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The role of protein phosphorylation during progamic phase of tobacco male gametophyte development
Fíla, Jan ; Honys, David (advisor) ; Paleček, Jan (referee) ; Smýkal, Petr (referee)
v angličtině (English abstract) Tobacco male gametophyte has a strongly dehydrated cytoplasm and represents a metabolically inactive stage. Upon cytoplasm rehydration, pollen grain becomes metabolically active and after the activation is finished, the pollen tube growth through a selected pollen aperture starts. The rehydration together with metabolic activation are accompanied by the regulation of translation and post-translational modifications (mainly phosphorylation) of the existing proteins. In this Ph.D. thesis, there were identified phosphopeptides from tobacco (Nicotiana tabacum) mature pollen, pollen activated in vitro 5 min and pollen activated in vitro 30 min. The total proteins from the above male gametophyte stages were extracted. The protein extract was trypsinized and the acquired peptide mixture was enriched by MOAC (metal oxide/hydroxide affinity chromatography) with titanium dioxide matrix. The enriched fraction was subjected to liquid chromatography coupled with tandem mass spectrometry (LC- MS/MS). Totally, there were identified 471 phosphopeptides, carrying 432 exactly localized phosphorylation sites. The acquired peptide identifications were mapped to 301 phosphoproteins that were placed into 13 functional categories, dominant of which were transcription, protein synthesis,...
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The role of ribosomal proteins in the plant development
Jirásková, Veronika ; Raabe, Karel (advisor) ; Smýkal, Petr (referee)
The translation is one of the fundamental cell processes, in which the protein is synthesized according to the sequence of the mRNA molecule. The foremost recognized element of the translation machinery is the ribosome, a molecule complex composed by rRNAs and ribosomal proteins. In plants, ribosomal proteins are encoded by more than one gene, which may lead to sub- functionalization and neo-functionalization of ribosomal protein paralogs in plant development or in the reaction to the contemporary environment. Assembly of ribosomal subunits from different ribosomal protein paralogs could lead to functionally distinct pools of ribosomes with specialized role in the translation and its regulation in plants. The aim of this work is to review current data regarding the individual ribosomal proteins function within the plant growth and development. Keywords translation, translation regulation, ribosome, ribosomal proteins
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Dynamics of cold regulated proteins during cold acclimation in cereals
Vítámvás, Pavel ; Prášil, Ilja (advisor) ; Honys, David (referee) ; Smýkal, Petr (referee)
The aim of this dissertation was to study the mechanism of cold acclimation via the dynamics of cold regulated proteins (such as WCS120 or DHN5) in different frost- tolerant wheat and barley cultivars. Mass spectrometry analysis of a total sample of proteins, soluble upon boiling, showed qualitative differences between cold-acclimated (e.g., 7 COR proteins) and non-acclimated (e.g., only 3 COR proteins) samples of the winter wheat Mironovskaya 808. Furthermore, by 2-DE or W-blot analysis, there were found quantitative differences in the accumulation of WCS120 proteins between cultivars, grown under different time, photoperiod, and/or temperature conditions. The higher levels of WCS120 proteins are associated with higher frost tolerance of cultivars, grown under constant and low temperature. However, the dynamics of WCS120 proteins during long-term cold-acclimation, with periods of de-acclimation and re- acclimation, demonstrated that plants with the same level of frost tolerance could be distinguished by the level of accumulation of the WCS120 proteins. These results indicated that developmental genes influence the ability to re-accumulate WCS120 proteins by the partial vernalization of plants, while the ability to induce high frost tolerance was only influenced by the saturation of vernalization....
