Biomedical subjects

A M Killary

Publications and source records attributed to A M Killary.

41 records · Page 3Linked to original sources

PubMed1986

Assignment of genes encoding dihydrofolate reductase and hexosaminidase B to Mus musculus chromosome 13.

The murine gene encoding dihydrofolate reductase (DHFR) has been localized to a particular mouse chromosome by complementation mapping. Microcells prepared from diploid mouse fibroblasts were fused with mutant hamster cells lacking the dihydrofolate reductase gene (Dhfr), and DHFR+ microcell hybrids were selected in medium lacking purines and pyrimidines. The complemented hybrids expressed wild-type levels of DHFR enzyme activity, and selectively retained a single mouse chromosome--chromosome 13. Genomic Southern blots of DNAs prepared from these hybrids and from an independently derived collection of clones isolated without selection on the DHFR phenotype confirmed the assignment of murine Dhfr gene sequences to chromosome 13. This assignment provides further evidence for the existence of genetic homology between regions of mouse chromosome 13 and human chromosome 5. In support of this view, we show that the gene encoding hexosamindase B, a chromosome 5 marker in man, also maps to mouse chromosome 13.

Animals↗

PubMed1985

Chromosomal assignment and trans regulation of the tyrosine aminotransferase structural gene in hepatoma hybrid cells.

The structural gene encoding liver-specific tyrosine aminotransferase (TAT; EC 2.6.1.5) was assigned to mouse chromosome 8 by screening a series of hybrid cell lines for retention of murine Tat-1 gene sequences by genomic Southern blotting. This assignment demonstrated that the Tat-1 structural gene was not syntenic with Tse-1, a chromosome 11-linked locus that negatively regulates TAT expression in trans (A. M. Killary and R. E. K. Fournier, Cell 38:523-534, 1984). We also showed that the fibroblast Tat-1 gene was systematically activated in hepatoma X fibroblast hybrids retaining fibroblast chromosomes 8 in the absence of chromosome 11 but was extinguished in cells retaining both fibroblast chromosomes. Thus, the TAT structural genes of both parental cell types were coordinately regulated in the intertypic hybrids, and the TAT phenotype of the cells was determined by the presence or absence of fibroblast Tse-1.

Animals↗

PubMed1984

Isolation of thymidine kinase-deficient rat hepatoma cells by selection with bromodeoxyuridine, Hoechst 33258, and visible light.

The photosensitivity of bromodeoxyuridine (BrdU)-substituted cells is known to be markedly enhanced by the fluorochrome Hoechst 33258. Since the incorporation of BrdU into nucleic acids depends upon its prior phosphorylation via thymidine kinase (TK; EC 2.7.1.21), cells deficient in TK activity are refractory to photoinduced killing. These observations suggested that combined treatment with BrdU, Hoechst 33258, and visible light would constitute an efficient selective strategy for the recovery of TK- mutant cells. In this report we describe a single-step selection protocol which reduced the survival of TK+ cells by a factor of 10(5) without affecting the viability of TK- mutants. This procedure was used to isolate H4IIEC3 -derived rat hepatoma cells deficient in TK activity. The properties of several TK- hepatoma clones are discussed.

Animals↗

PubMed1984

An isozyme-specific selective system for the recovery of mammalian cells deficient in hepatic alcohol dehydrogenase activity.

A selective system toxic towards mammalian cells expressing the liver-specific isozyme of alcohol dehydrogenase (L-ADH) has been developed. A number of alpha-unsaturated primary and secondary alcohols were assayed for their ability to serve as substrates for rat liver ADH and were screened for cytotoxicity towards L-ADH+ and L-ADH- cells. 1-Propen-3-ol and 1-penten-3-ol were identified as agents showing selective cytotoxicity. Reconstruction experiments demonstrated that 1-propen-3-ol at a concentration of 15 microM could be used to recover L-ADH- clones from mixed populations of L-ADH+ and L-ADH cells. Cells expressing the non-allelic S-ADH isozyme were not killed under these conditions. The selective system defined in this report is thus isozyme-specific.

1-Propanol↗

PubMed1984

A genetic analysis of extinction: trans-dominant loci regulate expression of liver-specific traits in hepatoma hybrid cells.

Extinction is an operational term that refers to the lack of expression of tissue-specific traits that is generally observed in hybrid cells formed by fusing dissimilar cell types. To define the genetic basis of this phenomenon, a series of rat hepatoma x mouse fibroblast hybrids has been isolated and characterized. We report here that the extinction of hepatic marker traits in these clones was strictly correlated with the retention of five particular fibroblast chromosomes (autosomes 8, 9, 10, 11, and 13). In order to dissect this correlation into its component parts, hepatoma microcell hybrids containing single, specific fibroblast chromosomes were constructed. Hepatoma clones retaining only fibroblast chromosome 11 were specifically extinguished for liver-specific tyrosine aminotransferase (TAT) expression, while expression of four other hepatic traits and of numerous constitutive markers was unaffected. Furthermore, removal of fibroblast chromosome 11 from the populations by back-selection resulted in reexpression of TAT activity to full parental levels. These data define and localize a genetic locus, tissue-specific extinguisher-1 (Tse-1), which regulates hepatic TAT expression in trans. We also provide evidence that human Tse-1 resides on the homologous chromosome (human chromosome 17), and that hybrids retaining active Tse-1 loci lack TAT-specific mRNA.

Animals↗