Jadwiga Litwińska

Is hydroxyl radical generated by the Fenton reaction in vivo

Yeast mutants deficient in activities of cytosolic superoxide dismutase and catalase A and T were exposed to four different kinds of oxygen stress. The response of the cells contradicts suggestions, that hydroxyl radical is formed in vivo through the Fenton reaction. The results suggest that superoxide radicals are directly responsible for cytotoxic effects of oxygen.

Superoxide dismutase deficiency and the toxicity of the products of autooxidation of polyunsaturated fatty acids in yeast

Mutants of Saccharomyces cerevisiae, deficient in cytosolic superoxide dismutase and catalase activities were used to study the role of various oxygen species in the process of lipid peroxidation in yeast cells. Lipid peroxidation does not occur normally in yeast, because this organism is unable to form fatty acids with more than one double bond, whereas under physiological conditions, only fatty acids with at least two double bonds undergo this process. The fatty acid content of cellular lipids was modified by growing the cells in anoxia in the presence of oleic or linolenic acid. Toxic effects of oxygen were observed almost exclusively in those cells of yeast mutants deficient in superoxide dismutase, which contain linolenic acid in cellular lipids. Hypersensitivity of the mutant cells, however, results mainly from toxic effects of the products of autooxidation of extracellular fatty acids. These facts suggest that superoxide dismutases are in some way involved in preventing toxic effects of the products of lipid peroxidation and to some extent prevent the process of lipid peroxidation.

Haemoprotein formation in yeast

SummaryA procedure was described for the isolation of mutants affected in the regulation of catalase activity. Two such mutants, cgr 1 and cgr 2 were obtained. Both of them show catalase activity that is resistant to repression by glucose, but is sensitive to anoxia to the same extent as the wild type.

Analysis of heme biosynthesis in catalase and cytochrome deficient yeast mutants.

SummaryMutants of Saccharomyces cerevisiae, described as catalase and cytochromes deficient (Pachecka et al., 1974), have been analyzed for heme biosynthesis ability. Some enzymatic activities involved in protoheme synthesis were measured in acellular extracts, whereas whole cells were analyzed for cytochrome spectra and for possible accumulation of porphyrin synthesis intermediates. A good correlation was found between these in vitro and in vivo studies. Results show that two mutants were impaired in 5-aminolevulinate synthesis, two mutants were devoid of uroporphyrinogen I synthetase activity and one mutant presented defects in coproporphyrinogen III oxidase activity.

Isolation and characterization of lactose non-utilizing mutants inAspergillus nidulans

SummaryA number ofAspergillus nidulans mutants unable to grow on lactose or growing very poorly on this sugar have been isolated. They may be divided into two major groups: to the first belong mutants in which β-galactosidase can be induced by galactose but not by lactose. Mutants of the second group are induced neither by lactose nor by galactose. Mutants of the first group showed an impaired lactose-permease system, while those of the second group most likely concern β-galactosidase structural or regulatory genes as they show a normal rate of lactose uptake. Genetic analysis revealed that mutants from the first group fall into three different loci and those from the second into four loci. No mutant has been found so far with the lactose-permease system and β-galactosidase simultaneously impaired, or with a constitutive level of either activity.The wild-type strain ofAspergillus nidulans grows on lactose as the sole carbon source. The two enzymes necessary for the utilization of lactose, that is lactose permease (which is likely to be a complex system) and β-galactosidase show an inductive response to lactose and galactose (Paszewskiet al., 1970). Mycelia grown on glucose show a low level of permease activity which rises 7–10-fold upon induction by lactose, and no activity of β-galactosidase. Induction of both enzymes is not time-coordinated — the induction of permease preceeds the induction of β-galactosidase. In contrast toNeurospora crassa (Bates and Woodward, 1964; Bateset al., 1967; Lester and Byers, 1965) only one type of β-galactosidase with pH optimum 7.5–7.6 was found inAspergillus nidulans.A number of mutants unable to grow on lactose or growing very poorly on this sugar have been isolated. Their genetic and enzymatic characterization is given in this paper.

Hemoproteid Formation in Yeast

SummaryA method of hemoproteid-deficient mutants isolation is described. 13 mutants lacking catalase belonging to 9 complementation groups were isolated. Mutants representing 7 loci were also unable to form cytochromes.Addition of δ-aminolevulinic acid does not restore catalase activity. Normal mendelian segregation in crosses with wild strain was observed only in three mutants. The rest give 4:0 segregation. After ethidium bromide treatment and induction of rho — strains normal segregation was observed in all mutants. The masking effect of normal mitochondrial activity on gene expression in mutant strains is discussed.

Effect of 5-aminolevulinic acid synthesis deficiency on expression of other enzymes of heme pathway in yeast.

Abstract Some enzymes of the heme pathway have been analysed in the nuclear mutant strains cat11 ( hemA-2 ) and cat7 ( hemA-1 ) of Saccharomyces cerevisiae , both in the rho + and rho − states, and in the mutant ole3 ( ole3 , rho + ) already described. The mutation leading to the absence of 5-aminolevulinic acid (ALA) synthase activity and to cytochrome deficiency was not expressed in cat11 rho + , and only partly in cat7 rho + . The rho − mutation unmasked the defects associated with the hemA mutations. When expressed, in ole3, cat11 rho − and cat7 rho − , the absence of ALA (and heme) synthesis led to a total loss of uroporphyrinogen I synthase activity and to an important increase of coproporphyrinogen III oxidase activity. Genetic analysis showed that hemA-1 and ole3 are two different unlinked loci.