Publications

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    ABSTRACT: In some RNA viruses (e.g. in brome mosaic virus, BMV), the same factor (intra- or intermolecular hybridization between viral RNA molecules) is capable of inducing two different processes: RNA silencing and RNA recombination. To determine whether there is some interplay between these two phenomena, we have examined if the BMV-based recombination vector containing a plant-genome-derived sequence can function as a gene-silencing vector. Surprisingly, we found that neither dsRNA forming during the replication of the BMV-based vector nor highly structured regions of its genome were effective RNAi triggers. Only mutants carrying a sequence complementary to the target mRNA functioned as gene silencing vectors and were steadily maintained in the infected plant. The constructs containing a sense sequence or inverted repeats did not induce gene silencing but instead were eliminated from the plant cells.
    Archives of Virology 11/2009; 155(2):169-79. · 2.03 Impact Factor
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    ABSTRACT: HYL1 is a nuclear protein involved in the processing of miRNAs but its exact function remains unknown. Arabidopsis thaliana hyl1 mutants exhibit hypersensitivity to ABA. We decided to answer the question whether ABA affects the HYL1 protein localization within the cell and show that it does not. We also studied the expression of HYL1 in different tissues and organs. In this paper we show for the first time the expression profile of the HYL1 protein using anti-HYL1 antibodies. The protein is present in seedlings and mature plants in all organs studied, with the highest amount in inflorescences. A. thaliana HYL1 protein has several repetitions of a 28-amino-acid sequence at the C-terminus that confer protein instability. Our bioinformatic analysis of HYL1 homologs in different Brassica species shows that this repetition is typical only for Arabidopsis. This may suggest a relatively late evolutionary acquisition of the C-terminal domain.
    Acta biochimica Polonica 09/2008; 55(3):517-24. · 1.19 Impact Factor
  • Lukasz Sobkowiak, Bogna Szarzyńska, Zofia Szweykowska-Kulińska
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    ABSTRACT: Micro RNAs are 21-24 nt long RNA molecules involved in the regulation of gene expression on mRNA level. Mature miRNA molecules associated with the RNA-induced silencing complex (RISC) guide specifically mRNA cleavage or inhibit the translation. In plants selective mRNA degradation in miRNA-dependent pathway is a widespread mechanism that occurs more frequently than translation inhibition. miRNA precursors, known as pri-miRNA, are processed in several steps to produce mature miRNA molecules. There are several plant proteins involved in miRNA processing: DCL1, HYL1, SE, HEN1 and HASTY. The role of these proteins is discussed.
    Postepy biochemii 02/2008; 54(3):308-16.
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    ABSTRACT: Previous studies using isozymes have established that the moss Plagiomnium curvatulum is an allopolyploid species that originated from hybridization and chromosome doubling of Plagiomnium elatum and Plagiomnium ellipticum. It has so far remained unknown which species is the donor of chloroplasts and mitochondria to P. curvatulum. Studies of organellar inheritance in bryophytes suggest that uniparental transmission was involved. Comparison of nucleotide sequence data of three chloroplast and three mitochondrial DNA markers from P. curvatulum and its parental species reveal that the chloroplast and mitochondrial sequences of P. curvatulum and P. ellipticum are identical and differ from homologous sequences of P. elatum. Seven samples from five different populations of P. curvatulum all gave the same result. Both mitochondria and chloroplasts of P. curvatulum were inherited from P. ellipticum. The nucleotide substitution rate calculated for homologous organellar nucleotide sequences is 4.57 times higher in chloroplasts than in mitochondria. Moreover, substitution rates differ considerably between chloroplast and mitochondrial homologous intron sequences when allopolyploid species and their parental haploids from different genera of bryophytes are compared.
    Taxon 01/2008; 57(1):145-152. · 2.78 Impact Factor
  • Agnieszka Dzikiewicz, Zofia Szweykowska-Kulińska
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    ABSTRACT: Nonsense-mediated mRNA decay (NMD) is a surveillance pathway that protects cells from potentially harmful effects of truncated proteins that would otherwise be translated from mRNAs bearing premature termination codons (PTC). NMD targets not only aberrant mRNAs possessing PTCs resulting from mutations, transcription errors or abnormal splicing but is also thought to be involved in regulating the expression of several wild-type transcripts (approximately 10% of the transcriptome). NMD is a conserved pathway present in all eukaryotic organisms studied so far (yeast, nematoda, flies, mammals and plants). The key players involved in NMD are also conserved among these species. However, details of NMD mechanism vary among eukaryotic organisms studied so far.
