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A Aguilera

Publications and source records attributed to A Aguilera.

At least 145 records · Page 8Linked to original sources
PubMed1988

Mitotic gene conversion of large DNA heterologies in Saccharomyces cerevisiae.

Gene conversion of large DNA heterologous fragments has been shown to take place efficiently in Saccharomyces cerevisiae. It has been found that a 2.6 kb LEU2 DNA fragment in a multicopy plasmid was replaced by a 3.1 kb PGI1 chromosomal DNA fragment, when both fragments were flanked by homologous DNA regions. Gene conversion was asymmetric in a total of 481 recombinants analyzed. In contrast, truncated PGI1 or LEU2 genes in multicopy plasmids, gave no recombinants that restored a complete plasmid copy of these genes in a total of 242 recombinants studied, confirming that a conversion tract is disrupted by a heterologous region. The asymmetry of the events detected suggest that gene conversion of large DNA heterologies involves a process whereby a gap first covers one heterologous fragment and then this is followed by new DNA synthesis using the other heterologous fragment as a template. Therefore, it is likely that large DNA heterologies are converted by a double-strand gap repair mechanism.

DNA, Fungal↗
PubMed1988

Genetic control of intrachromosomal recombination in Saccharomyces cerevisiae. I. Isolation and genetic characterization of hyper-recombination mutations.

Eight complementation groups have been defined for recessive mutations conferring an increased mitotic intrachromosomal recombination phenotype (hpr genes) in Saccharomyces cerevisiae. Some of the mutations preferentially increase intrachromosomal gene conversion (hpr4, hpr5 and hpr8) between repeated sequences, some increase loss of a marker between duplicated genes (hpr1 and hpr6), and some increase both types of events (hpr2, hpr3 and hpr7). New alleles of the CDC2 and CDC17 genes were recovered among these mutants. The mutants were also characterized for sensitivity to DNA damaging agents and for mutator activity. Among the more interesting mutants are hpr5, which shows a biased gene conversion in a leu2-112::URA3::leu2-k duplication; and hpr1, which has a much weaker effect on interchromosomal mitotic recombination than on intrachromosomal mitotic recombination. These analyses suggest that gene conversion and reciprocal exchange can be separated mutationally. Further studies are required to show whether different recombination pathways or different outcomes of the same recombination pathway are controlled by the genes identified in this study.

Cell Cycle↗
PubMed1987

Mutations suppressing the effects of a deletion of the phosphoglucose isomerase gene PGI1 in Saccharomyces cerevisiae.

A mutant with a deletion covering the phosphoglucose isomerase gene PGI1, allele pgil delta, can only grow on a medium containing fructose and low concentrations of glucose whereas growth is completely inhibited by glucose concentrations higher than 0.4%. This was used to select suppressor mutants restoring growth on synthetic media with 2% glucose as the sole carbon source. One complementation group, SPG1, was defined by recessive mutations. The ability to grow on glucose media was strictly dependent on functional mitochondria. The generation time of the selected mutants on YEP glucose was 6-8 h. No ethanol was formed from glucose and the levels of respiration were very high. These phenotypes were also observed in single pgil delta mutants when growing on fructose media supplemented with 0.4% glucose. The other glycolytic enzymes, the enzymes of the glucose-6-phosphate oxidation pathway as well as catabolite repression were normal in suppressed pgil delta mutants. The suppressor mutation alone caused no abnormal phenotype. The results suggest that the spg1 suppressor mutations allow S. cerevisiae pgil delta mutant strains to grow on glucose by using the Pentose-P cycle in combination with unusual strong respiration.

Chromosome Deletion↗
PubMed1986

Isolation and molecular analysis of the phosphoglucose isomerase structural gene of Saccharomyces cerevisiae.

The PGI1 gene of Saccharomyces cerevisiae coding for the glycolytic enzyme phosphoglucose isomerase has been cloned by complementation of a mutant strain (pgi1) with a strongly reduced phosphoglucose isomerase activity. A genomic library constructed in the yeast multicopy vector YEp13 (Nasmyth and Tatchell 1980) was used. Four plasmids containing an overlapping region of 4.1 kb were isolated and characterized by restriction endonuclease mapping. Southern analysis of genomic digests prepared with different restriction enzymes confirmed the same pattern for the chromosomal sequences. Transformants with the isolated plasmids had a phosphoglucose isomerase activity increased by a factor of 7. The cloned sequence hybridized to a constitutively synthesized 2.2 kb RNA in Northern analysis. The coding region includes a 2.05 kb EcoRI fragment common to all four inserts. A fragment including part of the PGI1 region was subcloned into vector YRp7 and used to induce integration at the PGI1 locus. Genetical and Southern analysis of stable transformants showed that single as well as tandem integration took place at this locus. This showed that the PGI1 gene had been isolated. Finally, and in contrast to the results of Kempe et al. (1974a, b) who reported three isoenzymes in yeasts, only one copy of the PGI1 gene per genome was found in several laboratory strains tested by Southern analysis.

