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T S Su

Publications and source records attributed to T S Su.

49 records · Page 3Linked to original sources
PubMed1986

Hepatitis B virus transcripts in a human hepatoma cell line, Hep 3B.

Hep 3B, a human hepatoma cell line was examined for its RNA hybridizable to the hepatitis B virus sequence. Using probes that covered different regions of the hepatitis B virus genome, five species of RNA were observed of sizes 4.0, 3.3, 2.9, 2.6 and 2.2 kilobases. The RNAs covered surface antigen gene, pre-S and X regions. None of them had a core antigen sequence. RNA with a 4.0 kilobase size was the most abundant. Using S1 nuclease analysis, its 5' end of hepatitis B virus sequence was mapped at pre-S region and its 3' end of viral sequence was mapped at DR region.

Antigens, Neoplasm↗
PubMed1985

Expression of c-myc gene in human hepatoma.

The level of c-myc transcript was examined in liver samples from seven hepatoma patients. Transcripts were detected in all the normal liver parts examined; in contrast, in two hepatoma parts, there was a dramatic reduction in c-myc transcripts. The restriction enzyme pattern of c-myc gene appeared the same among samples. The data suggest that c-myc gene expression might not be required for the maintenance of the tumor state in human liver carcinogenesis.

Carcinoma, Hepatocellular↗
PubMed1984

Genomic DNA-mediated gene transfer for argininosuccinate synthetase.

Canavanine-resistant (Canr) human cells overproduce argininosuccinate synthetase without the occurrence of gene amplification. Using calcium phosphate precipitation, genomic DNA from Canr human cells was used to carry out gene transfer into Chinese hamster cells, which do not express argininosuccinate synthetase activity. Growth in tissue culture medium with citrulline substituted for arginine was adequate to select enzyme-positive colonies. Six independent isolates were selected for detailed analysis by enzyme assay, Southern blotting, Northern blotting, and S1 nuclease analysis, the last of which distinguishes human and hamster mRNA. Five isolates were transferrants containing the human structural gene and synthesizing human enzyme. One isolate represented a cell line synthesizing Chinese hamster enzyme. The data document gene transfer of DNA fragments at least 80 kb in length, the low level of spontaneous activation of the argininosuccinate synthetase locus in Chinese hamster cells, the feasibility of this expression and selection system for DNA-mediated gene transfer, and a method for distinguishing the human and hamster gene products at an RNA level.

Animals↗
PubMed1984

Multiple, independent restriction site polymorphisms in human DNA detected with a cDNA probe to argininosuccinate synthetase (AS).

We have used a cDNA clone of the human urea cycle enzyme argininosuccinate synthetase (AS) to screen for restriction fragment length polymorphisms (RFLPs) using a large panel of restriction enzymes. The probe, pAS-1, detects from 15 to 27 human DNA fragments by Southern gel analysis. In addition to the structural locus on chromosome 9, AS-like sequences are found on at least 10 human chromosomes, including the X and Y. This large number of dispersed pseudogenes accounts for the multiplicity of hybridizing fragments detected with pAS-1. Of 37 restriction enzymes tested, 18 produced excellent digest patterns; of these 18 enzymes, three revealed high-frequency, independent RFLPs, testing a minimum of 16 individuals with each enzyme. The enzymes producing high-frequency polymorphisms are Hind III (allele frequencies .30 and .70), Hind II (.13 and .87), and Bam HI (.56 and .44). Most of the polymorphic alleles are found in Caucasians, American blacks, and Orientals. The RFLP detected with Hind III maps to chromosome 9 (9q11-q22), although not to the structural locus; the others are autosomal but otherwise unassigned. Two additional six-base enzymes and a four-base enzyme, Msp I, revealed further individual variation, but these variants have not been shown to segregate in families yet. We observed approximately 0.8% high-frequency RFLPs per cut site per enzyme or a minimum of .05% heterozygosity per nucleotide. pAS-1 should prove useful in assigning high-frequency RFLPs to several human chromosomes.

Adult↗
PubMed1984

Human chromosomal assignments for 14 argininosuccinate synthetase pseudogenes: cloned DNAs as reagents for cytogenetic analysis.