    Postepy biochemii 02/2006; 52(4):390-8.
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    ABSTRACT: All higher plant plastid genomes have six classes of tRNA genes containing introns. One of those is the tRNALysUUU gene, which encodes maturase protein. In the case of liverwort species from the genus Porella and mosses from the genus Plagiomnium, the maturase coding gene (matK) represents a truncated form of other plant matK genes: several subdomains of the reverse transcriptase-like domain and so-called domain X are not present in these ORFs. These ORFs probably represent pseudogenes of the matK gene. The analysis of codon usage within the matK gene revealed the presence of strong A/T pressure. The use of codons with the third letter being U or A varies from 71-93%. The comparison of maturase amino acid sequences at the family level shows a high identity between species. However, when liverwort and angiosperm maturase sequences are compared, the percentage of identity drops dramatically. The calculated values of the number of nucleotide substitutions vary considerably, even when liverwort species are compared pairwise. The phenetic tree of relationships between plant species on the basis of tRNALysUUU intron sequences concur with the generally accepted plant phylogeny.
    Cellular & Molecular Biology Letters 02/2004; 9(2):239-51. · 1.95 Impact Factor
  • Marta Sawczak, Zofia Szweykowska-Kulińska
    Postepy biochemii 02/2004; 50(1):69-81.
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    Zofia Szweykowska-Kulińska, Artur Jarmołowski, Marek Figlerowicz
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    ABSTRACT: Several years ago it was discovered that plant transformation with a transcribed sense transgene could shut down the expression of a homologous endogenous gene. Moreover, it was shown that the introduction into the cell of dsRNA (double-stranded RNA) containing nucleotide sequence complementary to an mRNA sequence causes selective degradation of the latter and thus silencing of a specific gene. This phenomenon, called RNA interference (RNAi) was demonstrated to be present in almost all eukaryotic organisms. RNAi is also capable of silencing transposons in germ line cells and fighting RNA virus infection. Enzymes involved in this process exhibit high homology across species. Some of these enzymes are involved in other cellular processes, for instance developmental timing, suggesting strong interconnections between RNAi and other metabolic pathways. RNAi is probably an ancient mechanism that evolved to protect eukaryotic cells against invasive forms of nucleic acids.
    Acta biochimica Polonica 02/2003; 50(1):217-29. · 1.19 Impact Factor
  • Andrzej Pacak, Zofia Szweykowska-Kulińska
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    ABSTRACT: Liverwort Pellia borealis is an allopolyploid species that originated after the hybridization and chromosome doubling of two cryptic species; Pellia epiphylla species N and Pellia epiphylla species S. A sequence comparison of chloroplast tRNAUCCGly, tRNAUUULys gene introns, the mitochondrial tRNAGCUSer gene intron, and the first intron of the coxIII gene in the case of three liverwort species studied revealed that the chloroplast and mitochondrial sequences are identical in P. borealis and P. epiphylla species N but different from homologous P. epiphylla species S sequences. Both mitochondria and chloroplasts of P. borealis were thus inherited from one parent- P. epiphylla species N. Studies on 14 different populations of P. borealis gave the same result. These are the first data on organellar transmission in liverworts, the earliest land plants. Moreover, we show that the intron sequences of some organellar genes, until now not used in any systematic studies, could be very good markers in studying taxonomic relationships in closely related species and reconstructing historical events.
    Journal of Molecular Evolution 02/2003; 56(1):11-7. · 2.15 Impact Factor
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    ABSTRACT: In order to establish the structure and sequence requirements for pseudouridine (Psi(35)) biosynthesis in Arabidopsis thaliana tRNA(Tyr) five mutants of nuclear pre-tRNA(Tyr) have been prepared and analyzed: DeltaI-tRNA(Tyr) transcript depleted of an intron, and 5UI, 7UI, 9UI and 12UI transcripts containing tracts of five, seven, nine and 12 U residues, respectively, instead of the wild type tRNA(Tyr) intron. The in vitro transcripts were incubated in a lupin seed extract containing Psi(35) synthase activity, and those containing an artificial intron composed of 12 or nine U residues turned out to be good substrates for Psi(35) synthase. The transcript with an intron composed of seven uridine residues was pseudouridylated up to 40%, whereas the remaining two were not pseudouridylated at all. The secondary structures of all transcripts were determined using enzymatic and chemical probes: S(1), V(1), T(1), A, P(1) and Pb(2+). All mutant pre-tRNAs show similar structural features: their anticodon arm contains a five base pair stem and a large loop which consists of five natural tRNA(Tyr) AC loop nucleotides to which five, seven, nine and 12 U residues are added. As the structure of the wild type pre-tRNA(Tyr) is different we propose that the role of its intron in the process of U(35) pseudouridylation is simply to expand the anticodon region to the required critical length.
