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Povzetek: In the field, plants are challenged by more than one biotic stressor at the same time. In this study, the molecular interactions between potato (Solanum tuberosum L.), Colorado potato beetle (Leptinotarsa decemlineata Say; CPB) and Potato virus YNTN (PVYNTN) were investigated through analyses of gene expression in the potato leaves and the gut of the CPB larvae, and of the release of potato volatile compounds. CPB larval growth was enhanced when reared on secondary PVYNTN-infected plants, which was linked to decreased accumulation of transcripts associated with the antinutritional properties of potato. In PVYNTN-infected plants, ethylene signalling pathway induction and induction of auxin response transcription factors were attenuated, while no differences were observed in jasmonic acid (JA) signalling pathway. Similarly to rearing on virus-infected plants, CPB larvae gained more weight when reared on plants silenced in JA receptor gene (coi1). Although herbivore-induced defence mechanism is regulated predominantly by JA, response in coi1-silenced plants only partially corresponded to the one observed in PVYNTN-infected plants, confirming the role of other plant hormones in modulating this response. The release of β-barbatene and benzyl alcohol was different in healthy and PVYNTN-infected plants before CPB larvae infestation, implicating the importance of PVYNTN infection in plant communication with its environment. This was reflected in gene expression profiles of neighbouring plants showing different degree of defence response. This study thus contributes to our understanding of plant responses in agro-ecosystems.
Ključne besede: insect midgut transcriptional response, gene expression, plant defence, volatile organic compounds, potato
Objavljeno v DiRROS: 02.08.2024; Ogledov: 87; Prenosov: 60
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Povzetek: Plant synthetic biology requires software tools to assist on the design of complex multi-genic expression plasmids. Here a vector design strategy to express genes in plants is formalized and implemented as a grammar in GenoCAD, a Computer-Aided Design software for synthetic biology. It includes a library of plant biological parts organized in structural categories and a set of rules describing how to assemble these parts into large constructs. Rules developed here are organized and divided into three main subsections according to the aim of the final construct: protein localization studies, promoter analysis and protein-protein interaction experiments. The GenoCAD plant grammar guides the user through the design while allowing users to customize vectors according to their needs. Therefore the plant grammar implemented in GenoCAD will help plant biologists take advantage of methods from synthetic biology to design expression vectors supporting their research projects.
Ključne besede: plasmid construction, plant genetics, gene expression
Objavljeno v DiRROS: 26.07.2024; Ogledov: 110; Prenosov: 69
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Povzetek: Background: Increased radioresistance due to previous irradiation or radiosensitivity due to human papilloma virus (HPV) infection can be observed in head and neck squamous cell carcinoma (HNSCC). The DNA-damage response of cells after exposure to DNA-damaging agents plays a crucial role in determining the fate of exposed cells. Tightly regulated and interconnected signaling networks are activated to detect, signal the presence of and repair the DNA damage. Novel therapies targeting the DNA-damage response are emerging; however, an improved understanding of the complex signaling networks involved in tumor radioresistance and radiosensitivity is needed. Materials and methods: In this study, we exposed isogenic human HNSCC cell lines with altered radiosensitivity to DNA-damaging agents: radiation, cisplatin and bleomycin. We investigated transcriptional alterations in the DNA-damage response by using a pathway-focused panel and reverse-transcription quantitative PCR. Results: In general, the isogenic cell lines with altered radiosensitivity significantly differed from one another in the expression of genes involved in the DNA-damage response. The radiosensitive (HPV-positive) cells showed overall decreases in the expression levels of the studied genes. In parental cells, upregulation of DNA-damage signaling and repair genes was observed following exposure to DNA-damaging agents, especially radiation. In contrast, radioresistant cells exhibited a distinct pattern of gene downregulation after exposure to cisplatin, whereas the levels in parental cells were unchanged. Exposure of radioresistant cells to bleomycin did not significantly affect the expression of DNA-damage signaling and repair genes. Conclusions: Our analysis identified several possible targets: NBN, XRCC3, ATR, GADD45A and XPA. These putative targets should be studied and potentially exploited for sensibilization to ionizing radiation and/or cisplatin in HNSCC. The use of predesigned panels of DNA-damage signaling and repair genes proved to offer a convenient and quick approach to identify possible therapeutic targets.
