Cudejková, Mária*
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BACKGROUND: When applied to a nutrition solution or agar media, the non-substituted aromatic cytokinins caused thickening and shortening of the primary root, had an inhibitory effect on lateral root branching, and even showed some negative effects on development of the aerial part at as low as a 10 nanomolar concentration. Novel analogues of aromatic cytokinins ranking among topolins substituted on N9-atom of adenine by tetrahydropyranyl or 4-chlorobutyl group have been prepared and tested in standardized cytokinin bioassays [1]. Those showing comparable activities with N(6)-benzylaminopurine were further tested in planta. METHODOLOGY/PRINCIPAL FINDINGS: The main aim of the study was to explain molecular mechanism of function of novel cytokinin derivatives on plant development. Precise quantification of cytokinin content and profiling of genes involved in cytokinin metabolism and perception in treated plants revealed several aspects of different action of m-methoxytopolin base and its substituted derivative on plant development. In contrast to standard cytokinins, N9- tetrahydropyranyl derivative of m-topolin and its methoxy-counterpart showed the negative effects on root development only at three orders of magnitude higher concentrations. Moreover, the methoxy-derivative demonstrates a positive effect on lateral root branching and leaf emerging in a nanomolar range of concentrations, in comparison with untreated plants. CONCLUSIONS/SIGNIFICANCE: Tetrahydropyranyl substitution at N9-position of cytokinin purine ring significantly enhances acropetal transport of a given cytokinins. Together with the methoxy-substitution, impedes accumulation of non-active cytokinin glucoside forms in roots, allows gradual release of the active base, and has a significant effect on the distribution and amount of endogenous isoprenoid cytokinins in different plant tissues. The utilization of novel aromatic cytokinin derivatives can distinctively improve expected hormonal effects in plant propagation techniques in the future.
- MeSH
- aktivace enzymů MeSH
- Arabidopsis účinky léků genetika růst a vývoj metabolismus MeSH
- biomasa MeSH
- cytokininy chemie farmakologie MeSH
- kořeny rostlin chemie účinky léků růst a vývoj MeSH
- kukuřice setá účinky léků genetika růst a vývoj metabolismus MeSH
- listy rostlin chemie účinky léků růst a vývoj MeSH
- oxidoreduktasy metabolismus MeSH
- proliferace buněk účinky léků MeSH
- regulace genové exprese u rostlin MeSH
- regulátory růstu rostlin chemie farmakologie MeSH
- semenáček účinky léků MeSH
- stanovení celkové genové exprese MeSH
- xylém chemie metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Recent technological advances have made next-generation sequencing (NGS) a popular and financially accessible technique allowing a broad range of analyses to be done simultaneously. A huge amount of newly generated NGS data, however, require advanced software support to help both in analyzing the data and biologically interpreting the results. In this article, we describe SATrans (Software for Annotation of Transcriptome), a software package providing fast and robust functional annotation of novel sequences obtained from transcriptome sequencing. Moreover, it performs advanced gene ontology analysis of differentially expressed genes, thereby helping to interpret biologically-and in a user-friendly form-the quantitative changes in gene expression. The software is freely available and provides the possibility to work with thousands of sequences using a standard personal computer or notebook running on the Linux operating system.
The fungus Claviceps purpurea is a biotrophic phytopathogen widely used in the pharmaceutical industry for its ability to produce ergot alkaloids (EAs). The fungus attacks unfertilized ovaries of grasses and forms sclerotia, which represent the only type of tissue where the synthesis of EAs occurs. The biosynthetic pathway of EAs has been extensively studied; however, little is known concerning its regulation. Here, we present the quantitative transcriptome analysis of the sclerotial and mycelial tissues providing a comprehensive view of transcriptional differences between the tissues that produce EAs and those that do not produce EAs and the pathogenic and non-pathogenic lifestyle. The results indicate metabolic changes coupled with sclerotial differentiation, which are likely needed as initiation factors for EA biosynthesis. One of the promising factors seems to be oxidative stress. Here, we focus on the identification of putative transcription factors and regulators involved in sclerotial differentiation, which might be involved in EA biosynthesis. To shed more light on the regulation of EA composition, whole transcriptome analysis of four industrial strains differing in their alkaloid spectra was performed. The results support the hypothesis proposing the composition of the amino acid pool in sclerotia to be an important factor regulating the final structure of the ergopeptines produced by Claviceps purpurea.