    Biochimica et Biophysica Acta 04/2002; 1574(2):137-44. · 4.66 Impact Factor
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    ABSTRACT: The sequencing of several complete genomes and the development of a DNA microarray technology are among the most important achievements of molecular biology. They gave the proper grounds for the development of modern functional genomics. However, there is one additional condition which needs to be satisfied to truely enable the study of how a genome works: a suitable method of selectively inducing and silencing the expression of each individual gene. The methods used so far have usually only permitted the influencing of gene expression through genetic manipulations at the DNA level (genetically modified plants). The discovery of RNA interference (RNAi) opens up completely new possibilities of research on the functioning of particular plant genes, without the necessity of altering the genome structure. In this case, interference takes place at the transcript level. Thus, at any given moment during plant development, the expression of a specific gene (or several genes) can be inhibited, even if it is important for the survival of the organism under study. To this end, a double-stranded RNA inducing the RNAi phenomenon has to be delivered into the plant cell. Here we describe the construction of four brome mosaic virus-based vectors, which, as our preliminary data indicate, can be used to transfer RNA into barley cells.
    Cellular & Molecular Biology Letters 02/2002; 7(2A):511-22. · 1.95 Impact Factor
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    Zofia Szweykowska-Kulińska, Andrzej Pacak, Kamila Jankowiak
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    ABSTRACT: A survey of fully-sequenced chloroplast genomes revealed that in land plants there are six tRNA genes that have introns. Moreover, the length of a particular tRNA gene intron remains relatively stable across species. However, in algae, the presence of chloroplast tRNA genes containing introns is exceptional. A survey of mitochondrial plant genomes revealed intron-containing tRNA genes are rather rare features, with the exception of tRNASerGCU genes in liverworts and peat-mosses. We isolated and sequenced one mitochondrial and three chloroplast intron-containing tRNA genes and a fragment of the mitochondrial coxIII gene containing the first intron from the following liverwort species: Pellia borealis, Pellia epiphylla-species N, Pellia epiphylla-species S and Porella baueri, Porella cordaeana, Porella platyphylla. We showed that, as in the case of higher plants, the rate of nucleotide substitution is lower in the mitochondrial genome than in the chloroplast genome. Moreover, the comparison of intron nucleotide sequences enabled us to show that in the case of one allopolyploid species, Pellia borealis, organelles were transmitted from one parent species, Pellia epiphylla-species N. In the case of another allopolyploid species, Porella baueri, organelles were also inherited from one parent species, Porella cordaeana. Therefore, organellar inheritance in liverworts seems to be uniparental. It remains clear that analysis of carefully chosen chloroplast and mitochondrial DNA sequences allowed us to reconstruct historical events.
    Cellular & Molecular Biology Letters 02/2002; 7(2A):403-16. · 1.95 Impact Factor
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    P Fiedorow, I Odrzykoski, J Szweykowski, Z Szweykowska-Kulińska
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    ABSTRACT: Species of the genus Pellia are similar to such an extent that their proper recognition based on morphological and anatomical features is difficult. To solve this problem isozyme methods were developed. As a result of these studies new cryptic species were recognized and new hypotheses concerning phylogenetic relationship in Pellia were formed. To examine hypotheses derived from isozyme data we decided to study species of the genus Pellia at the DNA level. Total DNA from all Polish Pellia species was isolated. The taxonomic and phylogenetic relationships in Pellia were examined using intergenic spacer sequences between nuclear tRNA(Leu) genes organised in tandem arrays. PCR (polymerase chain reaction) amplification of tRNA(Leu) gene spacers produced fragments of different sizes in all species and their restriction analysis showed species-specific patterns. PCR products were cloned and sequenced. Nucleotide sequence data were used for phylogenetic reconstruction. Sequence analysis confirmed previous results based on isozyme studies that P. endiviifolia- species A and species B as well as P. epiphylla- species S and species N (having different isozyme multilocus genotypes) are separate sibling species. Our results also confirmed the allopolyploid character of the only polyploid species in Pellia, P. borealis which was formed by hybridization of P. epiphylla- species N and P. epiphylla- species S cryptic species.