There are multiple, processed, dispersed pseudogenes for human argininosuccinate synthetase. Chinese hamster X human somatic cell hybrids were used to map DNA fragment groups corresponding to the single expressed gene and 14 pseudogene loci. Each chromosomal assignment was confirmed using hybrids containing very few human chromosomes and/or by demonstrating monosomic or trisomic dosage in human cell lines with chromosomal abnormalities. Pseudogenes were mapped to chromosomes 2cen-p25, 3q12-qter, 4q21-qter, 5 (two loci), 6, 7, 9p13-q11, 9q11-q22, 11q, 12, Xp22-pter, Xq22-q26, and Ycen-q11. DNA fragments from the expressed gene were mapped to 9q34-qter in agreement with the previous assignment for enzyme activity. A high-frequency restriction fragment length polymorphism mapped to 9q11-q22. The analyses emphasized the feasibility of using chromosomally abnormal human cell lines for confirmation and regionalization of gene-mapping assignments made using somatic-cell hybrids. Conversely, cloned DNA probes, once mapped and characterized, can be very valuable for determining the chromosomal composition of interspecies hybrids and the dosage of loci in human cells. The argininosuccinate synthetase cDNA is a convenient reagent for dosage analysis of 15 human loci on 11 different chromosomes. Improved reagents could be designed that would simplify Southern blot patterns by eliminating overlapping DNA fragments and providing a single DNA fragment for each locus.

Animals↗
PubMed1983

Sequence for human argininosuccinate synthetase cDNA.

The nucleotide sequence for human argininosuccinate synthetase cDNA was determined by analysis of six clones isolated from a single experiment. The sequence covered 1623 nucleotides including 76 bases of poly(A) and contained a 1236 nucleotide open reading frame encoding a protein of 46,434 daltons. In one cDNA isolate, a cloning artifact or perhaps RNA polymerase error involving addition of an A in a region of six A's within the coding sequence was documented. Single base variations in the 3' untranslated region were examined in detail since detection of DNA polymorphisms in the cDNAs could imply over-expression of both alleles at the active locus in canavanine-resistant cells, i.e. a trans-acting mechanism for enzyme overproduction. However, the sequence from five cDNAs suggested some single base artifacts, and DNA polymorphism remains uncertain. The occurrence of three tandem arginine codons in the 5' untranslated region of the cDNA suggested the possibility of an interaction of arginyl-tRNA with mRNA to regulate RNA processing or half-life as a mechanism for arginine-mediated repression.

Amino Acid Sequence↗
PubMed1983

Abnormal mRNA for argininosuccinate synthetase in citrullinaemia.

Citrullinaemia is a human inborn error of metabolism resulting from the deficiency of argininosuccinate synthetase. In a previous study of cultured skin fibroblasts from citrullinaemia patients, we showed that the presumed defects in DNA were not detectable by Southern blotting analysis, and that only 2 of 11 cell lines contained detectable enzyme antigen. All citrullinaemia cell lines contained hybridizable mRNA but slight size heterogeneity was noted. Here we report the extension of the analysis of the RNA using S1 nuclease mapping techniques. Among six cell lines examined, five showed an abnormality of mRNA detectable by S1 nuclease analysis. The data indicate that a minimum of three out of five non-consanguineous patients represent compound heterozygotes. The S1 nuclease detectable defects may represent deletions or rearrangements in the genomic DNA, or more probably represent examples of abnormal RNA splicing. The approach used here is useful for molecular analysis of genetic defects, for prenatal diagnosis, and for study of genetic variation.

Amino Acid Metabolism, Inborn Errors↗
PubMed1982

Dispersion of argininosuccinate-synthetase-like human genes to multiple autosomes and the X chromosome.

DNA sequences closely homologous to argininosuccinate synthetase are present at ten or more distinct locations in the human genome, including sites on chromosomes 6, 9 and X. Argininosuccinate synthetase thus represents one of the most widely dispersed multigene families described to date, the first instance of a multigene family associated with an enzyme of intermediary metabolism and, perhaps most striking, the first instance of a multigene family with members on both autosomes and sex chromosomes.

Argininosuccinate Synthase↗
PubMed1982

Molecular analysis of argininosuccinate synthetase deficiency in human fibroblasts.

