Increased translatable messenger ribonucleic acid for argininosuccinate synthetase in canavanine-resistant human cells

Su, Tsung‐Sheng; Beaudet, Arthur L.; O’Brien, William E.

doi:10.1021/bi00513a037

Cited by 67 publications

(27 citation statements)

References 22 publications

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“…It was previously shown that the human argininosuccinate synthetase gene was not subject to arginine-mediated repression in canavanine-resistant variants of RPMI 2650 cells (11,30). To determine whether the CAT minigenes were capable of showing the appropriate cell-specific regulation, we introduced the CAT minigenes into the Canr-1 cell line (31) using RSVneo as a selectable marker.…”

Section: Methodsmentioning

confidence: 99%

“…Argininosuccinate lyase, which converts argininosuccinate into arginine, is not subject to arginine-mediated repression (11,30). This indicates that the feedback regulation is specific for the argininosuccinate synthetase locus.…”

Section: Human Argininosuccinate Synthetase Minigenesmentioning

confidence: 99%

“…Argininosuccinate synthetase expression is also altered in cultured cells resistant to canavanine, a toxic arginine analog (12,30). Canavanine-resistant (Can9 cells can have up to 300-fold-higher levels of argininosuccinate synthetase as compared with wild-type cells.…”

Section: Human Argininosuccinate Synthetase Minigenesmentioning

confidence: 99%

“…Argininosuccinate synthetase plays a role in urea synthesis in liver and is involved in arginine biosynthesis in nonhepatic tissues. The negative-feedback regulation was first described in rodent and human cells by Schimke (27) and later in other mammalian cell lines such as hamster fibroblasts (9), human lymphoblasts (11), and a human squamous cell carcinoma cell line, RPMI 2650 (30). Cells grown in medium in which citrulline, a substrate for the argininosuccinate synthetase reaction, is substituted for arginine have 3-to 150-fold-higher levels of argininosuccinate synthetase activity relative to cells grown in medium containing arginine.…”

mentioning

confidence: 97%

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Human argininosuccinate synthetase minigenes are subject to arginine-mediated repression but not to trans induction.

Boyce¹,

Anderson²,

Rusk³

et al. 1986

Mol. Cell. Biol.

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The human argininosuccinate synthetase locus is subject to metabolite-mediated repression by arginine in some cultured cefl lines. To gain insight into the mechanism underlying this regulation, chloramphenicol acetyltransferase (CAT) minigenes under the transcriptional control of the human argininosuccinate synthetase promoter were constructed and tested for regulation. When the minigenes were introduced into RPMI 2650 cells, a human cell line that shows sixfold regulation of the argininosuccinate synthetase gene, CAT expression was repressed three-to fivefold when arginine was present in the culture medium. A minigene containing only 149 base pairs of 5'-flanking sequence was expressed at similar levels and regulated to the same degree as one having approximately 3 kilobases of 5'-flanking sequence. Therefore, the cis-acting sequences required for the arginine-mediated repression are likely to be located within the region of the transcription initiation site. The arginine-mediated repression of the CAT minigenes was not observed in canavanine-resistant variants of RPMI 2650 cells, and therefore they showed the appropriate cell-type specificity. Cultured cells having 200-foldincreased levels of argininosuccinate synthetase can be selected by growth in medium containing the arginine analog canavanine. It was previously demonstrated that the increased expression of argininosuccinate synthetase in canavanine-resistant human lymphoblasts was due to a trans-acting mechanism. To gain further support for a trans-acting mechanism, we tested our CAT minigenes for the trans induction in canavanineresistant variants of RPMI 2650 cells. Transfection of the CAT minigenes into RPMI 2650 cells and canavanine-resistant variants of this cell line yielded no difference in transient CAT expression. Furthermore, cloned canavanine-resistant variant cells having integrated copies of the CAT minigenes expressed CAT at similar levels as compared to the parental cell lines. Since these cell lines do exhibit argin'ine-mediated repression of CAT but not trans induction, these data indicate that the arginine-mediated repression is a regulatory event that occurs independently of the trans induction.There are many examples of eucaryotic genes that are regulated in trans by hormones (13, 16, 25, 26), metals (3, 13, 25, 29), and viral or cellular gene products (10,22). In most cases studied thus far, regulation occurs at the level of gene transcription such that a rapid change in transcription is concomitant with addition of the factor mediating the regulation. It has been shown for glucocorticoid hormone regulation of both cellular and viral genes that a hormonereceptor complex exerts its effect by binding to regions 5' to or within the regulated gene and thereby stimulating transcription. The cis-acting sequences involved in the regulation have been mapped by gene transfer with cloned DNA and by various physical techniques that are based on stable interactions between the cis-acting element and the regulator (20,21). Indirect evidence for c...

