Abstract
Albumin and alpha-fetoprotein (AFP), two major serum proteins, are synthesized predominantly in the liver and yolk sac of mammals. In the present paper we report on the developmental expression of the corresponding genes in nonhepatic rat tissues. Significant quantities of mature albumin and AFP mRNAs were revealed in kidney, pancreas, heart, and lung of fetal and/or newborn rats using dot blot and Northern blot assays. Very low levels of these mRNA sequences were also detected in adult kidney and pancreas using sensitive RNA-cDNA solution hybridization assays. In situ hybridization analysis revealed that the albumin and AFP gene transcripts are present in the tubular cells of the 20-day-old fetal kidney. In order to elucidate further the mechanisms governing this expression, we studied the chromatin structure and methylation pattern in the 5'-end of these two genes. A faint band, corresponding to a specific DNase I-hypersensitive site upstream from the albumin gene, was detected in the fetal and neonatal kidney nuclei but not in adult kidney. For both genes, a site CG, demethylation of which is correlated with expression in liver and hepatoma cell lines, is highly methylated in fetal kidney even though AFP and albumin genes are expressed. Taken together, these results show the presence of a cell population in the rat kidney that actively transcribes both the albumin and AFP genes. The expression of these genes may be mediated by mechanisms differing in at least some steps from those exerted in the liver.
Highlights
Albumin and a-fetoprotein (AFP), twomajor serum Serum albumin and a-fetoprotein (AFP)’ are two major proteins, are synthesized predominantly in the liver proteins characteristic of hepatic differentiation
In situ hybridization analysis revealed that the albumin and AFP gene transcripts are present in the tubular the prominent product of the adult liver, and it is already produced in large amounts by the hepatocytes of late gestation fetuses (1).AFP is actively synthesized by the hepatocytes of the fetal and newborn rat liver (2-4) and by the endodermal cells of the yolk sac (2, 5)
Albuminaa-nFdetoproGtEeeixnperession inTRiastsues mRNA sequences were about %O of that found in the 18 day Changes in Expression of Albumin and AFP Genes in fetal liver
Summary
The RNA bound to the DBM paper was hybridized to the 3ZP-labeledalbumin or AFP cDNA recombinant plasmids for 48 h a t 42'C in the same buffer with the exception that glycine was omitted and only 250 pg/ml salmon sperm DNA was used. After electrophoresis the RNA was transferred by blotting to DBM (28) or nitrocellulose paper and the hybridization was performed as reported elsewhere (29). Rat albumin and AFP cDNAs were synthesized from highly purified mRNA preparations (8)as reported elsewhere (31), using ["PI dCTP (250 Ci/mmol) as labeled substrate. RNA excess hybridizations with albumin and AFP 32P-labeled cDNAs were carried out as described previously (8, 31). DNA from kidney and liver nuclei was isolated according to the method described previously (37). Transfer of DNA fragments, filter prehybridization and hybridization, washing conditions, and autoradiography were performed as described previously (38). The specific activity of the resulting recombinant t3*P]DNAwas about 26 X 10' dpm/pg
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