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1

The minimal genome paradox

100%
Wegrzyn G.
Journal of Applied Genetics
|
2001
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vol. 42
|
issue 3
385-392
EN
The concept of a ?minimal genome? has appeared as an attempt to answer the question what the minimum number of genes or minimum amount of DNA to support life is. Since bacteria are cells bearing the smallest genomes, it has been generally accepted that the minimal genome must belong to a bacterial species. Currently the most popular chromosome in studies on a minimal genome belongs to Mycoplasma genitalium, a parasite bacterium whose total genetic material is as small as 580 kb. However, the problem is how we define life, and thus also a minimal genome. M. genitalium is a parasite and requires substances provided by its host. Therefore, if a genome of a parasite can be considered as a minimal genome, why not to consider genomes of bacteriophages? Going further, bacterial plasmids could be considered as minimal genomes. The smallest known DNA region playing the function of the origin of replication, which is sufficient for plasmid survival in natural habitats, is as short as 32 base pairs. However, such a small DNA molecule could not form a circular form and be replicated by cellular enzymes. These facts may lead to an ostensibly paradoxical conclusion that the size of a minimal genome is restricted by the physical size of a DNA molecule able to replicate rather, than by the amount of genetic information.
2

Mutagenic activity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine

64%
Ulanowska K., Wegrzyn G.
Journal of Applied Genetics
|
2006
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vol. 47
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issue 1
85-87
EN
MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) is a neurotoxin, which can damage dopaminergic neurons. It causes symptoms resembling those observed in patients suffering from Parkinson's disease, and hence this toxin is widely used in studies on animal models of this disorder. Mutagenicity of MPTP was also reported by some authors, but results obtained by others suggested that this compound is not mutagenic. Interestingly, those contrasting results were based on the same assay (the Ames test). Therefore, we aimed to test MPTP mutagenicity by employing a recently developed Vibrio harveyi assay, which was demonstrated previously to be more sensitive than the Ames test, at least for some mutagens. We found that MPTP showed a significant mutagenic activity. Moreover, MPTP mutagenicity was attenuated by methylxanthines, compounds that are known to form complexes with aromatic mutagens.
3

Is tRNA only a translation factor or also a regulator of other processes?

64%
Wegrzyn G., Wegrzyn A.
Journal of Applied Genetics
|
2008
|
vol. 49
|
issue 1
115-122
EN
Abstract. tRNA has been discovered as a factor playing a central role in the translation of genetic information (encoded in DNA and transcribed to mRNA) into amino acid sequences of proteins. However, subsequent studies led to the hypothesis that during evolution, tRNA originated in replication, not translation. Indeed, there are many examples of tRNA-like molecules playing roles in reactions other than translation, including replication of various replicons. In this review, we have focused on functions of tRNA molecules (not tRNA-like structures) outside of their direct roles in translation as factors for a passive transportation of amino acids into a ribosome and deciphering triplets of nucleotides in codons of mRNA. Interestingly, it appears that such tRNA-dependent reactions are effective only when tRNA is uncharged. The most spectacular examples come from bacterial cells and include induction of the stringent control, regulation of transcription of some operons, and control of replication of ColE1-type plasmids. Recent studies indicated that tRNA (not only pre-tRNA, shown previously to be capable of self-excision of intron sequences) can be responsible for specific cleavage of another transcript, a ColE1 plasmid-encoded RNA I, which is involved in the regulation of plasmid DNA replication initiation. If this reaction is not restricted to RNA I but represents a more general phenomenon, one might suspect a potential role for uncharged tRNA molecules in regulation of various processes, whose efficiency depends on tRNA-cleavable RNAs. This kind of regulation would provide a possibility for a cell to respond to different nutrition conditions resulting in different levels of tRNA aminoacylation.
4

Induction of light emission by luminescent bacteria treated with UV light and chemical mutagens