Cloning, Molecular↗
PubMed1986

Ethanol-sensitive mutants of Saccharomyces cerevisiae.

Saccharomyces cerevisiae mutants unable to grow at ethanol concentrations at which the wild type strain S288C does grow, have been isolated. Some of them show additional phenotypic alterations in colony size, temperature sensitivity and viability in ethanol, which cosegregate with the growth sensitivity in ethanol. 21 selected monogenic ethanol-sensitive mutants define 20 complementation groups, denominated ETA1 to ETA20, which indicates that there is a high number of genes involved in the ethanol tolerance/sensitivity mechanism. Out of 21 selected monogenic mutants, 20 are not altered in the glycolytic pathway since, when maintained in glucose-supplemented medium, they can produce as much ethanol as the wild type and at about the same velocity. Nor do any of the mutants seem to be altered in the lipid biosynthetic pathway since, whether grown in the absence or in the presence of ethanol, their concentration of fatty acids and ergosterol is similar to that of the wild type under the same conditions. Therefore growth sensitivity to ethanol does not seem necessarily to be related to carbohydrate or lipid metabolism.

Crosses, Genetic↗
PubMed1986

Deletion of the phosphoglucose isomerase structural gene makes growth and sporulation glucose dependent in Saccharomyces cerevisiae.

The structural gene PGI1 coding for phosphoglucose isomerase was replaced by the LEU2 gene in the genome of Saccharomyces cerevisiae. Plasmids carrying the LEU2 gene between genomic regions flanking the PGI1 gene were constructed and used to transform a PGI1/pgi1 diploid strain. Stable transformants lacking the PGI1 allele were isolated. Southern analysis of their meiotic products showed that haploid strains with a deletion of 1.6 kb within the 2.2 kb PGI1 coding region were viable. Thus, the PGI1 gene is not essential in yeasts. However, unlike pgi1 mutants with residual phosphoglucose isomerase activity, no growth was detected in the pgi1 delta haploid strains when fructose was supplied as sole carbon source. The wild-type growth rate could be restored by adding 0.1% glucose to the medium. Furthermore, pgi1 mutants with residual enzymatic activity grew very slowly on fructose-supplemented media containing up to 2% glucose. Strains carrying the deletion allele, however, failed to grow at glucose concentrations higher than 0.5%. Also the pgi1 delta strains did not grow in glucose as sole carbon source. On the other hand pgi1 delta/pgi1 delta diploid strains did not sporulate on the usual acetate medium. This defect could be alleviated by the addition of 0.05% glucose to the sporulation medium. Under these conditions the pgi1 delta mutants sporulated with an efficiency of 25% compared with the wild type. These results suggest that the phosphoglucose isomerase reaction is the only step catalysing the interconversion of glucose-6-P and fructose-6-P, glucose-6-P is essential in yeasts, and the oxidation of glucose-6-P through the glucose-6-P dehydrogenase reaction is not sufficient to support growth in yeasts.

Chromosome Deletion↗
PubMed1985

Role of mitochondria in ethanol tolerance of Saccharomyces cerevisiae.

The presence of active mitochondria and oxidative metabolism is shown to be essential to maintain low inhibition levels by ethanol of the growth rate (mu), fermentation rate (nu) or respiration rate (rho) of Saccharomyces cerevisiae wild type strain S288C. Cells which have respiratory metabolism show Ki (ethanol inhibition constant) values for mu, nu and rho, higher (Ki greater than 1 M) than those of "petite" mutants or "grande" strains grown in anaerobiosis (Ki = 0.7 M). In addition, the relationship between mu or nu and ethanol concentration is linear in cells with respiratory metabolism and exponential in cells lacking respiration. When functional mitochondria are transferred to "petite" mutants, the resulting strain shows Ki values similar to those of the "grande" strain and the inhibition of mu and nu by increasing ethanol concentrations becomes linear.

Anaerobiosis↗
PubMed1983

Selection of wine yeasts for growth and fermentation in the presence of ethanol and sucrose.

To optimize the conversion of carbohydrates to ethanol, strains of several Saccharomyces species were examined for the ability to grow and ferment in a range of sucrose and ethanol concentrations. A total of 632 wine yeasts, most of them isolated from wineries in Andalusia and Extremadura, southwestern Spain, were subjected to screening and selection. Growth and fermentative capacity in different ethanol and sucrose concentrations varied from one strain to another. There was no correlation between growth and fermentative capacity. The best 35 strains grew in 15% ethanol and fermented in 18% ethanol. Ethanol accumulated, although at a reduced rate, after the cells stopped growing. Most yeast strains were highly fermentative in 50% sucrose. Some of them effectively utilized the carbohydrates of the culture, yielding final ethanol concentrations of > 14%. Of the 35 selected strains, 16 were promising for genetic analysis and breeding because of their capacity to sporulate. These strains were homothallic, and their spores were viable. The meiotic products analyzed so far were also homothallic.

