Viral Hepatitis
Copyright ©The Author(s) 2004. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Oct 1, 2004; 10(19): 2805-2808
Published online Oct 1, 2004. doi: 10.3748/wjg.v10.i19.2805
Cytochrome C oxidase III interacts with hepatitis B virus X protein in vivo by yeast two-hybrid system
Dan Li, Xiao-Zhong Wang, Jie-Ping Yu, Zhi-Xin Chen, Yue-Hong Huang, Qi-Min Tao
Dan Li, Jie-Ping Yu, Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan 430060, Hubei Province, China
Xiao-Zhong Wang, Zhi-Xin Chen, Yue-Hong Huang, Department of Gastroenterology, Union Hospital of Fujian Medical University, Fuzhou 350001, Fujian Province, China
Qi-Min Tao, Research Institute of Hepatology, Beijing University, Beijing 100044, China
Author contributions: All authors contributed equally to the work.
Supported by Science and Technology Program of Fujian Province, No. 2001Z157
Correspondence to: Dr. Dan Li, Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan 430060, Hubei Province, China.
Telephone: +86-27-88043424-8508
Received: December 10, 2003
Revised: January 24, 2004
Accepted: February 1, 2004
Published online: October 1, 2004

AIM: To screen and identify the proteins which interact with hepatitis B virus (HBV) X protein in hepatocytes by yeast two-hybrid system and to explore the effects of X protein in the development of hepatocellular carcinoma (HCC).

METHODS: With HBV X gene amplified by polymerase chain reaction (PCR), HBV X bait plasmid, named pAS2-1-X, was constructed by yeast-two hybridization system3 and verified by auto-sequencing assay. pAS2-1-X was transformed into the yeast AH109, and X-BD fusion protein expressed in the yeast cells was detected by Western blotting. The yeast cells cotransformed with pAS2-1-X and normal human liver cDNA library were grown in selective SC/-trp-leu-his-ade medium. The second screen was performed with β-gal activity detection, and false positive clones were eliminated by segregation analysis, true positive clones were amplified, sequenced and analyzed with bioinformatics. Mating experiment was peformed to confirm the binding of putative proteins to X protein in the yeast cells.

RESULTS: Bait plasmid pAS2-1-X was successfully constructed and pAS2-1-X correctly expressed BD-X fusion protein in yeast AH109. One hundred and three clones grew in the selective SC/-trp-leu-his-ade medium, and only one clone passed through β-gal activity detection and segregation analysis. The inserted cDNA fragment showed high homology with Homo sapiens cytochrome C oxidase III (cox III). Furthermore, mating experiment identified that the binding of cox III to X protein was specific.

CONCLUSION: cox III protein is a novel protein that can interact with X protein in vivo by yeast two-hybrid system, and may contribute to the development of HCC through the interaction with X protein.

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