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Open Access Research article

Gankyrin gene deletion followed by proteomic analysis: insight into the roles of Gankyrin in Tumorigenesis and Metastasis

Xue Luo12, Liang Chen1, Jiang Dai1, Yanfei Gao1, Hongli Wang1, Na Wang1, Yongqiang Zhao1, Feng Liu1, Zhihong Sang1, Jie Wang1, Weihua Li1, Kun He1, Baofeng Jin1, Jianghong Man1, Wei Zhang3* and Qing Xia1*

Author Affiliations

1 National Center of Biomedical Analysis, 27 Taiping Road, Beijing, 100850, China

2 Department of Occupational Health, Third Military Medical University, 30 Gaotanyan Road, Chongqing, 400038, China

3 Department of Clinical Laboratory, No. 307 Hospital, Academy of Military Medical Sciences, Beijing, 100071, China

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BMC Medical Genomics 2012, 5:36  doi:10.1186/1755-8794-5-36

Published: 22 August 2012

Abstract

Background

Gankyrin was originally purified and characterized as the p28 component of the 26S proteasome, and later identified as an oncogenic protein in hepatocellular carcinomas (HCC). It has recently been found to be highly expressed in several other malignancies, and compelling evidence show gankyrin plays important roles in tumorigenesis. However, its mechanism of action remains unclear.

Methods

In order to further clarify the functions of gankyrin and better understand its molecular mechanisms, we generated a gankyrin null cell line, HCT116 gankyrin−/− , by targeted homologous recombination in human colon cancer cells, and then employed two-dimensional electrophoresis (2-DE) based proteomic approaches followed by MS identification to investigate alterations in the proteome due to the gankyrin knockout. Western blot and qRT-PCR assays were also used to examine the protein and mRNA levels of some identified proteins.

Results

Compared with wild-type control cells, gankyrin null cells were impaired in terms of their proliferation, migration and anchorage-independent growth. A total of 21 altered proteins were identified, which included 18 proteins that had not previously been reported to be related to gankyrin. Notably, eight metastasis-related proteins were identified. Western blot analyses confirmed that the changes in three examined proteins were consistent with 2-DE gel analysis.

Conclusions

In summary, we have generated a useful cell tool to clarify the functions of gankyrin. Our proteomic data provide novel information to better understand the roles and underlying mechanisms by which gankyrin is involved in tumorigenesis and cancer metastasis.