Transcriptional regulation of and by NFATc1 in Lymphocytes:
Gespeichert in:
| 1. Verfasser: | |
|---|---|
| Format: | Abschlussarbeit Buch |
| Sprache: | English |
| Veröffentlicht: |
2013
|
| Schlagworte: | |
| Online-Zugang: | Volltext http://d-nb.info/1044532459/34 Inhaltsverzeichnis |
| Beschreibung: | Zsfassung in dt. und engl. Sprache |
| Beschreibung: | 161 Blatt Ill., graph. Darst. |
| Format: | Langzeitarchivierung gewährleistet, LZA |
Internformat
MARC
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| 245 | 1 | 0 | |a Transcriptional regulation of and by NFATc1 in Lymphocytes |c Ronald Rudolf. Betreuer: E. Serfling |
| 246 | 1 | 3 | |a Transkriptionelle Regulation von und durch NFATc1 in Lymphozyten |
| 264 | 1 | |c 2013 | |
| 300 | |a 161 Blatt |b Ill., graph. Darst. | ||
| 336 | |b txt |2 rdacontent | ||
| 337 | |b n |2 rdamedia | ||
| 338 | |b nc |2 rdacarrier | ||
| 500 | |a Zsfassung in dt. und engl. Sprache | ||
| 502 | |a Würzburg, Univ., Diss., 2013 | ||
| 538 | |a Langzeitarchivierung gewährleistet, LZA | ||
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| 655 | 7 | |0 (DE-588)4113937-9 |a Hochschulschrift |2 gnd-content | |
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| 912 | |a ebook | ||
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Datensatz im Suchindex
| _version_ | 1804151768393711616 |
|---|---|
| adam_text | I. INTRODUCTION 6
1.1 THE ADAPTIVE IMMUNE SYSTEM 6
1.2 DEVELOPMENT AND FUNCTION OF T-LYMPHOCYTES 8
1.2.1 T-CELL DEVELOPMENT 8
1.2.2 T-CELL ACTIVATION 12
1.3 DEVELOPMENT AND FUNCTION OF B-LYMPHOCYTES 15
1.3.1 B-CELL DEVELOPMENT 15
1.3.2 B-CELL ACTIVATION AND FUNCTION 18
1.4 B-CELL LYMPHOMAS 20
1.4.1 B-CELL LYMPHOMA CLASSIFICATION 20
1.4.2 WEHI-231 B-LYMPHOMA CELLS 22
1.5 CELL DEATH 23
1.5.1 APOPTOSIS AND ACTIVATION INDUCED CELL DEATH 23
1.6 NFATC PROTEINS 25
1.6.1 NFAT ACTIVATION 26
1.6.2 THE NFAT FAMILY 29
1.6.3 NFATCL ISOFORMS 31
1.7 NFAT IN NON-LYMPHOID CELLS 33
1.7.1 NFAT IN BONE DEVELOPMENT 33
1.7.3 NFAT IN HEART-VALVE DEVELOPMENT 34
1.8 GOALS OF THE EXPERIMENTAL WORK 35
HTTP://D-NB.INFO/1047490951
CONTENT
II. MATERIAL AND METHODS 36
2.1 MATERIAL 36
2.1.1 MOUSE STRAINS 36
2.1.2 CELL CULTURE 37
2.1.3 BACTERIA 37
2.1.4 RETROVIRAL EXPRESSION VECTORS 39
2.1.5 PRIMERS 40
2.1.6 CHEMICALS 41
2.1.7 BUFFERS AND SOLUTIONS 43
2.1.8 ANTIBODIES 44
2.1.9 TECHNICAL EQUIPMENT 45
2.2 METHODS 46
2.2.1 DNA PURIFICATION 46
2.2.2 DNA QUANTIFICATION 46
2.2.3 DNA ELECTROPHORESIS 47
2.2.4 ISOLATION OF DNA FRAGMENTS 48
2.2.5 DNA CLONING 48
2.2.6 RETROVIRAL INFECTION OF CELL-LINES AND PRIMARY B-CELLS: 52
2.2.7 CHROMATIN IMMUNOPRECIPITATION 53
2.2.8 NEXT GENERATION SEQUENCING (PERFORMED BY TRON, TRANSLAT. ONCOLOGY,
UNIV.
