Chemical genetics: a dynamic tool to dissect jasmonic acid signaling in Arabidopsis thaliana
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| Format: | Abschlussarbeit Mikrofilm Buch |
| Sprache: | English |
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Köln
2016
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| Beschreibung: | 2 Mikrofiches (XI, 124 Seiten) Diagramme |
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Datensatz im Suchindex
| _version_ | 1804177443957768192 |
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| adam_text | Titel: Chemical genetics
Autor: Suliman, Mohamed Nady Sayed
Jahr: 2016
Contents
Abstract:................................I
Zusammenfassung............................III
Abriviations:................................IX
1. Introduction..................................1
1.1. Jasmonic acid QA] and its functional role in plants......................................................................1
1.2. JA biosynthesis..................................1
1.3. Metabolic modifications of JA.........................................................................................................2
1.3. Molecular mechanism of JA signaling............................................................................................4
1.3.1. Perception of the bioactive JA-Ile................................-...............................................................4
1.3.2. Repression of JA signaling via JAZs and other co-repressors....................................................4
1.3.3. Positive regulators of JA signaling...............................................................................................6
1.4. Chemical genetics and phytohormones.........................................................................................8
1.4.1. Phasel: primary screening and putative hit compounds identification................................9
1.4.2. Phase 2: Confirmation and validation of the putative hit compounds...................................10
1.4.2. Phase 3: Target identification strategies.................................................................................11
1.5. JA signaling: what is still missing?........................................................13
1.6. Aims of this work:..................................20
2. RESULTS................................................................................................................................................22
2.1. Establishing conditions for chemical screening..........................................................................22
2.4.1. Impact of compounds on 1A dependent responses................................................................25
2.4.2. Impact of the chosen hit compounds on JA-independent pathways......................................27
2.5. Characterization and target identification..................................................................................44
2.5.1. Characterization of BML-259...................................................................................................44
2.5.2. Characterization of nocodazole...............................................................................................56
2.5.3. Characterization of L.-cycloserine............................................................................................68
3. Discussion..............................................................................................................................................79
3.1. The significance of investigating JA signaling using chemical genetics.....................................79
3.2. Potential benefits of establishing a robust screening campaign................................................79
3.3. Chemical genetics aided identification of novel JA signaling modulators.................................81
3.4. The significance of the CDK inhibitor BML-259 as a JA signaling inhibitor..............................82
3.5. Characterizing BML-mediated effect as an inhibitor of JA signaling.........................................82
3.6. A CDK is likely a target of the selective JA signaling inhibitor BML-259...................................84
3.7. Understanding how BML-259 inhibits JA signaling....................................................................84
3.8. Identifying the target of BML-259 using alternative approaches..............................................86
3.9. Nocodazole encompasses a novel feature as a selective JA signaling inhibitor........................87
3.10. Dissecting nocodazole s mode of action as a JA signaling inhibitor........................................88
3.11. Using alternative tools to identify the target of ncodazole......................................................90
3.12. L-cycloserine selectively inhibits JA signaling..........................................................................91
3.13. How is L-cycloserine inhibiting JA signaling?...........................................................................92
3.14. Other ways to identify molecular target of L-cycloserine........................................................93
3.15. Concluding remarks...........................................................95
4. Materials.................................................97
4.1. Plant materials............................................97
4.2. Chemicals.......................................................................................................................................97
4.3. Media..............................................................................................................................................98
4.4. Buffers and solutions.....................................................................................................................98
5. Methods.................................................................................................................................................98
5.1. Maintenance and cultivation of Arabidopsis plants....................................................................98
5.2. Maintenance and cultivation of Arabidopsis plants in growth chamber..................................99
5.3. Chemical treatment of Arabidopsis plants..................................................................................99
5.4. Chemical screen and chemical treatments..................................................................................99
5.5. Biochemical methods....................................................................................................................99
5.5.1. Isolation of total protein extract from Arabidopsis.................................................................99
5.5.2. Denaturing SDS-polyacrylamide gel electrophoresis (SDS-PAGE).........................................100
5.5.3. Immunublot analysis..............................................................................................................100
5.6. Molecular biological methods.....................................................................................................100
5.6.1. RNA extraction.......................................................................................................................100
5.6.2. cDNA synthesis......................................................................................................................100
5.6.3. Quantitative real-time PCR (qRT-PCR)...................................................................................101
5.6.4. qRT-PCR program:..................................................................................................................101
5.7.1. Reporter assays......................................................................................................................102
5.8. Monitoring the oxidative burst...................................................................................................104
5.9. Quantification of anthocyanin....................................................................................................104
5.10. In vitro quantification of proteasome activity........................................................................104
5.11. Yeast two-hybrid assay.............................................................................................................105
5.12. Extraction and quantification of the jasmonates....................................................................105
References...............................................................................................................................................106
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| author | Suliman, Mohamed Nady Sayed 1983- |
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| discipline | Biologie |
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| language | English |
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| spelling | Suliman, Mohamed Nady Sayed 1983- Verfasser (DE-588)1114924415 aut Chemical genetics a dynamic tool to dissect jasmonic acid signaling in Arabidopsis thaliana vorgelegt von Mohamed Nady Sayed Suliman Köln 2016 2 Mikrofiches (XI, 124 Seiten) Diagramme txt rdacontent h rdamedia he rdacarrier 24x Dissertation Universität zu Köln 2016 (DE-588)4113937-9 Hochschulschrift gnd-content B:DE-101 application/pdf http://d-nb.info/1120962307/04 Inhaltsverzeichnis HBZ Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=029669412&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Suliman, Mohamed Nady Sayed 1983- Chemical genetics a dynamic tool to dissect jasmonic acid signaling in Arabidopsis thaliana |
| subject_GND | (DE-588)4113937-9 |
| title | Chemical genetics a dynamic tool to dissect jasmonic acid signaling in Arabidopsis thaliana |
| title_auth | Chemical genetics a dynamic tool to dissect jasmonic acid signaling in Arabidopsis thaliana |
| title_exact_search | Chemical genetics a dynamic tool to dissect jasmonic acid signaling in Arabidopsis thaliana |
| title_full | Chemical genetics a dynamic tool to dissect jasmonic acid signaling in Arabidopsis thaliana vorgelegt von Mohamed Nady Sayed Suliman |
| title_fullStr | Chemical genetics a dynamic tool to dissect jasmonic acid signaling in Arabidopsis thaliana vorgelegt von Mohamed Nady Sayed Suliman |
| title_full_unstemmed | Chemical genetics a dynamic tool to dissect jasmonic acid signaling in Arabidopsis thaliana vorgelegt von Mohamed Nady Sayed Suliman |
| title_short | Chemical genetics |
| title_sort | chemical genetics a dynamic tool to dissect jasmonic acid signaling in arabidopsis thaliana |
| title_sub | a dynamic tool to dissect jasmonic acid signaling in Arabidopsis thaliana |
| topic_facet | Hochschulschrift |
| url | http://d-nb.info/1120962307/04 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=029669412&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| work_keys_str_mv | AT sulimanmohamednadysayed chemicalgeneticsadynamictooltodissectjasmonicacidsignalinginarabidopsisthaliana |
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