- MeSH
- biotechnologie MeSH
- Claviceps genetika metabolismus MeSH
- exprese genu MeSH
- fungální proteiny genetika metabolismus MeSH
- geny hub MeSH
- jednonukleotidový polymorfismus MeSH
- námelové alkaloidy biosyntéza MeSH
- oxidační stres MeSH
- peptidsynthasy genetika metabolismus MeSH
- průmyslová mikrobiologie MeSH
- sekvence aminokyselin MeSH
- sekvenční homologie aminokyselin MeSH
- stanovení celkové genové exprese MeSH
- Publikační typ
- časopisecké články MeSH
Pathogen-derived cytokinins (CKs) have been recognized as important virulence factor in several host-pathogen interactions and it was demonstrated multiple times that phytopathogenic fungi form CKs via the tRNA degradation pathway. In contrast to previous studies, the focus of this study is on the second step of CK formation and CK degradation to improve our understanding of the biosynthesis in fungi on the one hand, and to understand CK contribution to the infection process of Claviceps purpurea on the other hand. The ergot fungus Claviceps purpurea is a biotrophic phytopathogen with a broad host range including economically important crops causing harvest intoxication upon infection. Its infection process is restricted to unfertilized ovaries without causing macroscopic defense symptoms. Thus, sophisticated host manipulation strategies are implicated. The cytokinin (CK) plant hormones are known to regulate diverse plant cell processes, and several plant pathogens alter CK levels during infection. C. purpurea synthesizes CKs via two mechanisms, and fungus-derived CKs influence the host-pathogen interaction but not fungus itself. CK deficiency in fungi with impact on virulence has only been achieved to date by deletion of a tRNA-ipt gene that is also involved in a process of translation regulation. To obtain a better understanding of CK biosynthesis and CKs' contribution to the plant-fungus interaction, we applied multiple approaches to generate strains with altered or depleted CK content. The first approach is based on deletion of the two CK phosphoribohydrolase (LOG)-encoding genes, which are believed to be essential for the release of active CKs. Single and double deletion strains were able to produce all types of CKs. Apparently, log gene products are dispensable for the formation of CKs and so alternative activation pathways must be present. The CK biosynthesis pathway remains unaffected in the second approach, because it is based on heterologous overexpression of CK-degrading enzymes from maize (ZmCKX1). Zmckx1 overexpressing C. purpurea strains shows strong CKX activity and drastically reduced CK levels. The strains are impaired in virulence, which reinforces the assumption that fungal-derived CKs are crucial for full virulence. Taken together, this study comprises the first analysis of a log depletion mutant that proved the presence of alternative cytokinin activation pathways in fungi and showed that heterologous CKX expression is a suitable approach for CK level reduction.
Together with auxins, cytokinins are the main plant hormones involved in many different physiological processes. Given this knowledge, cytokinin levels can be manipulated by genetic modification in order to improve agronomic parameters of cereals in relation to, for example, morphology, yield, and tolerance to various stresses. The barley (Hordeum vulgare) cultivar Golden Promise was transformed using the cytokinin dehydrogenase 1 gene from Arabidopsis thaliana (AtCKX1) under the control of mild root-specific β-glucosidase promoter from maize. Increased cytokinin degradation activity was observed positively to affect the number and length of lateral roots. The impact on morphology depended upon the recombinant protein's subcellular compartmentation. While assumed cytosolic and vacuolar targeting of AtCKX1 had negligible effect on shoot growth, secretion of AtCKX1 protein to the apoplast had a negative effect on development of the aerial part and yield. Upon the application of severe drought stress, all transgenic genotypes maintained higher water content and showed better growth and yield parameters during revitalization. Higher tolerance to drought stress was most caused by altered root morphology resulting in better dehydration avoidance.
- MeSH
- aklimatizace genetika fyziologie MeSH
- biotechnologie MeSH
- cytokininy genetika metabolismus MeSH
- fenotyp MeSH
- fyziologický stres MeSH
- geneticky modifikované rostliny MeSH
- ječmen (rod) genetika růst a vývoj fyziologie MeSH
- kořeny rostlin fyziologie MeSH
- metabolické sítě a dráhy MeSH
- období sucha MeSH
- oxidoreduktasy genetika MeSH
- proteiny huseníčku genetika fyziologie MeSH
- regulátory růstu rostlin genetika metabolismus MeSH
- rekombinantní proteiny genetika metabolismus MeSH
- rostlinné geny MeSH
- rostlinné proteiny genetika MeSH
- stanovení celkové genové exprese MeSH
- upregulace MeSH
- Publikační typ
- časopisecké články MeSH
