Functional analysis of a novel DNA polymorphism of a tandem repeated sequence in the asparagine synthetase gene in acute lymphoblastic leukemia cells.
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Declined asparagine synthetase mRNA expression and enhanced sensitivity to asparaginase in HL-60 cells committed to monocytic differentiation.
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During 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced differentiation of human promyelocytic leukemia HL-60 cells toward maturing monocytes/macrophages, asparagine synthetase (ASNS) mRNA expression declined time and dose-dependently. The effect of TPA was inhibited by inhibitors for PKC and MEK 1/2, but not by those for JNK and p38 MAPK. Combination treatment with TPA and asparaginase synergistically enhanced the growth retardation accompanied by apoptotic cell death characterized by internucleosomal DNA fragmentation. These data suggest the possible involvement of MEK1/2 MAPK in the inhibitory effect of TPA on ASNS mRNA expression and that the induction of the down-regulation of ASNS (via MEK1/2 activation) may be a new strategy for the treatment of leukemia blast cells. (+info)
Dark-induced and organ-specific expression of two asparagine synthetase genes in Pisum sativum.
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Nucleotide sequence analysis of cDNAs for asparagine synthetase (AS) of Pisum sativum has uncovered two distinct AS mRNAs (AS1 and AS2) encoding polypeptides that are highly homologous to the human AS enzyme. The amino-terminal residues of both AS1 and AS2 polypeptides are identical to the glutamine-binding domain of the human AS enzyme, indicating that the full-length AS1 and AS2 cDNAs encode glutamine-dependent AS enzymes. Analysis of nuclear DNA shows that AS1 and AS2 are each encoded by single genes in P.sativum. Gene-specific Northern blot analysis reveals that dark treatment induces high-level accumulation of AS1 mRNA in leaves, while light treatment represses this effect as much as 30-fold. Moreover, the dark-induced accumulation of AS1 mRNA was shown to be a phytochrome-mediated response. Both AS1 and AS2 mRNAs also accumulate to high levels in cotyledons of germinating seedlings and in nitrogen-fixing root nodules. These patterns of AS gene expression correlate well with the physiological role of asparagine as a nitrogen transport amino acid during plant development. (+info)
A mammalian temperature-sensitive mutation affecting G1 progression results from a single amino acid substitution in asparagine synthetase.
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ts11 is a temperature-sensitive (ts) mutant isolated from the BHK-21 Syrian hamster cell line that is blocked in the G1 phase of the cell cycle at the non-permissive temperature (39.5 degrees C). We previously showed that the human gene encoding asparagine synthetase (AS) transformed ts11 cells to a ts+ phenotype and that ts11 cells were auxotrophic for asparagine at 39.5 degrees C. We show here that ts11 cells exhibit a ts phenotype for AS activity, and that the ts11 AS was much heat-labile than the wt enzyme. We have isolated AS cDNAs from wt BHK and ts11 cells and found that wt, but not ts11 AS cDNAs were capable of transformation. The deduced amino acid sequence of Syrian hamster AS showed 95% identity to the human protein as well as the same number of residues. The inability of the ts11 AS cDNAs to transform was due to a single base change, a C to T transition, that would result in the substitution of leucine with phenylalanine at a residue located in the C-terminal fourth of the enzyme. Thus the ts11 mutation identifies a mutated, thermolabile AS. (+info)
Nucleotide sequence of Escherichia coli asnB and deduced amino acid sequence of asparagine synthetase B.
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The Escherichia coli asparagine synthetase B gene (asnB) has been cloned into a temperature-sensitive, low copy plasmid, pOU71, as shown by the complementation of an E. coli asparagine auxotroph, E. coli JE6279. The nucleotide sequence of asnB and the flanking sequences were determined. The proposed coding region for the gene is 1662 nucleotides in length, and the deduced amino acid sequence of the coding region results in a protein that has a molecular weight of 62,666 and contains 554 amino acids. A promoter region is identified based on the transcription start site that was determined by primer extension experiments. Homology studies of the asnB protein sequence with the human asparagine synthetase and E. coli asparagine synthetase A protein show that there is a high degree of homology with only the human asparagine synthetase. A purF type glutamine amide transfer domain was identified upon inspection of the amino-terminal amino acid sequence of the asparagine synthetase B protein. (+info)