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Section: Methodsmentioning

confidence: 99%

Section: Human Argininosuccinate Synthetase Minigenesmentioning

confidence: 99%

Section: Human Argininosuccinate Synthetase Minigenesmentioning

confidence: 99%

mentioning

confidence: 97%

See 2 more Smart Citations

Human argininosuccinate synthetase minigenes are subject to arginine-mediated repression but not to trans induction.

Boyce¹,

Anderson²,

Rusk³

et al. 1986

Mol. Cell. Biol.

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“…Similar metabolite regulation of AS was observed in cultured human lymphoblasts (10, 11) and in a human epitheloid cell line, RPMI-2650 (18). A second aspect of regulation of AS in cultured cells involves overproduction of the enzyme in cell variants selected for resistance to the toxic arginine analog canavanine (11,18 (Fig. 1).…”

mentioning

confidence: 99%

Arginine-mediated regulation of an argininosuccinate synthetase minigene in normal and canavanine-resistant human cells.

Jackson¹,

O’Brien²,

Beaudet³

1986

Mol. Cell. Biol.

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Regulation of argininosuccinate synthetase (AS) was studied by using minigenes containing 3 kilobases of DNA upstream from the TATAA box and 9 kilobases downstream (including the first four exons of the AS gene) ligated to either the cDNA for AS or to the chloramphenicol acetyltransferase (CAT) gene. Unlike the endogenous AS gene, expression of the CAT minigene was not elevated in Canrl cells, which overproduce AS compared with parental RPMI-2650 cells. Expression of the CAT minigene in both stable and transient analyses was four-to five-fold higher in RPMI-2650 cells grown in citrulline medium than in cells grown in arginine medium. Although endogenous AS activity is not subject to metabolite regulation in Canrl cells and expression of the CAT minigene in Can 1 cells was not increased when cells were grown in citrulline medium, expression of the CAT minigene was 10-to 22-fold greater when intracellular arginine pools were depleted by transient starvation for arginine and citrulline.Relatively few systems are available for the study of metabolite regulation of gene expression in cultured animal cells. Schimke (16) reported low levels of argininosuccinate cynthetase (AS) in cultured cells grown in medium with a nigh concentration of arginine. In contrast, AS activity was increased when cells were grown in medium with citrulline s:ibstituted for arginine or in medium with a low concentratiorn of arginine. Similar metabolite regulation of AS was observed in cultured human lymphoblasts (10, 11) and in a human epitheloid cell line, RPMI-2650 (18). A second aspect of regulation of AS in cultured cells involves overproduction of the enzyme in cell variants selected for resistance to the toxic arginine analog canavanine (11,18 (Fig. 1). The constructions used portions of the pSVOcat plasmid (8), which lacks the simian virus 40 viral promotor and enhancer elements. The CAT construction should synthesize authentic CAT protein rather than a fusion protein due to the presence of termination * Corresponding author. codons in the AS reading frame, to the 5' side of the CAT coding sequences. A small segment of DNA between the two HindIll sites in intron 1 was lost from both constructions.Expression of pMJ311 in Chinese hamster cells. The ability of the AS minigene to direct the synthesis of functional AS was tested in Chinese hamster cells, which do not express endogenous AS activity. RJK88 cells (7) were cotransfected with pMJ311 and pSV2gpt (14) by a modification of the calcium phosphate coprecipitation method (9). Stable transformants were selected either in citrulline medium or in HAT (hypoxanthine-aminopterin-thymidine) medium (7) to evaluate expression of AS with and without selection pressure. The activities were similar irrespective of the selection scheme (1.59 + 0.65 mU/mg of protein) and fell within the range of AS activity seen in cultured human fibroblasts (1). mRNA initiation sites. RNase protection experiments were performed to determine whether the CAT minigene transcripts were correctly initiated compared with...

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Gene Amplification

Tlsty

2002

Encyclopedia of Molecular Biology

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Increased translatable messenger ribonucleic acid for argininosuccinate synthetase in canavanine-resistant human cells

Cited by 67 publications

References 22 publications

Human argininosuccinate synthetase minigenes are subject to arginine-mediated repression but not to trans induction.

Human argininosuccinate synthetase minigenes are subject to arginine-mediated repression but not to trans induction.

Arginine-mediated regulation of an argininosuccinate synthetase minigene in normal and canavanine-resistant human cells.

Gene Amplification

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