Up-regulation and subcellular localization of hnRNP A2/B1 in the development of hepatocellular carcinoma.
BACKGROUND
Hepatocellular carcinoma (HCC) is one of the world's leading causes of death among cancer patients. It is important to find a new biomarker that diagnoses HCC and monitors its treatment. In our previous work, we screened a single-chain antibody (scFv) N14, which could specifically recognize human HepG2 HCC cells but not human non-cancerous liver LO2 cells. However, the antigen it recognized in the cells remained unknown.


METHODS
Recombinant scFv N14 antibody was expressed as an active antibody. Using this antibody with a combination of immunological and proteomic approaches, we identified the antigen of scFv N14 antibody as the heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1). The expression of hnRNP A2/B1 in HCC cells was then investigated by semi-quantitative RT-PCR and immunohistochemistry.


RESULTS
We found that the up-regulation of hnRNP A2/B1 was measured at both transcriptional and translational levels in rat HCC cells but not in rat hepatic cells. We also found that in various human hepatic tissues, hnRNP A2/B1 was highly expressed in both human hepatitis virus positive liver tissues and human HCC tissues but not in normal liver tissues. Interestingly, we observed that the localization of hnRNP A2/B1 in HCC cells was altered during the development of HCC. In human hepatitis virus infected tissues hnRNP A2/B1 resides exclusively in the nuclei of hepatocytes. However, when the HCC progressed from a well differentiated to a poorly differentiated stage, hnRNP A2/B1 was increasingly localized in the cytoplasm. In contrast, the HCC tissues with hnRNP A2/B1 highly expressed in the nucleus decreased.


CONCLUSIONS
This work is the first to show that hnRNP A2/B1 is the antigen specifically recognized by the scFv N14 antibody in HCC cells. The over-expression of hnRNP A2/B1 was confirmed in cultured human and rat HCC cell lines, human virus related hepatitis liver tissues and human HCC tissues. The increased localization of hnRNP A2/B1 in the cytoplasm of HCC cells was revealed during the dedifferentiation of hepatocellular carcinoma. Therefore, we suggest that the increased expression and cytoplasmic localization of hnRNP A2/B1 can be used as a diagnostic biomarker to assess the risk of human liver cancer.
Cui
Huaqing
H
State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, No 33 Life Science Park Road, Beijing 102206, China.
Wu
Feng
F
Sun
Yanling
Y
Fan
Guocai
G
Wang
Qingming
Q
D000595:Amino Acid Sequence; D000818:Animals; D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism* / Q000473:pathology; D005260:Female; D005455:Fluorescent Antibody Technique; D022781:Hepatocytes / Q000276:immunology / Q000378:metabolism / Q000473:pathology; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D007124:Immunoenzyme Techniques; D008099:Liver / Q000378:metabolism / Q000473:pathology; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D015854:Up-Regulation
D014408:Biomarkers, Tumor; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D012333:RNA, Messenger; D057127:Single-Chain Antibodies; C439768:hnRNP A2