    Gene 02/2001; 262(1-2):309-15. · 2.20 Impact Factor
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    J Maceluch, M Kmieciak, Z Szweykowska-Kulińska, A Jarmołowski
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    ABSTRACT: Rat Nap57 and its yeast homologue Cbf5p are pseudouridine synthases involved in rRNA biogenesis, localized in the nucleolus. These proteins, together with H/ACA class of snoRNAs compose snoRNP particles, in which snoRNA guides the synthase to direct site-specific pseudouridylation of rRNA. In this paper we present an Arabidopsis thaliana protein that is highly homologous to Cbf5p (72% identity and 85% homology) and NAP57 (67% identity and 81% homology). Moreover, the plant protein has conserved structural motifs that are characteristic features of pseudouridine synthases of the TruB class. We have named the cloned and characterized protein AtNAP57 (Arabidopsis thaliana homologue of NAP57). AtNAP57 is a 565 amino-acid protein and its calculated molecular mass is 63 kDa. The protein is encoded by a single copy gene located on chromosome 3 of the A. thaliana genome. Interestingly, the AtNAP57 gene does not contain any introns. Mutations in the human DKC1 gene encoding dyskerin (human homologue of yeast Cbf5p and rat NAP57) cause dyskeratosis congenita a rare inherited bone marrow failure syndrome characterized by abnormal skin pigmentation, nail dystrophy and mucosal leukoplakia.
    Acta biochimica Polonica 02/2001; 48(3):699-709. · 1.19 Impact Factor
  • J Pieńkowska, Z Szweykowska-Kulińska
    Postepy biochemii 02/2001; 47(3):232-42.
  • M Kmieciak, Z Szweykowska-Kulińska, A Jarmołowski
    Postepy biochemii 02/1999; 45(4):270-84.
  • P Fiedorow, Z Szweykowska-Kulińska
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    ABSTRACT: We have established a new type of genetic marker in Pellia liverworts that can be used to differentiate between closely related species within this genus. The Pellia intergenic sequences spanning the tandemly repeated nuclear tRNA(Leu) genes were amplified using the PCR technique and, in the case of Pellia neesiana, the tRNA(Leu) tandem region was cloned and sequenced. The length comparison of the intergenic regions allowed for the identification of a number of Pellia species. Additional experiments suggest that this marker can be used to differentiate other species of liverworts and mosses.
    Plant Molecular Biology 01/1999; 38(6):1257-61. · 3.52 Impact Factor
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    A Pacak, P Fiedorow, J Dabert, Z Szweykowska-Kulińska
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    ABSTRACT: The random amplified polymorphic DNA (RAPD) technique was used to study the genomic relationships of three liverworts from the genus Pellia: P.epiphylla, P. borealis and P. neesiana. Altogether 150 characters (150 DNA fragments obtained using PCR) were scored. These characters were used to create a matrix of pairwise distances between all the pairs of taxa. Both distance (UPGMA, Fitch-Margoliash and Neighbor-Joining) and binary character-state (Wagner and Camin-Sokal parsimony and compatibility) methods were applied for trees' construction. Our results strongly support distinction of the recently discovered sibling species of P.epiphylla - species N and P.epiphylla - species S, which have an allopatric distribution in Poland (N - North, S - South Poland). Moreover, our data also supports the hypothesis of a hybrid origin (alloploid) of the polyploid P. borealis from P. epiphylla-NxP.epiphylla-S. P.neesiana was excluded as a donor of either of the genomes of P. borealis.
    Genetica 11/1998; 104(2):179-87. · 1.68 Impact Factor
  • J Pieńkowska, Z Szweykowska-Kulińska
    Postepy biochemii 02/1998; 44(2):102-13.
  • J Pieńkowska, J Wrzesiński, Z Szweykowska-Kulińska
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    ABSTRACT: Plant cytoplasmic tyrosine tRNA was pseudouridylated at three different positions: 35, 39 and 55. These pseudouridines were introduced by three different enzymes--pseudouridine synthases. Variants of the Arabidopsis thaliana pre-tRNA(Tyr) were constructed that allow to monitor specifically pseudouridylation at different nucleotide positions. Using such RNAs to assay pseudouridine synthesis we have prepared an extract from Lupinus luteus cv. Ventus seeds containing activities of at least psi35 and psi55 synthases. This is the first report describing the preparation of the lupin seed extract that specifically modifies plant pre-tRNA(Tyr) transcribed by T7 RNA polymerase. U35 is converted to psi35 only in an intron-dependent manner, while pseudouridylation of U55 is insensitive to the presence or absence of an intron.
    Acta biochimica Polonica 02/1998; 45(3):745-54. · 1.19 Impact Factor

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