Extinction of liver-specific functions in hybrids between differentiated and dedifferentiated rat hepatoma cells

Deschatrette, Jean; Weiss, Mary C.

doi:10.1007/bf01538451

Cited by 47 publications

(26 citation statements)

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“…The extinction of this specialized protein in hybrids between differentiated and dedifferentiated variants has also been observed by Weiss and her co-workers (5,13). That an active albumin gene can be shut off after fusion with an inactive cell implies the existence of a trans-acting negative regulatory signal in the nonproducing parent.…”

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confidence: 58%

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A domain of methylation change at the albumin locus in rat hepatoma cell variants.

Orlofsky¹,

Chasin²

1985

Mol. Cell. Biol.

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A well-differentiated rat hepatoma cell line, Fu5-5, yields variant clones whose rate of secretion of serum albumin ranges from 40 to <0.08 pLg of albumin/mg of cell protein per 48 h. Clones were classified as high producers (10 to 40 ,ug/mg per 48 h), intermediate producers (1 to 10 ,Lg/mg per 48 h), low producers (0.1 to 1.0 ,ug/mg per 48 h), and null variants (<0.1 ,ug/mg per 48 h). Albumin synthetic rates are proportional to secretion rates and range from 0.9 to <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.Stably differentiated cell lines provide attractive material for the study of the regulation of specialized functions. In these systems, a small number of functions can be isolated from the complexity of regulatory interactions in the animal, and homogeneous material for biochemical analysis is readily obtained. A particularly valuable feature is the feasibility of isolating subclones that show heritable variation from the parental phenotype. The elucidation of the molecular basis for such events promises to shed light on the mechanism by which heritable variation (i.e., differentiation) arises in the organism.The rat cell line Fu5-5, derived from a well-differentiated hepatoma (54), has been extensively studied by Weiss and co-workers, who have demonstrated its capacity to undergo a number of regulatory events leading to heritable changes in specialized functions (for review, see reference 55). We have recently described the isolation of a number of subclones from this line that are altered in various liver-specific functions (57, 60). In particular, variants affected in serum albumin synthesis were studied. These clones produce albumin at characteristic rates that vary over a range of three orders of magnitude, from 10% of that found in adult liver to <0.1% (undetectable). Most of this variation is also observable at the level of translat...

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confidence: 58%

“…The extinction of this specialized protein in hybrids between differentiated and dedifferentiated variants has also been observed by 13 citrulline in place of arginine for growth. This clone was one of several whose cells exhibited an altered, more elongated morphology compared with the parental line.…”

mentioning

confidence: 92%

A domain of methylation change at the albumin locus in rat hepatoma cell variants.

Orlofsky¹,

Chasin²

1985

Mol. Cell. Biol.

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“…p4 and H5 cells fail to express the entire group of seven hepatic functions that we study, whereas FaoflC2 cells continue to express one of them; no striking karyotypic modification characterizes these clones (10). The isolation and characterization of the HF and pF hybrids have been described (11,12).…”

Section: Introductionmentioning

confidence: 70%

“…2: reexpression of the Fao phenotype is associated with restoration of the Fao repeat length. Cells of the variant clone p4, like those of H5, not only fail to express the entire group of hepatic functions but also extinguish their expression when fused with the differentiated cells of origin (11). Fig.…”

Section: Methodsmentioning

confidence: 92%

Chromatin repeat length correlates with phenotypic expression in hepatoma cells, their dedifferentiated variants, and somatic hybrids.

Sperling¹,

Weiss²

1980

Proc. Natl. Acad. Sci. U.S.A.

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“…In striking contrast, in dedifferentiated H5 hepatoma cells (12) or in fibroblasts, this promoter is 100-fold less active. The homologous region of the mouse albumin gene was shown to be required for tissue-specific transcription in vitro, using nuclear extracts from liver or other organs (15).…”

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confidence: 84%

The rat albumin promoter: cooperation with upstream elements is required when binding of APF/HNF1 to the proximal element is partially impaired by mutation or bacterial methylation.

et al. 1989

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Mol. Cell. Biol. 9:4750-4758, 1989) [29]). Third, additional sequences sometimes prove to be required in order to obtain the correct expression pattern in transgenic mice (18). This is the case for the albumin gene (31): in transgenic mice, abundant and liver-specific albumin gene transcription requires the presence of a region located between -8.5 and -10.4 kilobases in addition to the promoter.We have previously shown that the 150 base pairs upstream of the rat albumin gene constitutes a fully functional promoter in the differentiated hepatoma cell line H4II (32), where it is as active as the sinian virus 40 early promoter (20). In striking contrast, in dedifferentiated H5 hepatoma cells (12) or in fibroblasts, this promoter is 100-fold less active. The homologous region of the mouse albumin gene was shown to be required for tissue-specific transcription in vitro, using nuclear extracts from liver or other organs (15). We have shown that the tissue-specific promoter activity obtained after transfection relies on six independent positive elements, four of which bind a characterized trans-acting factor in vitro. Distal element II (DEII), at nucleotide -120, binds NF1/CTF, DEI at -90 binds C/EBP20 (1, 5, 27), a CCAAT box at -83 binds ACFINFY (33), and a proximal element (PE) at -60 binds the APF factor (4), also designated HNF-1, LF-B1, or HP1 in other contexts. Present only in hepatocytes and differentiated hepatoma cells, this last factor also participates in the transcriptional control of several other liver-specific genes, including al-antitrypsin (28), pyruvate kinase (S. Vaulont, personal communication), AFP (14,17), and transthyretin (8) genes.The strict tissue distribution of the APF/HNF1 factor, combined with the presence of its target sequence in a 4759

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Extinction of liver-specific functions in hybrids between differentiated and dedifferentiated rat hepatoma cells

Cited by 47 publications

References 20 publications

A domain of methylation change at the albumin locus in rat hepatoma cell variants.

A domain of methylation change at the albumin locus in rat hepatoma cell variants.

Chromatin repeat length correlates with phenotypic expression in hepatoma cells, their dedifferentiated variants, and somatic hybrids.

The rat albumin promoter: cooperation with upstream elements is required when binding of APF/HNF1 to the proximal element is partially impaired by mutation or bacterial methylation.

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