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Dynamics of cold regulated proteins during cold acclimation in cereals
Vítámvás, Pavel ; Prášil, Ilja (advisor) ; Honys, David (referee) ; Smýkal, Petr (referee)
The aim of this dissertation was to study the mechanism of cold acclimation via the dynamics of cold regulated proteins (such as WCS120 or DHN5) in different frost- tolerant wheat and barley cultivars. Mass spectrometry analysis of a total sample of proteins, soluble upon boiling, showed qualitative differences between cold-acclimated (e.g., 7 COR proteins) and non-acclimated (e.g., only 3 COR proteins) samples of the winter wheat Mironovskaya 808. Furthermore, by 2-DE or W-blot analysis, there were found quantitative differences in the accumulation of WCS120 proteins between cultivars, grown under different time, photoperiod, and/or temperature conditions. The higher levels of WCS120 proteins are associated with higher frost tolerance of cultivars, grown under constant and low temperature. However, the dynamics of WCS120 proteins during long-term cold-acclimation, with periods of de-acclimation and re- acclimation, demonstrated that plants with the same level of frost tolerance could be distinguished by the level of accumulation of the WCS120 proteins. These results indicated that developmental genes influence the ability to re-accumulate WCS120 proteins by the partial vernalization of plants, while the ability to induce high frost tolerance was only influenced by the saturation of vernalization....
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Identification and expression of genes engaged in flowering of a model plant Chenopodium rubrum.
Cháb, David ; Štorchová, Helena (advisor) ; Kovařík, Aleš (referee) ; Smýkal, Petr (referee)
Summary: Study presentedin this PhD thesis focusedon the molecularbasis of flowering inductionin a short-dayplantChenopodiumrubrum.We lookedfor respectivehomologsof CONSTANS (CO), FLOWERING LOCUS T (FZ) and LEAFT (LF"I) genes,which act as crucialregulatorsin thephotoperiod-dependentsignalpathvtayinArabidopsisthaliana. We identifiedtwoFT-like (FTL) genesCTFTL| aCTFTL2 ditreringin theirexpression pattemsin tefraploidC. rubrun. CTFTLI showedrhythmicexpressionpeakingin midday. ElevatedexpressionoÍCrFTL| wascorrelatedwithfloweringunderpermissivephotoperiodic treatnents,whereasit was not expressedat all undernon inductivephotoperiďic regimes' CrFTL2 showedconstitutiveexpression.CrFTL| verylikely playsa Íoleas a floral inducer, butthefunctionof CTFTL2remarnsunknown. Two CO-lil&e (COZ) genes CTCOLI a CTCOL2 identified inC. rubrum are altemativelysplicedandproducetwovariantsoftranscripts.Oneofthem wasstandardwith oneintronlocatedin conservativesite,theotheronehadanadditionalintronconespondingto the90bp regionofthe first exon.This typeofaltemativesplicinghasnotbeendescribedin other lnown CoL gercs. All forms of fanscripts show allmost identicď rhythmic Eanscriptionalpattsmspeakingat the end of the night and differ only in the level of individualmRNA. Light stronglyinhibitednanscriptionofboth CrCOtr genes. We have ďso...
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The role of protein phosphorylation during progamic phase of tobacco male gametophyte development
Fíla, Jan ; Honys, David (advisor) ; Paleček, Jan (referee) ; Smýkal, Petr (referee)
v angličtině (English abstract) Tobacco male gametophyte has a strongly dehydrated cytoplasm and represents a metabolically inactive stage. Upon cytoplasm rehydration, pollen grain becomes metabolically active and after the activation is finished, the pollen tube growth through a selected pollen aperture starts. The rehydration together with metabolic activation are accompanied by the regulation of translation and post-translational modifications (mainly phosphorylation) of the existing proteins. In this Ph.D. thesis, there were identified phosphopeptides from tobacco (Nicotiana tabacum) mature pollen, pollen activated in vitro 5 min and pollen activated in vitro 30 min. The total proteins from the above male gametophyte stages were extracted. The protein extract was trypsinized and the acquired peptide mixture was enriched by MOAC (metal oxide/hydroxide affinity chromatography) with titanium dioxide matrix. The enriched fraction was subjected to liquid chromatography coupled with tandem mass spectrometry (LC- MS/MS). Totally, there were identified 471 phosphopeptides, carrying 432 exactly localized phosphorylation sites. The acquired peptide identifications were mapped to 301 phosphoproteins that were placed into 13 functional categories, dominant of which were transcription, protein synthesis,...
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