51%
Czyz A., Plata K., Wegrzyn G.
Journal of Applied Genetics
|
2002
|
vol. 43
|
issue 3
377-389
EN
Intensity of light emission by luminescent bacteria in response to UV irradiation and chemical mutagens was tested. We demonstrated that luminescence of six strains of marine bacteria (belonging to four species: Photobacterium leiognathi, P. phosphoreum, Vibrio fischeri and V. harveyi) is significantly increased by UV irradiation relatively shortly after dilution of cultures. Such a stimulation of luminescence was abolished in cells treated with chloramphenicol 15 min before UV irradiation, indicating that effective gene expression is necessary for UV-mediated induction of light emission. These results suggest that stimulation of luminescence in UV-irradiated bacterial cells may operate independently of the quorum sensing regulation. A significant induction of luminescence was also observed upon treatment of diluted cultures of all investigated strains with chemical mutagens: sodium azide (SA), 2-methoxy-6-chloro-9-(3-(2-chloroethyl)aminopropylamino)acridine ? 2HCl (ICR-191), 4-nitro-o-phenylenediamine (NPD), 4-nitroquinolone-N-oxide (NQNO), 2-aminofluorene (2-AF), and benzo[?]pyrene. These results support the proposal that genes involved in bioluminescence belong to the SOS regulon. The use of bacterial luminescence systems in assays for detection of mutagenic compounds is discussed in the light of this proposal.
5

Optimisation of the microbiological mutagenicity assay based on genetically modified Vibrio harveyi strains

51%
Podgorska B., Chec E., Ulanowska K., Wegrzyn G.
Journal of Applied Genetics
|
2005
|
vol. 46
|
issue 2
241-246
EN
Recently, we have developed a novel assay designed for detection of mutagenic pollution of the marine environment. This assay is based on the use of a series of genetically modified strains (named BB7, BB7M, BB7X and BB7XM) of a marine bacterium Vibrio harveyi. Sensitivity of the V. harveyi mutagenicity assay was found to be similar to, or even somewhat higher than, that of the commonly used Ames test. Subsequent studies indicated that this assay may be useful in assessment of mutagenic contamination of the marine environment. Nevertheless, we assumed that improvement of this assay is still possible, and thus we aimed to optimise its procedures. Here we present our research on the optimisation of the V. harveyi mutagenicity assay, which indicated that different tester strains used in this assay give the best results depending upon the experimental conditions employed. Incubation of bacteria in a buffer, rather than in a nutrient broth, containing a mutagen, increased the efficiency of the assay with BB7 and BB7M strains, but had a deleterious effect in the case of BB7X and BB7XM. The latter couple of strains revealed higher mutagenicity in the plate assay, as compared to the liquid medium assay. However, the opposite effect was observed for BB7 and BB7M. Low-dose (1 J m?2) UV irradiation, as well as 30 min incubation in 0.1 M CaCl2, had no significant effect on the efficiency of the assay when using BB7 and BB7M, whereas the number of mutagen-induced mutants of BB7X and BB7XM strains increased about two times under these conditions. Our previous experiments indicated that various tester strains revealed different sensitivity to particular mutagens. Thus, a series of strains should be used in the assay. Results presented in this report show that different conditions should be used for two pairs of the tester strains: BB7 and BB7M, and BB7X and BB7XM.
6

The use of the Vibrio harveyi luminescence mutagenicity assay as a rapid test for preliminary assessment of mutagenic pollution of marine sediments

51%
Podgorska B., Pazdro K., Wegrzyn G.
Journal of Applied Genetics
|
2007
|
vol. 48
|
issue 4
409-412
EN
Mutagenic pollution of the natural environment is currently one of the most serious environmental problems. It includes the pollution of marine sediments. Therefore, rapid detection of the presence of mutagens is an important issue. Recently, we have developed a novel microbiological assay for rapid assessment of mutagenicity of samples from the natural environment. This assay is based on bioluminescence of a mutant Vibrio harveyi strain, and was shown to be useful in testing samples of marine water and plant tissues. Here we demonstrate the usefulness of this assay in preliminary assessment of mutagenic pollution of marine sediments. Mutagenicity of environmental samples taken from the Baltic Sea, is documented and compared here with a commercially available standard sediment sample (IAEA 383), which contains known amounts of mutagenic compounds. The whole procedure, from obtaining a sample in the laboratory to getting final results, is very short (less than 4 h).
7

