Abstract
The expression of the gene of α-fetoprotein (AFP) was studied in AH-66 hepatoma cells and adult rat liver cells. With purified AFP mRNA as a template, [3H]-labeled AFP complementary deoxyribonucleic acid (cDNA) (around 1000 nucleotides) was synthesized by the use of reverse transcriptase and was used for assaying the nucleotide sequence involved in AFP gene and in its transcripts by the hybridization technique. Upon treatment with DNase I, the AFP gene in AH-66 nuclei was digested preferentially, whereas that in rat liver nuclei was not. These results show that the AFP gene exists in a relaxed conformation in AH-66 but takes a more condensed structure in rat liver. Micrococcal nuclease selectively cleaved the AFP gene in AH-66, as did DNase I. It also preferentially recognized the AFP gene in rat liver, although to a lesser extent than that in AH-66. Chromosomal proteins extracted from AH-66 and ratliver chromatin were divided into four fractions with QAE-Sephadex A-25 (CP0, 0.1, 0.25 and 3 respectively). Liver chromatin was dissociated and reconstituted in the presence of each of AH-66 CP0-3 and, conversely, AH-66 chromatin was reconstituted in the presence of each of liver CP0-3. Reconstituted chromatins were transcribed by E. coli RNA polymerase and the transcripts were annealed to AFP cDNA. AH-66 CP0.1 activated the AFP gene transcription strongly, while none of the liver CP0-3 inhibited the expression of the gene.