Journal Article↗
PubMed1981

Laser flow cytofluorometric analysis of HTC cells synchronized with hydroxyurea, nocodazole and aphidicolin.

The technique of laser flow cytofluorometry has been used to monitor the arrival in G1 and the subsequent progression through the cell cycle of HTC cells accumulated in metaphase with colcemid alone or after treatment with hydroxyurea and Nocodazole. Under the experimental conditions used in this study, the latter procedure gives much better results, avoiding in particular the extensive formation of micronucleated cells. Aphidicolin, an inhibitor of DNA polymerase, in combination with Nocodazole, provides a useful method to tightly synchronize these cells at the G1/S border.

Animals↗
PubMed1976

Vaccinia virus: the possibilities of its oncogenicity in humans.

Contrary to the earlier generally accepted view that vaccinia virus replicates in the cytoplasm only of suitable target cells, steadily accumulating data show that the viral genome spends a limited period of time in the nucleus. This, together with the many cases where a close association has been suspected or established between skin cancer and vaccination, suggests that vaccinia virus may, under certain yet undefined physiological conditions, act as an oncogenic virus in humans.

Cell Nucleus↗
PubMed1976

Study of messenger-like RNA extracted from HeLa cells polysomes. II. Identification of the types of RNA eluted from methylated albumin kieselguhr columns.

Together with the elution pattern of pure messenger RNA molecules of various origin, the labelling kinetics of rapidly labelled heterogeneously sedimenting RNA (HSRNA) extracted from polysomes of HeLa cells have been studied by chromatogrphy on columns made of methylated bovine serum albumin adsorbed on kieselguhr. HSRNA is eluted within three peaks-IP, Q2P and TDP-following in that order the increase of NaC1 concentration in the eluting buffer. Besides peak TDP which results from an experimental artefact, our data suggest that the appearance of peaks IP and Q2P reflects the absence and presence respectively of polyadenylic acid stretches in these molecules. Within peak Q2P, the critical factor affecting the order of elution is the size of the polyadenylic acid stretch.

HeLa Cells↗
PubMedDate not supplied

Protein-losing enteropathy is associated with peritoneal functional abnormalities in peritoneal dialysis patients.

OBJECTIVE: To evaluate the relationship between acquired peritoneal transport disorders and the presence of protein-losing enteropathy (PLE), and their contribution to the protein malnutrition in peritoneal dialysis (PD) patients. PATIENTS AND METHODS: We studied 31 clinically stable PD patients that received a fat overload diet for 3 days. We measured intestinal absorption of fecal fat (normal < 6 g/24-hour stool) and nitrogen (normal < 2 g/24-hr stool), intestinal protein permeability [fecal clearance of alpha1-antitrypsin (Calpha1AT) (normal < 12 mL/24-hr stool)], and nutritional markers [normalized protein nitrogen appearance (nPNA), half-life medium-term proteins, and body mass index]. Peritoneal solute transport was measured by mass transfer coefficient (MTC), and water transport by peritoneal ultrafiltration (UF) capacity. To define protein maldigestion it was necessary to find high fecal nitrogen values with normal Calpha1AT; PLE was defined when both values were elevated. RESULTS: High fecal nitrogen (mean 2.1+/-1 g/24-hr stool) and fat (mean 5.8+/-3.6 g/24-hr stool) were found in 15 patients; 6 patients had high Calpha1AT levels (PLE). These 6 patients showed a worse nutritional status: lower albumin (3.57+/-0.57 g/dL vs 3.98+/-0.38 g/dL, p < 0.05) and transferrin (243+/-70 mg/dL vs 272+/-44.3 mg/dL, p < 0.05), as well as lower triglycerides (131.3+/-31.7 mg/dL vs 187+/-116 mg/dL, p< 0.05). Higher urea MTCs were found in 10 patients, normal in 7, and lower in 14. Higher creatinine MTCs were found in 8 patients, normal in 15, and lower in 8. Normal peritoneal UF capacity was found in 25 and lower in 6 patients. These 6 patients showed higher urea and creatinine MTCs and Calpha1AT. A positive linear correlation between Calpha1AT, urea MTC (r = 0.56, p < 0.01), and creatinine MTC (r = 0.46, p < 0.01) was found. A similar situation occurred between Calpha1AT, fecal fat (r = 0.45, p < 0.05), and fecal nitrogen (r = 0.43, p < 0.05). Thirteen patients with previous history of peritonitis showed higher Calpha1AT than those without peritonitis (10.2+/-8 mL/24-hr stool vs 5.2+/-4.4 mL/24-hr stool, p < 0.05). CONCLUSIONS: We confirm that protein and fat malabsorption, maldigestion, and PLE are present in some PD patients. Higher fecal Calpha1AT is associated with malnutrition and poorer showings of the viability markers of peritoneal membrane function.

Adult↗