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Computational prediction of non-enzymatic RNA degradation patterns

100%
Rybarczyk A., Jackowiak P., Figlerowicz M., Blazewicz J.
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2016
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vol. 63
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issue 4
745-751
EN
Since the beginning of the 21st century, an increasing interest in the research of ribonucleic acids has been observed in response to a surprising discovery of the role that RNA molecules play in the biological systems. It was demonstrated that they do not only take part in the protein synthesis (mRNA, rRNA, tRNA) but also are involved in the regulation of gene expression. Several classes of small regulatory RNAs have been discovered (e.g. microRNA, small interfering RNA, piwiRNA). Most of them are excised from specific double-stranded RNA precursors by enzymes that belong to the RNaseIII family (Drosha, Dicer or Dicer-like proteins). More recently, it has been shown that small regulatory RNAs are also generated as stable intermediates of RNA degradation (the so called RNA fragments originating from tRNA, snRNA, snoRNA etc.). Unfortunately, the mechanisms underlying biogenesis of the RNA fragments remain unclear. It is thought that several factors may be involved in the formation of the RNA fragments. The most important are the specific RNases, RNA-protein interactions and RNA structure. In this work, we focus on the RNA primary and secondary structures as factors influencing the RNA stability and consequently the pattern of RNA fragmentation. Earlier, we identified the major structural factors affecting non-enzymatic RNA degradation. Now, based on these data, we developed a new branch-and-cut algorithm that is able to predict the products of large RNA molecules' hydrolysis in vitro. We also present the experimental data that verify the results generated using this algorithm.
2
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AmiRNA Designer - new method of artificial miRNA design

88%
Mickiewicz A., Rybarczyk A., Sarzynska J., Figlerowicz M., Blazewicz J.
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2016
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vol. 63
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issue 1
71-77
EN
MicroRNAs (miRNAs) are small non-coding RNAs that have been found in most of the eukaryotic organisms. They are involved in the regulation of gene expression at the post-transcriptional level in a sequence specific manner. MiRNAs are produced from their precursors by Dicer-dependent small RNA biogenesis pathway. Involvement of miRNAs in a wide range of biological processes makes them excellent candidates for studying gene function or for therapeutic applications. For this purpose, different RNA-based gene silencing techniques have been developed. Artificially transformed miRNAs (amiRNAs) targeting one or several genes of interest represent one of such techniques being a potential tool in functional genomics. Here, we present a new approach to amiRNA*design, implemented as AmiRNA Designer software. Our method is based on the thermodynamic analysis of the native miRNA/miRNA* and miRNA/target duplexes. In contrast to the available automated tools, our program allows the user to perform analysis of natural miRNAs for the organism of interest and to create customized constraints for the design stage. It also provides filtering of the amiRNA candidates for the potential off-targets. AmiRNA Designer is freely available at http://www.cs.put.poznan.pl/arybarczyk/AmiRNA/.
3
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RNA-Seq-based analysis of differential gene expression associated with hepatitis C virus infection in a cell culture

64%
Hojka-Osinska A., Budzko L., Zmienko A., Rybarczyk A., Maillard P., Budkowska A., Figlerowicz M., Jackowiak P.
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2016
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vol. 63
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issue 4
789-798
EN
Hepatitis C virus (HCV) infection is one of the major causes of chronic liver diseases. Unfortunately, the mechanisms of HCV infection-induced liver injury and host-virus interactions are still not well recognized. To better understand these processes we determined the changes in the host gene expression that occur during HCV infection of Huh-7.5 cells. As a result, we identified genes that may contribute to the immune and metabolic cellular responses to infection. Pathway enrichment analysis indicated that HCV induced an increased expression of genes involved in mitogen-activated protein kinases signaling, adipocytokine signaling, cell cycle and nitrogen metabolism. In addition, the enrichment analyses of processes and molecular functions revealed that the up-regulated genes were mainly implicated in the negative regulation of phosphorylation. Construction of the pathway-gene-process network enabled exploration of a much more complex landscape of molecular interactions. Consequently, several essential processes altered by HCV infection were identified: negative regulation of cell cycle, response to endoplasmic reticulum stress, response to reactive oxygen species, toll-like receptor signaling and pattern recognition receptor signaling. The analyses of genes whose expression was decreased upon HCV infection showed that the latter were engaged in the metabolism of lipids and amino acids. Moreover, we observed disturbance in the cellular antiviral defense. Altogether, our results demonstrated that HCV infection elicits host response that includes a very wide range of cellular mechanisms. Our findings significantly broaden the understanding of complex processes that accompany HCV infection. Consequently, they may be used for developing new host-oriented therapeutic strategies.
4
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Translational and structural analysis of the shortest legume ENOD40 gene in Lupinus luteus

52%
Podkowinski J., Zmienko A., Florek B., Wojciechowski P., Rybarczyk A., Wrzesinski J., Ciesiolka J., Blazewicz J., Kondorosi A., Crespi M., Legocki A.
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issue 1
89-102
EN
Two early nodulin 40 (enod40) genes, ENOD40-1, the shortest legume ENOD40 gene, and ENOD40-2, were isolated from Lupinus luteus, a legume with indeterminate nodules. Both genes were expressed at similar levels during symbiosis with nitrogen-fixing bacteria. ENOD40 phylogeny clustered the L. luteus genes with legumes forming determinate nodules and revealed peptide similarities. The ENOD40-1 small ORF A fused to a reporter gene was efficiently expressed in plant cells, indicating that the start codon is recognized for translation. The ENOD40-1 RNA structure predicted based on Pb(II)-induced cleavage and modeling revealed four structurally conserved domains, an absence of domain 4 characteristic for legumes of indeterminate nodules, and interactions between the conserved region I and a region located upstream of domain 6. Domain 2 contains Mg(II) ion binding sites essential for organizing RNA secondary structure. The differences between L. luteus and Glycine max ENOD40 RNA models suggest the possibility of a switch between two structural states of ENOD40 transcript.
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