A domain of methylation change at the albumin locus in rat hepatoma cell variants.
A well-differentiated rat hepatoma cell line, Fu5-5, yields variant clones whose rate of secretion of serum albumin ranges from 40 to less than 0.08 micrograms of albumin/mg of cell protein per 48 h. Clones were classified as high producers (10 to 40 micrograms/mg per 48 h), intermediate producers (1 to 10 micrograms/mg per 48 h), low producers (0.1 to 1.0 micrograms/mg per 48 h), and null variants (less than 0.1 micrograms/mg per 48 h). Albumin synthetic rates are proportional to secretion rates and range from 0.9 to less than 0.002% of total protein synthesis as measured by pulse-labeling. Steady-state albumin mRNA levels were measured by filter hybridization of fragmented, end-labeled mRNA and by Northern blotting. Message levels are proportional to albumin synthetic rates except for a high producer in which albumin mRNA is less elevated than the synthetic rate. The extent of methylation was quantitated at each of 24 CpG-containing sites or site clusters at the albumin locus. These sites span a region that contains the albumin gene as well as 10 kilobases of the 5' flank and 1 kilobase of the 3' flank. An 8-kilobase region is described, with boundaries in the 5' flank and in the middle of the gene, within which all 11 sites examined showed a correlation of undermethylation with the high-producer phenotype. In contrast, 12 of 13 sites outside of this region showed no phenotype correlation. Null variants derived from a high producer underwent de novo methylation of this domain. Six independent hybrid clones derived from the cross of a high producer with a null variant showed extinction of albumin production and hypermethylation of the domain. Apparently these cells retain the capacity for the de novo methylation of these specific sites.
Orlofsky
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A
Chasin
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LA
D000418:Albumins / Q000235:genetics*; D000818:Animals; D002460:Cell Line; D004262:DNA Restriction Enzymes; D005786:Gene Expression Regulation; D006822:Hybrid Cells / Q000502:physiology; D008114:Liver Neoplasms, Experimental / Q000503:physiopathology; D008745:Methylation; D010641:Phenotype; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D014158:Transcription, Genetic
D000418:Albumins; D012333:RNA, Messenger; D004262:DNA Restriction Enzymes