Analysis of the inductive effect of the genomic equivalent of HALF1 sequence in the reversion of rat dedifferentiated hepatoma cells.
HALF1 (human activator of liver function 1) is a closed DNA duplex implicated in reversion of rat dedifferentiated C2 hepatoma cells to a well-differentiated state. A copy of HALF1 is found in high-molecular-weight DNA in the human genome. The genomic equivalent of HALF1 and its flanking sequences [gH(5'-3') fragment] have previously been cloned and sequenced. To analyze the ability of the gH(5'-3') fragment to induce reversion process of C2 cells, two series of transfections were performed: (1) cotransfection of gH(5'-3') and plasmid pSVneo1 and (2) transfection of gH(5'-3') inserted into pSVneo1. The frequency of reversion was enhanced in transfected cells from the first experiment whereas no revertants were obtained from transfected cells in the second one. DNA analysis of the revertant clones revealed that reversion is associated with the transient presence of nonintegrated gH(5'-3') molecules. C2 cells were also transfected with gH(5'-3') cloned in pSV2dhfr. In the products of this transfection, the genesis of revertants correlated with amplification process of the gH(5'-3') sequence. We conclude that the presence of integrated copies of gH(5'-3'), even in high copy number, is not sufficient to induce the reversion process. We propose that extrachromosomal forms of gH(5'-3'), either given to cells or formed during amplification cycles, are involved in the reversion process of C2 cells.
Boccaccio
C
C
INSERM U347, Le Kremlin-Bicêtre, France.
Meunier-Rotival
M
M
Deschatrette
J
J
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D002454:Cell Differentiation; D002999:Clone Cells; D003001:Cloning, Molecular; D017931:DNA Primers; D004270:DNA, Circular / Q000235:genetics* / Q000378:metabolism; D004273:DNA, Neoplasm / Q000235:genetics* / Q000378:metabolism; D015894:Genome, Human; D006801:Humans; D008114:Liver Neoplasms, Experimental; D008727:Methotrexate / Q000633:toxicity; D010957:Plasmids; D016133:Polymerase Chain Reaction; D051381:Rats; D015183:Restriction Mapping; D014162:Transfection; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D004270:DNA, Circular; D004273:DNA, Neoplasm; D008727:Methotrexate