Ključne besede: DNA-damaging agents, gene expression, head and neck cancer, squamous cell carcinoma
Objavljeno v DiRROS: 24.07.2024; Ogledov: 119; Prenosov: 86
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Povzetek: The abscission of tomato leaves occurs in the petiole abscission zone, and its late stage includes two spatially divided processes: cell separation and programmed cell death (PCD). Both of these processes are regulated by ethylene. The last step in ethylene biosynthesis is conversion of 1-aminocyclopropane-1-carboxylic acid to ethylene, which is catalysed by the enzyme 1-aminocyclopropane-1-carboxylate oxidase (ACO); however, the location of ACO in the leaf petiole abscission zone is not known. The tomato gene LeLX encodes ribonuclease LX, which is a marker for PCD and is induced by ethylene during abscission, but its association with ACO has not been explored. In a tomato transgenic line 1-7 with inhibited expression of LeLX showing delayed leaf abscission, the morphology and ultrastructure of the leaf petiole abscission zone was examined. In this zone of the cv.’VF36’ and of a transgenic line 1-7, spatiotemporal differences in expression of LeACO1 and LeACO4 were analysed and ACO protein was detected immunohistochemically. In comparison to wild-type plants, there were no obvious morphological and ultrastructural features in the abscission zone of plants of a transgenic line 1-7 before and after abscission induction. LeACO1 expression was low before abscission induction, and increased 24 h after induction, although with no apparent spatial pattern. In contrast, LeACO4 was expressed before abscission induction, and its transcript level declined 24 h after induction on the distal side of the abscission zone fracture. In the LeLX-inhibited transgenic line, there were no significant differences in LeACO1 and LeACO4 expression in the petiole abscission zone, in comparison to wild-type plants. In addition, the ACO protein was immunolocalised to the vascular tissues that traverse the petiole abscission zone in plants of wild type and of a transgenic line 1-7; and additionally in the plane of future abscission zone fracture of transgenic-line plants. The results suggest temporal differential expression of the LeACO genes in tomato leaf petioles and vascular localisation of ACO1 protein. Additionally, the results indicate that expression of LeACO genes is not affected by suppression of the LeLX expression.
Ključne besede: abscission, 1-aminocyclopropane-1-carboxylate oxidase, ethylene, gene expression, localization, RNase LX
Objavljeno v DiRROS: 24.07.2024; Ogledov: 102; Prenosov: 51
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Povzetek: The use of omics is gaining importance in the field of nanoecotoxicology; an increasing number of studies are aiming to investigate the effects and modes of action of engineered nanomaterials (ENMs) in this way. However, a systematic synthesis of the outcome of such studies regarding common responses and toxicity pathways is currently lacking. We developed an R-scripted computational pipeline to perform reanalysis and functional analysis of relevant transcriptomic data sets using a common approach, independent from the ENM type, and across different organisms, including Arabidopsis thaliana, Caenorhabditis elegans, and Danio rerio. Using the pipeline that can semiautomatically process data from different microarray technologies, we were able to determine the most common molecular mechanisms of nanotoxicity across extremely variable data sets. As expected, we found known mechanisms, such as interference with energy generation, oxidative stress, disruption of DNA synthesis, and activation of DNA-repair but also discovered that some less-described molecular responses to ENMs, such as DNA/RNA methylation, protein folding, and interference with neurological functions, are present across the different studies. Results were visualized in radar charts to assess toxicological response patterns allowing the comparison of different organisms and ENM types. This can be helpful to retrieve ENM-related hazard information and thus fill knowledge gaps in a comprehensive way in regard to the molecular underpinnings and mechanistic understanding of nanotoxicity.
Ključne besede: gene expression, nanomaterials
Objavljeno v DiRROS: 19.07.2024; Ogledov: 79; Prenosov: 103
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