We have analyzed cultured skin fibroblasts derived from patients with argininosuccinate synthetase deficiency for alterations in gene structure, mRNA content, and protein structure. Genomic DNA was digested with the endonucleases EcoRI or HindIII, and the fragments were analyzed by Southern blotting and hybridization with a cDNA probe for argininosuccinate synthetase. The blot pattern is complex because there are at least 10 copies of argininosuccinate synthetase-like genes scattered over multiple human chromosomes. All nine patients studied showed patterns of DNA fragments that were indistinguishable from the normal control cell lines, and despite the possibility that the complexity could mask some changes, major deletions of the active gene(s) were not present. Blot hybridization of RNA indicated the presence of hybridizable mRNA of approximately normal size in seven of seven individuals examined with a suggestion of some heterogeneity. Analysis of enzyme antigen by protein transfer from NaDodSO4 containing polyacrylamide gels revealed considerable heterogeneity. This analysis revealed no cross-reacting material (CRM) in nine cell lines, CRM of normal molecular weight in one cell line, and CRM of reduced molecular weight in one cell line. These findings suggest that the genes for argininosuccinate synthetase in most citrullinemia patients are transcribed and produce stable mRNA. These mRNA either are not translated, or the translation product (enzyme) is rapidly degraded or is immunologically nonreactive. Defective gene expression in this disorder appears to involve abnormal mRNA, which may be altered by point mutations, frame shift mutations, deletions, insertions or particularly by abnormal RNA processing.

Argininosuccinate Synthase↗
PubMed1981

Cloning of cDNA for argininosuccinate synthetase mRNA and study of enzyme overproduction in a human cell line.

Previous studies of the human cell line RPMI-2650 (wild type) and its canavanine-resistant variants have demonstrated differences in argininosuccinate synthetase activity as follows: canavanine-resistant much greater than wild type grown in citrulline greater than wild type grown in arginine (Su, T.-S., Beaudet, A. L., and O'Brien, W. E. (1981) Biochemistry 20, 2956-2960). A recombinant plasmid containing a 1.55-kilobase insert complementary to the mRNA for human argininosuccinate synthetase was isolated by the combined use of differential colony hybridization and immunoprecipitation of the products of plasmid-selected mRNA translation. Both blot and dot hybridization analysis of polyadenylated RNA indicated a major mRNA species of 1.67 kilobase in all cells, and the levels of mRNA correlated well with the levels of enzyme activity: canavanine-resistant, 180; wild type grown in citrulline, 7; and wild type grown in arginine, 1. One major mRNA species of 1.67 kilobase and one minor species of 2.68 kilobase were observed in wild type and canavanine-resistant cell lines. Reassociation kinetics of pAS1 with genomic DNA from human liver, canavanine-resistant cells, and wild type cells were not significantly different. Blot hybridization of genomic DNA revealed no detectable differences between wild type cells, canavanine-resistant cells, and human leukocytes. The data demonstrated that there were multiple copies, perhaps 10 or more, of argininosuccinate synthetase-like sequences in human DNA and that the canavanine-resistant phenotype was not due to gene amplification.

Argininosuccinate Synthase↗
PubMed1981

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

The level of argininosuccinate synthetase activity in the human tissue culture cell line RPMI 2650 was 6-fold higher when citrulline was substituted for arginine in the culture medium. Canavanine-resistant (Canr) variants were isolated and had enzyme activity up to 25 nmol min-1 (mg of protein)-1 or 180-fold higher than that of the wild-type cells grown in arginine. The differences in enzyme activity were paralleled by differences in the amount of enzyme determined immunologically. The micrograms of enzyme per milligrams of protein, determined by complement fixation, were 0.03 for wild-type cells grown in arginine, 0.29 for wild-type cells grown in citrulline, and 6.73 for a Canr variant. In vivo labeling studies suggested increased synthesis of argininosuccinate synthetase in Canr cells, and in vitro translation of poly(adenylic acid) [poly(A)] messenger ribonucleic acid (mRNA) from wild-type and Canr cells confirmed a quantitatively compatible increase in translatable poly(A) mRNA for the enzyme in Canr cells. No precursor for the enzyme was recognized by using in vitro translation, and the poly(A) mRNA for the enzyme had a sedimentation value of 16 S by sucrose-gradient analysis. The levels of argininosuccinate synthetase activity in the Canr cells were similar to those found in normal liver.

Animals↗