ARHI (aplasia Ras homologue member I; also known as DIRAS3) is an imprinted tumour suppressor gene, the expression of which is lost in the majority of breast and ovarian cancers. Unlike its homologues Ras and Rap, ARHI functions as a tumour suppressor. Our previous study showed that ARHI can interact with the transcriptional activator STAT3 (signal transducer and activator of transcription 3) and inhibit its nuclear translocation in human breast- and ovarian-cancer cells. To identify proteins that interact with ARHI in nuclear translocation, in the present study, we performed proteomic analysis and identified several importins that can associate with ARHI. To further explore this novel finding, we purified 10 GST (glutathione transferase)–importin fusion proteins (importins 7, 8, 13, β1, α1, α3, α5, α6, α7 and mutant α1). Using a GST-pulldown assay, we found that ARHI can bind strongly to most importins; however, its binding is markedly reduced with an importin α1 mutant that contains an altered NLS (nuclear-localization signal) domain. In addition, an ARHI N-terminal deletion mutant exhibits greatly reduced binding to all importins compared with wild-type ARHI. In nuclear-import assays, the addition of ARHI blocked nuclear localization of phosphorylated STAT3. ARHI also inhibits the interaction of Ran–importin complexes with GFP (green fluorescent protein) fusion proteins that contain an NLS domain and a β-like import receptor-binding domain, thereby blocking their nuclear localization. By conducting GST-pulldown assays, we found that ARHI could compete for Ran-importin binding. Thus ARHI-induced disruption of importin-binding to cargo proteins, including STAT3, could serve as an important regulatory mechanism that contributes to the tumour-suppressor function of ARHI.
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Research Article|
December 15 2009
ARHI (DIRAS3), an imprinted tumour suppressor gene, binds to importins and blocks nuclear import of cargo proteins
Shaoyi Huang;
Shaoyi Huang
*Department of Experimental Therapeutics, The University of Texas, M.D. Anderson Cancer Center, Houston, TX 77030, U.S.A.
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In Soon Chang;
In Soon Chang
†Department of Biochemistry and Molecular Biology, The University of Texas, M.D. Anderson Cancer Center, Houston, TX 77030, U.S.A.
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Wenbo Lin;
Wenbo Lin
§School of Life Sciences, Xiamen University, Xiamen, Fujian 361005, People's Republic of China
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Wenduo Ye;
Wenduo Ye
§School of Life Sciences, Xiamen University, Xiamen, Fujian 361005, People's Republic of China
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Robert Z. Luo;
Robert Z. Luo
*Department of Experimental Therapeutics, The University of Texas, M.D. Anderson Cancer Center, Houston, TX 77030, U.S.A.
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Zhen Lu;
Zhen Lu
*Department of Experimental Therapeutics, The University of Texas, M.D. Anderson Cancer Center, Houston, TX 77030, U.S.A.
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Yiling Lu;
Yiling Lu
‡Department of Systems Biology, The University of Texas, M.D. Anderson Cancer Center, Houston, Texas 77030, U.S.A.
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Ke Zhang;
Ke Zhang
†Department of Biochemistry and Molecular Biology, The University of Texas, M.D. Anderson Cancer Center, Houston, TX 77030, U.S.A.
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Warren S.-L. Liao;
Warren S.-L. Liao
*Department of Experimental Therapeutics, The University of Texas, M.D. Anderson Cancer Center, Houston, TX 77030, U.S.A.
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Tao Tao;
Tao Tao
§School of Life Sciences, Xiamen University, Xiamen, Fujian 361005, People's Republic of China
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Robert C. Bast, Jr;
*Department of Experimental Therapeutics, The University of Texas, M.D. Anderson Cancer Center, Houston, TX 77030, U.S.A.
1To whom correspondence should be addressed (email yinhuay@gmail.com).
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Xiaomin Chen;
Xiaomin Chen
†Department of Biochemistry and Molecular Biology, The University of Texas, M.D. Anderson Cancer Center, Houston, TX 77030, U.S.A.
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Yinhua Yu
Yinhua Yu
1
*Department of Experimental Therapeutics, The University of Texas, M.D. Anderson Cancer Center, Houston, TX 77030, U.S.A.
∥Obstetrics and Gynecology Hospital of Fudan University, Shanghai 200011, People's Republic of China
1To whom correspondence should be addressed (email yinhuay@gmail.com).
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Publisher: Portland Press Ltd
Received:
January 13 2009
Revision Received:
April 22 2009
Accepted:
May 13 2009
Accepted Manuscript online:
May 13 2009
Online ISSN: 1573-4935
Print ISSN: 0144-8463
© The Authors Journal compilation © 2010 Biochemical Society
2010
Biosci Rep (2010) 30 (3): 159–168.
Article history
Received:
January 13 2009
Revision Received:
April 22 2009
Accepted:
May 13 2009
Accepted Manuscript online:
May 13 2009
Citation
Shaoyi Huang, In Soon Chang, Wenbo Lin, Wenduo Ye, Robert Z. Luo, Zhen Lu, Yiling Lu, Ke Zhang, Warren S.-L. Liao, Tao Tao, Robert C. Bast, Xiaomin Chen, Yinhua Yu; ARHI (DIRAS3), an imprinted tumour suppressor gene, binds to importins and blocks nuclear import of cargo proteins. Biosci Rep 1 June 2010; 30 (3): 159–168. doi: https://doi.org/10.1042/BSR20090008
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