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Query: UMLS:C0038187 (starvation)
 24,951 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cells often acquire resistance to the antiproliferative agents methotrexate (MTX) or N-phosphonacetyl-L-aspartate (PALA) through amplification of genes encoding the target enzymes dihydrofolate reductase or carbamylphosphate synthetase/aspartate transcarbamylase/dihydroorotase (CAD), respectively. We showed previously that Syrian hamster BHK cells resistant to selective concentrations of PALA (approximately 3 x ID50) arise at a rate of approximately 10(-4) per cell per generation and contain amplifications of the CAD gene as ladder-like structures on one of the two B9 chromosomes, where CAD is normally located. We now find that BHK cells resistant to high concentrations of PALA (approximately 15 x ID50) appear only after prior exposure to selective concentrations of PALA for approximately 72 h. Furthermore, in contrast to untreated cells, BHK cells pretreated with selective concentrations of MTX give colonies in high concentrations of PALA, and cells pretreated with selective concentrations of PALA give colonies in high concentrations of MTX or 5-fluorouracil. As judged by measuring numbers of cells and metaphase cell pairs, BHK cells do not arrest completely when starved for pyrimidine nucleotides by treatment with selective concentrations of PALA for up to 72 h. We propose that DNA damage, caused when cells fail to stop DNA synthesis promptly under conditions of dNTP starvation, stimulates amplification throughout the genome by mechanisms--such as bridge-breakage-fusion cycles--that are triggered by broken DNA. Amplified CAD genes were analyzed by fluorescence in situ hybridization both in cells where amplification was induced by PALA pretreatment and in cells in which the amplification occurred spontaneously, before selection with PALA. The ladder-like structures that result from bridge-breakage-fusion cycles were observed in both cases.
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PMID:Inefficient growth arrest in response to dNTP starvation stimulates gene amplification through bridge-breakage-fusion cycles. 886 64

The establishment of culture conditions suitable for inducing differentiation of Toxoplasma gondii tachyzoites into parasites resembling the latent bradyzoite form has opened this important developmental transition to experimental analysis. In order to develop a genetic marker suitable for positive and negative selection during parasite differentiation. the T. gondii HXGPRT gene was placed under control of 5' flanking sequences derived from two bradyzoite-specific genes: BAG1 and LDH2. Random transgene integration at undefined genomic loci resulted in modest regulation (approximately 5-6-fold induction) above relatively high background levels (approximately 4% of wild-type controls). Integration of transgenes at a defined genomic position was achieved by targeting the uracil phosphoribosyl transferase (UPRT) locus using flanking homologous sequences and fluorouracil selection. This strategy was found to provide the added advantage of enhancing bradyzoite induction frequencies under conditions of pyrimidine starvation (low CO2). Constructs integrated in the direction of normal UPRT transcription exhibited moderate levels of inducibility, but transgenes integrated in the opposite direction were dramatically induced under differentiation conditions: 50-100-fold above the very low levels observed in tachyzoites (< 1% control). Positive selection (using mycophenolic acid) was shown to inhibit tachyzoites but not bradyzoites, while negative selection (using 8-azahypoxanthine) inhibited bradyzoites only. Stage-specific regulation of the HXGPRT selectable marker should permit genetic selections for the identification of mutants in the bradyzoite differentiation process.
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PMID:Stage-specific expression of a selectable marker in Toxoplasma gondii permits selective inhibition of either tachyzoites or bradyzoites. 927 73

To understand the regulation and expression of pyrimidine biosynthesis in plants, we have examined the effect of the metabolic inhibitor 5-fluoroorotic acid (FOA) on uridine-5'-monophosphate synthase (UMPSase) expression in cell cultures of Nicotiana plumbaginifolia. UMPSase is the rate-limiting step of pyrimidine biosynthesis in plants. Addition of FOA causes an up-regulation of UMPSase enzyme activity in cell cultures after a lag phase of several days. Western-blot analysis demonstrated that the up-regulation in enzyme activity was caused by increased expression of the UMPSase protein. Northern-blot analysis demonstrated a higher level of UMPSase mRNA in the FOA-induced tissues than in control tissues. Run-on transcriptional assays showed that the UMPSase gene was transcriptionally activated after FOA treatment. The mechanism of toxicity of FOA is through thymine starvation. We found that addition of thymine abrogated the FOA-mediated up-regulation of UMPSase. In addition, methotrexate and aminopterin, which affect thymine levels by inhibiting dihydrofolate reductase, also up-regulate UMPSase in N. plumbaginifolia cells.
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PMID:Uridine 5'-Monophosphate Synthase Is Transcriptionally Regulated by Pyrimidine Levels in Nicotiana plumbaginifolia 949 Jul 73

