Mismatch repair in the antimutator Escherichia coli mud

Antimutators are genetic mutants that produce mutations at reduced rates compared to the wild type strain. They are interesting because they may provide insights into the mechanisms by which spontaneous mutations occur. We have investigated a reported antimutator strain of Escherichia coli termed mud for its possible mechanism. The mud strain exhibits a decrease in both spontaneous mutagenesis and mutability with alkylated agents and base analogs. These types of DNA lesions are known to be the substrates for the E. coli methyl-directed mismatch repair encoded by the mutHLSU system. We investigated whether the putative antimutator effect results from the increased expression or activity of the mutHLSU system. To directly measure the mismatch repair capacity of mud cells, we have transfected them with phage lambda heteroduplexes and scored the fraction of mixed (unrepaired) infective centers. This transfection system has been used routinely to assay mismatch repair capacity in E. coli and other organisms. No difference between mud and wild type cells is observed. From the results of the experiments we conclude that the reported antimutator effect of mud does not result from enhanced mismatch repair capacity. This conclusion is consistent with recently published evidence that the mud effect does not represent a real antimutator effect, but is an artifact due to impaired growth of mud cells under certain selective conditions.     

Efficient Haploid Induction in Microspore Suspension Culture of Aesculus hippocastanum and Karyotype Analysis

Suspension culture was more efficient method for haploid production than anther culture. All analysed androgenic regenerants originating from embryogenic microspores in suspension culture of Aesculus hippocastanum L. had a haploid number of chromosomes (n=20), while 50 % of those derived from anther culture were diploids. This revised version was published online in July 2006 with corrections to the Cover Date.     

Changes in midgut and brain proteins in Morimus funereus larvae depending on nutritive substrate

The response of Morimus funereus larvae to total starvation and refeeding with qualitatively different nutritive substrates (artificial diets supplemented with yeast as a source of B complex vitamins or with a digestibility reducer-tannic acid) was examined in this paper. Refeeding resulted in a compensatory increase of larval growth. Feeding and refeeding with qualitatively different nutritive substrates affected both quality and quantity of midgut and brain proteins. The observed differences suggest the possible switching of enzyme isoforms in M. funereus midgut and changes in synthesis/secretion of neurohormones, depending on food presence and its nutritional value.     

Sexual dimorphism in life history plasticity in the gypsy moth (Lymantria dispar L.)

Sexual differences in reaction norms of life history traits (larval development time--LDT, pupal weight--PW and adult longevity--L) were investigated in the gypsy moth reared on young or old oak leaves during the first larval instar. Sexual dimorphism was revealed for genetic variation in reaction norms that was expressed only for LDT in males, and PW and L in females. Higher mean plasticity of longevity was found in males compared to females indicating that the sexes are exposed to divergent selective pressures. Greater dependence of males on energy resources (carbohydrates and lipids) may account for the observed differences.     

RecFOR function is required for DNA repair and recombination in a RecA loading-deficient recB mutant of Escherichia coli

The RecA loading activity of the RecBCD enzyme, together with its helicase and 5' --> 3' exonuclease activities, is essential for recombination in Escherichia coli. One particular mutant in the nuclease catalytic center of RecB, i.e., recB1080, produces an enzyme that does not have nuclease activity and is unable to load RecA protein onto single-stranded DNA. There are, however, previously published contradictory data on the recombination proficiency of this mutant. In a recF(-) background the recB1080 mutant is recombination deficient, whereas in a recF(+) genetic background it is recombination proficient. A possible explanation for these contrasting phenotypes may be that the RecFOR system promotes RecA-single-strand DNA filament formation and replaces the RecA loading defect of the RecB1080CD enzyme. We tested this hypothesis by using three in vivo assays. We compared the recombination proficiencies of recB1080, recO, recR, and recF single mutants and recB1080 recO, recB1080 recR, and recB1080 recF double mutants. We show that RecFOR functions rescue the repair and recombination deficiency of the recB1080 mutant and that RecA loading is independent of RecFOR in the recB1080 recD double mutant where this activity is provided by the RecB1080C(D(-)) enzyme. According to our results as well as previous data, three essential activities for the initiation of recombination in the recB1080 mutant are provided by different proteins, i.e., helicase activity by RecB1080CD, 5' --> 3' exonuclease by RecJ- and RecA-single-stranded DNA filament formation by RecFOR.     

