Biological activities of a putative truncated hepatitis B virus X gene product fused to a polylysin stretch.
Truncated transcripts terminating within the HBx frame have been recognized previously in tumor and liver tissue of HCC patients. In this study biological activities of a predicted truncated HBx fused to a polylysin stretch (HBtx-polylysin) and of full length HBx were compared in NIH3T3 cells transfected with respective cDNA plasmids. Transactivation of a co-transfecting reporter gene and influence on neor DNA mediated transformation to G418 resistance were determined. In comparison to full length HBx the data indicate for HBtx-polylysin a lower transactivating capacity and as judged by the yield of colonies on a solid surface, a lower capacity to stimulate neor DNA mediated transformation. In soft agar the outgrowth into G418 resistant colonies was dependent on co-transfecting HBx cDNA. In providing this condition HBtx-polylysin had a much higher relative activity than full length HBx. Large cointegrants consisting of the plasmids carrying truncated HBx cDNA and neor DNA respectively were identified by chromosomal in situ hybridization. Based on Southern blot analyses extended concatemeres of the HBx cDNA plasmid constituted a main part of the cointegrants. Expression of truncated HBx cDNA was followed on the RNA and the protein level. The presence of this cDNA could be correlated to a compact spindle like cell appearance, its loss after prolonged passaging in the absence of G418 to a concomitant reversion to the phenotype of the NIH3T3 cell. Interspersed selection for G418 resistance stabilized the morphologically transformed phenotype. These results provide a basis to manipulate expression of truncated HBx and to recognize thereby processes leading to transformation.
Rakotomahanina
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CK
Department of Virus-Wirtzell-Wechselwirkungen, Deutsches Krebsforschungszentrum, Heidelberg, Germany.
Hilger
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C
Fink
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Zentgraf
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Schröder
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CH
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D002471:Cell Transformation, Neoplastic; D018076:DNA, Complementary / Q000235:genetics; D015870:Gene Expression; D017403:In Situ Hybridization; D051379:Mice; D008969:Molecular Sequence Data; D010957:Plasmids; D011107:Polylysine / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics; D015534:Trans-Activators / Q000235:genetics*; D015533:Transcriptional Activation; D014162:Transfection; D054334:Viral Regulatory and Accessory Proteins
D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; D011107:Polylysine