Ubiquitin ligase activity of XIAP in cellular inflammatory response:
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| Format: | Abschlussarbeit Mikrofilm Buch |
| Sprache: | English |
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Köln
2016
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| Beschreibung: | 1 Mikrofiche (IV, 92 Seiten) Illustrationen, Diagramme |
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Datensatz im Suchindex
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| adam_text | Titel: Ubiquitin ligase activity of XIAP in cellular inflammatory response
Autor: Broxtermann, Pia Nora
Jahr: 2016
Table of contents
Abbreviations.......................................................................................................Ill
Abstract..................................................................................................................1
Zusammenfassung................................................................................................2
1 Introduction........................................................................................................4
1.1 Inhibitor of Apoptosis Proteins...................................................................................4
1.1.1 X-linked inhibitor of apoptosis.............................................................................6
1.2 lAPs in programmed cell death pathways..................................................................8
1.2.1 Different types of programmed cell death...........................................................8
1.2.2 lAPs as regulators of programmed cell death pathways...................................11
1.3 lAPs in cancer..........................................................................................................12
1.3.1 lAPs in migration and metastasis......................................................................13
1.3.2 Targeting lAPs in cancer...................................................................................14
1.4 lAPs: ubiquitin ligases regulating cell death and inflammation................................15
1.4.1 lAPs as ubiquitin ligases...................................................................................15
1.4.2 lAPs in inflammatory signalling.........................................................................17
1.5 Aim of the study.......................................................................................................20
2. Material and methods.....................................................................................21
2.1 Chemicals...............................................................................................................21
2.2 Antibodies................................................................................................................21
2.3 DNA Constructs.......................................................................................................22
2.4 Cell culture...............................................................................................................25
2.5 Transaction.............................................................................................................26
2.6 RNA interference.....................................................................................................26
2.7 NF-kB Luziferase Reporter Gene Assay..................................................................26
2.8 Enzyme-linked immunosorbent assay (ELISA)........................................................26
2.9 RNA isolation...........................................................................................................27
2.10 Reverse transcription.............................................................................................27
2.11 quantitative PGR (qPCR).......................................................................................27
2.12 PrimePCR™ Assay................................................................................................28
2.13 Sample preparation and Western blotting..............................................................28
2.14 Proteome Profiler™ Human Cytokine Array Kit, Panel A......................................30
2.15 Immunoprecipitation (IP)........................................................................................30
2.16 Immunofluorescence..............................................................................................30
2.17 Immunohistochemistry...........................................................................................31
2.18 Bacterial infection...................................................................................................32
2.19 Statistical analyses................................................................................................32
2.20 Mouse experiments................................................................................................32
2.20.1 Generation of HA-mXIAP knock-in Mice.........................................................32
2.20.2 Animal care.....................................................................................................33
2.20.3 DNA-isolation from tail cuts.............................................................................33
2.20.4 Extraction of proteins from mouse organs.......................................................33
2.21 Technical equipment and software........................................................................34
3 Results..............................................................................................................35
3.1 Elevated XIAP expression results in the formation of large cytoplasmic protein
complexes......................................................................................................................35
3.2 XIAP oligomerisation promotes auto-ubiquitination and complex formation............40
3.3 XIAP oligomerisation is not inhibited by cytoplasmic SMAC or SMAC mimetics and
does not interfere with XIAP-caspase 3 interaction.......................................................46
3.4 XIAP ubiquitin ligase activity leads to NF-kB activation and up-regulation of IL-8
transcription and secretion.............................................................................................49
3.5 IL-8 secretion is regulated by XIAP mediated RIPK2 ubiquitination........................53
3.6 NOD stimulation leads to oligomerisation of XIAP via the RING domain and
formation of cytoplasmic protein complexes..................................................................58
3.7 XIAP contributes to IL-8 secretion and neutrophil infiltration in Melanoma..............62
3.8 Generation and characterisation of XIAP knock in mouse.......................................63
4 Discussion........................................................................................................68
4.1 Enhanced XIAP expression results in large cytoplasmic complexes independently of
caspase binding ability...................................................................................................68
4.2 Oligomerisation of the RING domain enables activation of the E3 ubiquitin ligase of
XIAP followed by auto-ubiquitination.............................................................................70
4.3 Enhanced XIAP expression promotes inflammatory signalling and IL-8 secretion via
RIPK2 ubiquitination......................................................................................................71
4.4 Activation of NOD signalling leads to XIAP complex formation and IL-8 secretion
mediated by XIAP..........................................................................................................72
4.5 XIAP level impacts on IL-8 secretion in melanoma..................................................74
4.6 Generation and characterisation of a conditional HA-mXIAP knock in mouse line.. 76
4.7 Conclusion...............................................................................................................78
5 References........................................................................................................80
6 Appendix...........................................................................................................87
6.1 Bio-Rad PrimePCR™ Assay....................................................................................87
6.2 Proteome Profiler™ Human Cytokine Array Kit, Panel A........................................89
6.3 Vector maps.............................................................................................................90
6.4 XIAP Open reading frame........................................................................................91
6.5 Melanoma Patients..................................................................................................91
6.6 Erklarung..................................................................................................................92
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| spelling | Broxtermann, Pia Nora Verfasser (DE-588)1101961872 aut Ubiquitin ligase activity of XIAP in cellular inflammatory response vorgelegt von Pia Nora Broxtermann Köln 2016 1 Mikrofiche (IV, 92 Seiten) Illustrationen, 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/1108812074/04 Inhaltsverzeichnis HBZ Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=029230880&sequence=000002&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Broxtermann, Pia Nora Ubiquitin ligase activity of XIAP in cellular inflammatory response |
| subject_GND | (DE-588)4113937-9 |
| title | Ubiquitin ligase activity of XIAP in cellular inflammatory response |
| title_auth | Ubiquitin ligase activity of XIAP in cellular inflammatory response |
| title_exact_search | Ubiquitin ligase activity of XIAP in cellular inflammatory response |
| title_full | Ubiquitin ligase activity of XIAP in cellular inflammatory response vorgelegt von Pia Nora Broxtermann |
| title_fullStr | Ubiquitin ligase activity of XIAP in cellular inflammatory response vorgelegt von Pia Nora Broxtermann |
| title_full_unstemmed | Ubiquitin ligase activity of XIAP in cellular inflammatory response vorgelegt von Pia Nora Broxtermann |
| title_short | Ubiquitin ligase activity of XIAP in cellular inflammatory response |
| title_sort | ubiquitin ligase activity of xiap in cellular inflammatory response |
| topic_facet | Hochschulschrift |
| url | http://d-nb.info/1108812074/04 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=029230880&sequence=000002&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
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