Substrate deprivation therapy: a new hope for patients suffering from neuronopathic forms of inherited lysosomal storage diseases

51%
Jakobkiewicz-Banecka J., Wegrzyn A., Wegrzyn G.
Journal of Applied Genetics
|
2007
|
vol. 48
|
issue 4
383-388
EN
Lysosomal storage diseases are a group of disorders caused by defects in enzymes responsible for degradation of particular compounds in lysosomes. In most cases, these diseases are fatal, and until recently no treatment was available. Introduction of enzyme replacement therapy was a breakthrough in the treatment of some of the diseases. However, while this therapy is effective in reduction of many somatic symptoms, its efficacy in the treatment of the central nervous system is negligible, if any, mainly because of problems with crossing the blood-brain-barrier by intravenously administered enzyme molecules. On the other hand, there are many lysosomal storage diseases in which the central nervous system is affected. Results of very recent studies indicate that in at least some cases, another type of therapy, called substrate deprivation therapy (or substrate reduction therapy) may be effective in the treatment of neuronopathic forms of lysosomal storage diseases. This therapy, based on inhibition of synthesis of the compounds that cannot be degraded in cells of the patients, has been shown to be effective in several animal models of various diseases, and recent reports demonstrate its efficacy in the treatment of patients suffering from Niemann-Pick C disease and Sanfilippo disease.
8

A modified procedure for quantitative analysis of mtDNA, detecting mtDNA depletion

51%
Weglewska A., Jakobkiewicz-Banecka J., Wegrzyn G.
|
EN
Quantitative analysis of mitochondrial DNA (mtDNA) is crucial for proper diagnosis of diseases that are caused by or associated with mtDNA depletion. However, such a quantitative characterization of mtDNA is not a simple procedure and requires several laboratory steps at which potential errors can accumulate. Here, we describe a modified procedure for quantitative human mtDNA analysis. The procedure is based on using two PCR-amplified, fluorescein-labeled DNA probes, complementary to mtDNA (detection probe) and chromosomal 18S rDNA (reference probe), both of similar length. Thus, equal amounts of these probes can be used and, contrary to previously published procedures, no mtDNA purification (apart from total DNA isolation) or 18S rDNA cloning is necessary for probe preparation. Two separate hybridizations (each with one probe) are suggested instead of one hybridization with both probes; this decreases background signals and enables adjustment of the strength of specific signals from both probes, which is useful in the subsequent densitometric analysis after superimposing of both pictures. Using different DNA amounts for reactions, we have proved that the procedure is quantitative in a broad range of sample DNA concentrations. Moreover, we were able to detect mtDNA depletion unambiguously in tissue samples from patients suffering from diseases caused by dysfunction of mtDNA.
9

Bacteriophage contamination: is there a simple method to reduce its deleterious effects in laboratory cultures and biotechnological factories?

39%
Los M., Czyz A., Sell E., Wegrzyn A., Neubauer P., Wegrzyn G.
Journal of Applied Genetics
|
2004
|
vol. 45
|
issue 1
111-120
EN
Infection of bacterial cultures by bacteriophages as well as prophage induction in the host cells are serious problems in both research and biotechnological laboratories. Generally, prevention strategies (like good laboratory/factory hygiene, sterilisation, decontamination and disinfection) are necessary to avoid bacteriophage contamination. However, it is well known that no matter how good the laboratory/factory practice and hygiene are, bacteriophage infections occur from time to time. The use of immunised or resistant bacterial strains against specific phages may be helpful, but properties of the genetically modified strains resistant to phages are often worse (from the point of view of a researcher or a biotechnological company) than those of the parental, phage-sensitive strains. In this article we review recent results that may provide a simple way to minimise deleterious effects of bacteriophage infection and prophage induction. It appears that low bacterial growth rates result in a significant inhibition of lytic development of various bacteriophages. Moreover, spontaneous prophage induction is less frequent in slowly growing bacteria.
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