Extracellular ATP, when added as a single dose at concentrations higher than 0.1 mM to the culture medium, was growth inhibitory or even cytotoxic for human epidermoid carcinoma cells (A431). Adenosine at the same concentrations was much less potent. The molecular mechanism underlying the inhibitory effect of extracellular ATP has been investigated. The cytostatic as well as the cytotoxic effects of ATP could be prevented by supplying uridine as a pyrimidine source and, alternatively, by simultaneous addition of dipyridamole, which inhibits the uptake of adenosine. The data suggest that the long-term production and continuous uptake of adenosine, which is enzymatically generated from the ATP in the medium, led to an intracellular nucleotide imbalance with pyrimidine starvation. This triggered suicidal processes ending up in apoptosis of the cells. The tumor cells have been adapted to extracellular ATP with the aim to obtain cells which are more resistant to ATP. Therefore, growing cells were periodically treated with extracellular ATP. These cells were characterized by an enlargement of cell size, a decreased proliferation rate, and a reduced but not abolished sensitivity to cytostatic and cytotoxic ATP doses. The calcium response of adapted cells was shortened. The nucleotide hydrolyzing ectoenzyme activities (ecto-ATPase, ecto-ADPase, ecto-AMPase, ecto-Ap4Aase) were simultaneously upregulated. All phenotypic alterations of the adapted cells disappeared after cultivation for several generations in the absence of extracellular ATP. Considering ATP as a potential chemotherapeutic agent the adaptive phenomena of treated cells might be important.
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PMID:Nucleotide metabolizing ectoenzymes are upregulated in A431 cells periodically treated with cytostatic ATP leading to partial resistance without preventing apoptosis. 973 53

Saccharomyces cerevisiae responds to pyrimidine starvation by increasing the expression of four URA genes, encoding the enzymes of de novo pyrimidine biosynthesis, three- to eightfold. The increase in gene expression is dependent on a transcriptional activator protein, Ppr1p. Here, we investigate the mechanism by which the transcriptional activity of Ppr1p responds to the level of pyrimidine biosynthetic intermediates. We find that purified Ppr1p is unable to promote activation of transcription in an in vitro system. Transcriptional activation by Ppr1p can be observed, however, if either dihydroorotic acid (DHO) or orotic acid (OA) is included in the transcription reactions. The transcriptional activation function and the DHO/OA-responsive element of Ppr1p localize to the carboxyl-terminal 134 amino acids of the protein. Thus, Ppr1p directly senses the level of early pyrimidine biosynthetic intermediates within the cell and activates the expression of genes encoding proteins required later in the pathway. These results are discussed in terms of (i) regulation of the pyrimidine biosynthetic pathway and (ii) a novel mechanism of regulating gene expression.
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PMID:Activation of transcription by metabolic intermediates of the pyrimidine biosynthetic pathway. 985 11

In UV-irradiated excision-proficient (uvr+) Escherichia coli, pre-induced by simultaneous pre-starvation for thymine (T) and amino acids (AAs), and/or a low UV pre-dose applied after prestarvation for AAs, pyrimidine dimer excision (PDE) is reduced without an adequate increase of UV sensitivity and UV mutagenesis. The unexcised lesions are tolerated by a putative repair pathway that is uvr dependent but does not involve excision. The process consists of PDE inhibition, which requires outer membrane protease OmpT, and subsequent pyrimidine dimer (PD) toleration, which may be mediated by interaction with a sister duplex using a number of SOS and stress-inducible proteins.
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PMID:A non-excision uvr-dependent DNA repair pathway of Escherichia coli (involvement of stress proteins). 986 97

The six biochemical steps of the de novo pyrimidine biosynthesis pathway are conserved in all known organisms. However, in animals and fungi, unlike prokaryotes, at least the first two activities are grouped on a multifunctional enzyme. Here, we report cloning, mapping and transcriptional characterization of some pyrimidine biosynthesis genes in the filamentous fungus Aspergillus nidulans. The first two steps of the pathway are performed by a multifunctional enzyme comprising the activities of carbamoyl phosphate synthetase (CPSase) and aspartate transcarbamylase (ATCase). This polypeptide is encoded by a 7 kbp cluster gene, pyrABCN, which has a high degree of nucleotide identity with the Ura2 gene in Saccharomyces cerevisiae. The enzyme of the third step, dihydroorotase (DHOase), is encoded by a separate locus, pyrD. However, the pyrABCN gene apparently contains an evolutionary remnant of a DHOase-encoding sequence, similarly to the Ura2 gene of Saccharomyces cerevisiae. The pyrABCN gene is transcribed as a single 7 kb mRNA species. The level of transcripts of pyrABCN, pyrD and, to a lesser degree, pyrF genes responds to the presence of exogenous pyrimidines and to the conditions of pyrimidine starvation. Derepression of pyrABCN and pyrD under pyrimidine starvation is noticeably enhanced in pyrE mutants that accumulate dihydroorotic acid. The pyrABCN gene maps to the distal portion of the right arm of the chromosome VIII, whereas the pyrD gene, in contrast to early genetic data, is closely linked to the brlA gene and located to the right of it. Our data on mitotic recombination should help to verify the genetic map of the chromosome VIII. Comparison of amino acid sequences of active dihydroorotases with related enzymes and with their non-functional homologues in yeast and Aspergillus indicates that the active dihydroorotases from fungi are more similar to ureases and enzymes of the pyrimidine degradation pathway. The 'silent' dihydroorotase domains of the multifunctional enzymes from fungi and active DHOase domains of the multifunctional enzymes in higher eukaryotes are more closely related to bacterial dehydroorotases.
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PMID:Structural and transcriptional analysis of the pyrABCN, pyrD and pyrF genes in Aspergillus nidulans and the evolutionary origin of fungal dihydroorotases. 1041 50

