Upregulation of the truncated basic hair keratin 1(hHb1‐ΔN) in carcinoma cells by Epstein‐Barr virus (EBV)

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Abstract
To investigate the role of Epstein‐Barr virus (EBV) in epithelial tumors, we compared the expression pattern of cellular genes in the EBV‐infected gastric carcinoma cell line, NU‐GC‐3, and its uninfected control. Subtractive suppression hybridization (SSH) was combined with high‐density DNA array screening to identify differentially expressed genes. We have discovered that EBV infection upregulated a truncated variant of human basic hair keratin 1 (hHb1‐ΔN), a gene that had previously been identified in metastatic breast carcinoma. We verified the differential expression of hHb1‐ΔN in 3 independent EBV‐positive and ‐negative NU‐GC‐3 clones by Northern blotting. We further verified the EBV‐dependent upregulation of hHb1‐ΔN in 3 other carcinoma cell lines (AGS, TWO3 and DLD1) by RT‐PCR. Inhibition of CpG methylation by 5‐Aza‐CdR induced hHb1‐ΔN mRNA expression in the EBV‐negative clones but did not alter the expression in the EBV‐positive clones. The expression of hHb1‐ΔN protein was detectable by immunofluorescence and Western blotting in EBV‐positive but not in EBV‐negative NU‐GC‐3 clones after proteasome inhibitor (MG132) treatment. hHb1‐ΔN protein formed fibrous structures in the cytoplasm and accumulated in distinct nuclear bodies in the euchromatic areas of the cell nucleus. We suggest that the unstable hHb1‐ΔN protein may inhibit some of the functions of the keratin cytoskeleton and/or interfere with transcription regulation. It also may establish a link between EBV and the low differentiated or anaplastic status of the carcinomas that carry the virus.

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