The activity of antioxidant enzymes and the content of uncoupling protein-1 in the brown adipose tissue of hypothyroid rats: comparison with effects of iopanoic acid

The activity of antioxidant enzymes, copper-zinc superoxide dismutase (CuZnSOD), manganese superoxide dismutase (MnSOD) and catalase (CAT), as well as that of the mitochondrial FAD-dependent alpha-glycerophosphate dehydrogenase (alpha-GPD) in the rat interscapular brown adipose tissue (IBAT) were studied after the treatment with methimazole (MMI) for three weeks or with iopanoic acid (IOP) for five days. Besides, the mitochondrial concentration of uncoupling protein-1 (UCP-1) and the activity of catecholamine degrading enzyme monoamine oxidase (MAO) in the IBAT as well as the activity of the catecholamine synthesizing enzyme, dopamine beta-hydroxylase (DBH) in rat serum were examined. Judging by the significantly enhanced level of serum DBH, which is an index of sympathetic activity, and that of IBAT MAO, the increase in MnSOD and CAT activities in the IBAT of hypothyroid (MMI-treated) rats seems to be due to elevated activity of sympathetic nervous system (SNS). However, CuZnSOD activity is not affected by SNS. On the contrary, IOP, which is a potent inhibitor of T4 deiodination into T3 producing "local" hypothyroidism, did not change either SNS activity or activities of IBAT antioxidant enzyme. However, both treatments significantly decreased IBAT UCP-1 content and alpha-GPD activity suggesting that the optimal T3 concentration in the IBAT is necessary for maintaining basal levels of these key mitochondrial parameters.     

Ontogenetic profile of ecto-ATPase activity in rat hippocampal and caudate nucleus synaptic plasma membrane fractions

An ontogenetic study of ecto-ATPase activity and the content of enzyme proteins was assessed in the caudate nucleus and hippocampal synaptic plasma membranes isolated from rats at various ages (15, 30, 90, 180 and 365 days). The ontogenetic profile revealed that the enzyme activities in both brain areas were the highest on day 30 and 365, while the ecto-ATPase protein abundance was the highest on day 15 after birth. Possible explanation for obtained ontogenetic profile and the discrepancy between activity and abundance may reside in the fact that ecto-ATPase during development could exert additional roles other than those related to metabolism of ATP. It is likely that ecto-ATPase, regulating the concentration of ATP and adenosine in synaptic cleft, has important role in the processes of brain development and aging.     

Effects of estradiol benzoate and progesterone on superoxide dismutase activity in the thymus of rats

The activity of mitochondrial superoxide dismutase (MnSOD) and cytosol superoxide dismutase (CuZnSOD) was measured in corresponding subcellular fractions prepared from the thymi of intact and chronically gonadectomized (GX) rats of both sexes, as well as of GX male and female rats injected subcutaneously with a single dose of 5 microg estradiol benzoate (EB) and/or 2 mg progesterone (P). Animals were sacrificed 2 h or 24 h following hormone treatment. In the females, the activity of MnSOD in the thymus was stable during the estrous cycle and did not change after ovariectomy. Treatment of GX females with estradiol benzoate resulted 2 h later in a significant elevation of MnSOD activity, whereas 24 h later the activity returned back to control values. On the other hand, treatment of GX females with progesterone had no effect on the MnSOD activity. However, combined hormone treatment, in which EB injection preceded progesterone injection by one hour, enhanced the effect on MnSOD activity similar to that of estradiol benzoate alone. The activity of CuZnSOD in cycling rats was increased in proestrus, whereas removal of the ovaries kept the values at low diestrus and estrus levels. Contrary to MnSOD, CuZnSOD activity did not change after EB treatment of GX females, while progesterone increased the enzyme activity at 2 h and 24 h after hormone treatment. However, combined EB+P treatment proved to be ineffective. In the males, neither MnSOD nor CuZnSOD activity was affected by the removal of testes or by progesterone treatment of GX animals. Only EB injection to GX rats significantly increased CuZnSOD activity 24 h later.