The molecular and biological consequences of UV-B radiation were investigated by studying five species of marine bacteria and one enteric bacterium. Laboratory cultures were exposed to an artificial UV-B source and subjected to various post-UV irradiation treatments. Significant differences in survival subsequent to UV-B radiation were observed among the isolates, as measured by culturable counts. UV-B-induced DNA photodamage was investigated by using a highly specific radioimmunoassay to measure cyclobutane pyrimidine dimers (CPDs). The CPDs determined following UV-B exposure were comparable for all of the organisms except Sphingomonas sp. strain RB2256, a facultatively oligotrophic ultramicrobacterium. This organism exhibited little DNA damage and a high level of UV-B resistance. Physiological conditioning by growth phase and starvation did not change the UV-B sensitivity of marine bacteria. The rates of photoreactivation following exposure to UV-B were investigated by using different light sources (UV-A and cool white light). The rates of photoreactivation were greatest during UV-A exposure, although diverse responses were observed. The differences in sensitivity to UV-B radiation between strains were reduced after photoreactivation. The survival and CPD data obtained for Vibrio natriegens when we used two UV-B exposure periods interrupted by a repair period (photoreactivation plus dark repair) suggested that photoadaptation could occur. Our results revealed that there are wide variations in marine bacteria in their responses to UV radiation and subsequent repair strategies, suggesting that UV-B radiation may affect the microbial community structure in surface water.
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PMID:Marine bacterial isolates display diverse responses to UV-B radiation. 1047 81

The regulation of the activity of purine transporters in two protozoan species, Crithidia fasciculata and Trypanosoma brucei brucei, was investigated in relation to purine availability and growth cycle. In C. fasciculata, two high-affinity purine nucleoside transporters were identified. The first, designated CfNT1, displayed a K(m) of 9.4 +/- 2.8 microM for adenosine and was inhibited by pyrimidine nucleosides as well as adenosine analogues; a second C. fasciculata nucleoside transporter (CfNT2) recognized inosine (K(m) = 0.38 +/- 0.06 microM) and guanosine but not adenosine. The activity of both transporters increased in cells at mid-logarithmic growth, as compared to cells in the stationary phase, and was also stimulated 5-15-fold following growth in purine-depleted medium. These increased rates were due to increased Vmax values (K(m) remained unchanged) and inhibited by cycloheximide (10 microM). In the procyclic forms of T. b. brucei, adenosine transport by the P1 transporter was upregulated by purine starvation but only after 48 h, whereas hypoxanthine transport was maximally increased after 24 h. The latter effect was due to the expression of an additional hypoxanthine transporter, H2, that is normally absent from procyclic forms of T. b. brucei and was characterised by its high affinity for hypoxanthine (K(m) approximately 0.2 microM) and its sensitivity to inhibition by guanosine. The activity of the H1 hypoxanthine transporter (K(m) approximately 10 microM) was unchanged. These results show that regulation of the capacity of the purine transporters is common in different protozoa, and that, in T. b. brucei, various purine transporters are under differential control.
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PMID:Differential regulation of nucleoside and nucleobase transporters in Crithidia fasciculata and Trypanosoma brucei brucei. 1074 14

The release of amino acids from their vacuolar store was studied in situ, i.e. in cells with selectively permeabilized plasma membrane and functionally intact vacuoles. As we previously described [Roos et al., J. Biol. Chem. 272 (1997) 15849-15855], this transport process is regulated by extravacuolar adenylates at their physiological concentrations. We now show, using our test object Penicillium cyclopium, that not only purine but also pyrimidine nucleotides are involved in the control of efflux of vacuolar phenylalanine. At 0.1 mM adenosine or guanosine phosphates inhibit, whereas cytidine or uridine phosphates stimulate the rate of efflux. At 1 mM the same nucleotides have no measurable impact on efflux but abolish the effects of other nucleotides present at 0.1 mM. This argues for at least two interacting binding sites with different nucleotide affinities. The minimum structural requirement for any of the observed effects is a non-cyclic ribonucleoside monophosphate. In intact cells, cytosolic concentrations of ATP (representing purine nucleotides) and CTP (representing pyrimidine nucleotides) are 1-2 mM and 0.05-0.2 mM, respectively. ATP is therefore assumed to dominate transport control and allow optimum efflux (and uptake) rates. Short-time starvation of carbon and nitrogen adjusts CTP and ATP at levels that cause declining efflux rates. During prolonged starvation both nucleotides fall below their transport-controlling concentrations and thus allow increasing rates of efflux from the still maintained vacuolar pool. Hence, efflux control under nutrient limitation includes an interplay of purine and pyrimidine nucleotides which precisely regulates the release of vacuolar amino acids and enables flexible adjustment to either amino acid saving or cell survival.
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PMID:Selective regulatory effects of purine and pyrimidine nucleotides on vacuolar transport of amino acids. 1099 56


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