACK1 promotes the proliferation, migration and invasion capacity of mouse hepatocellular carcinoma cell line in vitro probably by PI3K/Rac1 signaling pathway. Biomed Pharmacother. 67:313–319. 2013. View Article : Google Scholar : PubMed/NCBI

16 

Sleeman JP: The lymph node as a bridgehead in the metastatic dissemination of tumors. Recent Results Cancer Res. 157:55–81. 2000. View Article : Google Scholar : PubMed/NCBI

17 

Kawada K and Taketo MM: Significance and mechanism of lymph node metastasis in cancer progression. Cancer Res. 71:1214–1218. 2011. View Article : Google Scholar : PubMed/NCBI

18 

Kabilova TO, Kovtonyuk LV, Zonov EV, Ryabchikova EI, Popova NA, Nikolin VP, Kaledin VI, Zenkova MA, Vlassov VV and Chernolovskaya EL: Immunotherapy of hepatocellular carcinoma with small double-stranded RNA. BMC Cancer. 14:3382014. View Article : Google Scholar : PubMed/NCBI

19 

Abdel-Rahman O: Revisiting oxaliplatin-based regimens for advanced hepatocellular carcinoma. Curr Oncol Rep. 16:3942014. View Article : Google Scholar : PubMed/NCBI

20 

Quetglas IM, Moeini A, Pinyol R and Llovet JM: Integration of genomic information in the clinical management of HCC. Best Pract Res Clin Gastroenterol. 28:831–842. 2014. View Article : Google Scholar : PubMed/NCBI

21 

Chuang SC, La Vecchia C and Boffetta P: Liver cancer: Descriptive epidemiology and risk factors other than HBV and HCV infection. Cancer Lett. 286:9–14. 2009. View Article : Google Scholar : PubMed/NCBI

22 

Shi J, Meng L, Sun MZ, Guo C, Sun X, Lin Q and Liu S: CRKL knockdown promotes in vitro proliferation, migration and invasion, in vivo tumor malignancy and lymph node metastasis of murine hepatocarcinoma Hca-P cells. Biomed Pharmacother. 71:84–90. 2015. View Article : Google Scholar : PubMed/NCBI

23 

Lin Q, Sun MZ, Guo C, Shi J, Chen X and Liu S: CRKL overexpression suppresses in vitro proliferation, invasion and migration of murine hepatocarcinoma Hca-P cells. Biomed Pharmacother. 69:11–17. 2015. View Article : Google Scholar : PubMed/NCBI

24 

Liu S, Guo C, Wang J, Wang B, Qi H and Sun MZ: ANXA11 regulates the tumorigenesis, lymph node metastasis and 5-fluorouracil sensitivity of murine hepatocarcinoma Hca-P cells by targeting c-Jun. Oncotarget. 7:16297–16310. 2016.PubMed/NCBI

25 

Hou L, Li Y, Jia YH, Wang B, Xin Y, Ling MY and Lü S: Molecular mechanism about lymphogenous metastasis of hepatocarcinoma cells in mice. World J Gastroenterol. 7:532–536. 2001. View Article : Google Scholar : PubMed/NCBI

26 

Levin-Arama M, Abraham L, Waner T, Harmelin A, Steinberg DM, Lahav T and Harlev M: Subcutaneous compared with intraperitoneal KetamineXylazine for anesthesia of mice. J Am Assoc Lab Anin Sci. 55:794–800. 2016.

27 

Feliciello I and Chinali G: A modified alkaline lysis method for the preparation of highly purified plasmid DNA from Escherichia coli. Anal Biochem. 212:394–401. 1993. View Article : Google Scholar : PubMed/NCBI

28 

Yamada S, Yanamoto S, Kawasaki G, Rokutanda S, Yonezawa H, Kawakita A and Nemoto TK: Overexpression of CRKII increases migration and invasive potential in oral squamous cell carcinoma. Cancer Lett. 303:84–91. 2011. View Article : Google Scholar : PubMed/NCBI

29 

Liu R, Wang Q, Xu G, Li K, Zhou L and Xu B: The adaptor protein CrkII regulates IGF-1-induced biological behaviors of pancreatic ductal adenocarcinoma. Tumour Biol. 37:817–822. 2016. View Article : Google Scholar : PubMed/NCBI

30 

Askari M and Darabi M, Jahanzad E, Mostakhdemian Hosseini Z, Musavi Chavoshi M and Darabi M: Immunohistochemichal assessment of the CrkII proto-oncogene expression in common malignant salivary gland tumors and pleomorphic adenoma. J Dent Res Dent Clin Dent Prospects. 9:29–34. 2015. View Article : Google Scholar : PubMed/NCBI

31 

Sun MZ, Liu S, Tang J, Wang Z, Gong X, Sun C and Greenaway F: Proteomics analysis of two mice hepatocarcinoma ascites syngeneic cell lines with high and low lymph node metastasis rates provide potential protein markers for tumor malignancy attributes to lymphatic metastasis. Proteomics. 9:3285–3302. 2009. View Article : Google Scholar : PubMed/NCBI

32 

Klemke RL, Leng J, Molander R, Brooks PC, Vuori K and Cheresh DA: CAS/Crk coupling serves as a ‘molecular switch’ for induction of cell migration. J Cell Biol. 140:961–972. 1998. View Article : Google Scholar : PubMed/NCBI

33 

Kiyokawa E, Hashimoto Y, Kurata T, Sugimura H and Matsuda M: Evidence that DOCK180 up-regulates signals from the CrkII-p130(Cas) complex. J Biol Chem. 273:24479–24484. 1998. View Article : Google Scholar : PubMed/NCBI

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June 2019
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Copy and paste a formatted citation
APA
Zhou, Z., Sun, X., Guo, C., Sun, M., & Liu, S. (2019). CRKII overexpression promotes the in vitro proliferation, migration and invasion potential of murine hepatocarcinoma Hca‑P cells. Oncology Letters, 17, 5169-5174. https://doi.org/10.3892/ol.2019.10194
MLA
Zhou, Z., Sun, X., Guo, C., Sun, M., Liu, S."CRKII overexpression promotes the in vitro proliferation, migration and invasion potential of murine hepatocarcinoma Hca‑P cells". Oncology Letters 17.6 (2019): 5169-5174.
Chicago
Zhou, Z., Sun, X., Guo, C., Sun, M., Liu, S."CRKII overexpression promotes the in vitro proliferation, migration and invasion potential of murine hepatocarcinoma Hca‑P cells". Oncology Letters 17, no. 6 (2019): 5169-5174. https://doi.org/10.3892/ol.2019.10194