The temperature-sensitive RLA209-15 fetal rat hepatocyte line grown at the nonpermissive temperature (40°C, normal phenotype) produces authentic rat α-fetoproteins (AFPs) of 69K and 73K (fetal AFPs) which are encoded by a 2.2-kb mRNA. These cells also produce low levels of a 1.7-kb AFP mRNA and a 65K variant AFP when grown at the permissive temperature (33°C, transformed phenotype). Hybrid-selected translation demonstrates that the 1.7-kb AFP mRNA encodes the 65K variant AFP. Northern blot hybridization and S1 nuclease analyses indicate that the 1.7-kb mRNA lacks sequences present in the first seven 5′ exons of the 2.2-kb AFP mRNA. However, the 1.7- and 2.2-kb AFP mRNAs share common sequences extending from the beginning of the eighth exon (corresponding to nucleotide 873 of the fetal AFP mRNA) to the 3′ end. Primer extension analysis suggests that the 1.7-kb RNA contains additional sequences 5′ to the common regions shared by both AFP mRNAs. We have previously shown that adult rat liver produces a 1.7-kb AFP mRNA; we now report the isolation of a cDNA (ARFP5) encoding this variant AFP mRNA from an adult rat liver cDNA library. Restriction endonuclease mapping and sequence analysis of ARFP5 confirm that the 1.7- and 2.2-kb AFP mRNAs share similar sequences at the 3′ region (approximately 1.1 kb). However, ARFP5 contains an additional 90 bp variant AFP mRNA-specific 5′ sequence which is located in the seventh intron of the rat AFP gene. In addition to RLA209-15 and adult rat liver, a rat hepatoma cell line (CRL 1548) expresses the 1.7-kb but not the 2.2-kb AFP mRNA. The 1.7-kb AFP mRNA in CRL1548 cells, like the variant AFP mRNA of RLA209-15 cells, hybridizes to a probe containing either the 3′ sequence of the fetal AFP mRNA or the 90 bp 5′ sequence of ARFP5.
|Original language||English (US)|
|Number of pages||8|
|State||Published - 1988|
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