MAINZ) 57
1
CONTENT
2.2.9 PROTEIN (CO-)IMMUNOPRECIPITATION (CO-IP) 58
2.2.10 FLOW CYTOMETRY 58
2.2.11 REAL-TIME PCR (QRT-PCR) 59
2.2.12 CONFOCAL-MICROSCOPY: 60
III. RESULTS 61
3.1 NFATCL INDUCTION IN LYMPHOCYTES 61
3.1.1 NFATCL INDUCTION IN PERIPHERAL T-CELLS 61
3.1.2 NFATCL INDUCTION IN PERIPHERAL B-CELLS 63
3.2 REGULATION OF NFATCL TRANSCRIPTION IN LYMPHOCYTES 66
3.2.1 CHARACTERIZATION OF TWO REGULATORY ELEMENTS WITHIN THE NFATCL
LOCUS 66
3.2.2 REGULATION OF NFATCL EXPRESSION BY THE TWO INTRONIC ELEMENTS EL
AND E2 IN PRIMARY
LYMPHOCYTES 68
3.2 NFATCL-AVIDIN FUSION PROTEINS 71
3.2.1 CLONING OF EXPRESSION PLASMID PEGZ-NFATCL/AA-AVI 71
3.2.2 CLONING OF EXPRESSION PLASMIDS PEGZ-NFATCL/PC-AVI AND
PEGZ-NFATCL/PC-
AVIFL 73
3.3 BACTERIAL ARTIFICIAL CHROMOSOME NFATCL EX.9-AVIDIN 77
3.3.1 CLONING OF THE SHUTTLE-VECTOR PLD53.SC2 NFATCL EX.9-AVIDIN 77
3.4 FUNCTIONAL ANALYSIS OF BIOTINYLATABLE NFATCL PROTEIN ISOFORMS 81
3.4.1 TRANSIENT EXPRESSION OF NFATCL-AVI PROTEINS IN HEK 293T-CELLS 81
3.4.2 RETROVIRAL INFECTION OF WEHI-231 B-LYMPHOMA CELLS 83
2
CONTENT
3.4.3 TRANSCRIPTION INDUCTION OF NFATCL/A- ANDNFATCL/P-ISOFORMS IN
WEHI-231 CELLS
85
3.4.4 EXPRESSION OF NFATCL/AA-AVI AND NFATCL/PC-AVI PROTEINS IN WEHI-231
CELLS
OVEREXPRESSING NFATCL/AA-AVI OR NFATCL/PC-AVI 87
3.4.5 RETROVIRAL INFECTIONS OF PRIMARY B-CELLS WITH PEGZ-NFATCL/AA-AVI
91
3.5 APOPTOSIS
AND CELL-DEATH
OF WEHI-231 CELLS
OVEREXPRESSING INDIVIDUAL NFATCL PROTEINS
92
3.5.1 BCR-TRIGGERED INDUCTION OF APOPTOSIS IN WEHI-231 CELLS 92
3.5.2 CELL-DEATH IN WEHI-231 B-LYMPHOMA CELLS 93
3.6 TRANSCRIPTOME SEQUENCING OF WEHI 231 CELLS OVEREXPRESSING
NFATCL/AA-AVI AND
NFATCL/PC-AVI 94
3.6.1 QUANTIFICATION OF NFATCL/A- AND NFATCL/P-ISOFORM TRANSCRIPT
EXPRESSION 94
3.6.2 TRANSCRIPTIONAL REGULATION BY NFATCL/AA AND NFATCL/PC IN WEHI-231
B-
LYMPHOMA CELLS /§7
3.6.3 GENE CLUSTERING OF TRANSCRIPTS FROM WEHI-231 CELLS OVEREXPRESSING
NFATCL/AA-
AVI OR NFATCL/PC-AVI 99
3.6.4 STIMULATION-INDEPENDENT GENE-INDUCTION AND REPRESSION IN WEHI-231
CELLS
OVEREXPRESSING NFATCL/AA OR NFATCL/PC 104
3.7 CHIP ASSAYS USING WEHI-231 CELLS OVERXPRESSING NFATCL/AA-AVI OR
NFATCL/PC-AVI
109
3.7.1 CHIP ASSAYS USING WEHI-231 CELLS OVEREXPRESSING NFATCL/AA-AVI,
NFATCL/PC-
AVI AND/OR BIRA 109
3.7 CHLP-SEQ 112
3.7.1 DNA EXTRACTION AND CHLP-SEQ 112
3
3.7.2 CHIP SEQUENCING (TRON) AND DATA ANALYSIS
IV. DISCUSSION
113
118
4.1 NFATCL INDUCTION IN LYMPHOCYTES 118
4.1.1 NFATCL INDUCTION IN PERIPHERAL T-CELLS 118
4.1.2 NFATCL INDUCTION IN PERIPHERAL B-CELLS 119
4.2 REGULATION OF NFATCL TRANSCRIPTION IN LYMPHOCYTES 120
4.2.1 CHARACTERIZATION OF TWO POTENTIAL REGULATORY ELEMENTS WITHIN THE
NFATCL LOCUS... 121
4.3 DESIGN AND CLONING OF BIOTINYLATABLE NFATCL-ISOFORMS 122
4.4 NFATCL EX.9-AVIDIN BAC CLONING 123
4.5 FUNCTIONAL ANALYSIS OF BIOTINYLATABLE NFATCL-ISOFORMS IN WEHI-231
B-CELLS 124
4.5.1 TRANSIENT EXPRESSION AND BIOTINYLATION OF AVIDIN-TAGGED
NFATCL-ISOFORMS IN
HEK293T-CELLS 124
4.5.2 EXPRESSION OF NFATCL AVIDIN-TAGGED ISOFORMS IN WEHI-231 B-LYMPHOMA
CELLS
125
4.6 APOPTOSIS AND CELL-DEATH OF WEHI-231 B-LYMPHOMA CELLS 128
4.7 TRANSCRIPTOME-SEQUENCING OF WEHI-231 CELLS OVEREXPRESSING NFATCL/AA
AND
NFATCL/PC 129
4.7.1 QUANTIFICATION OF NFATCL/A- AND NFATCL/(I-ISOFORM TRANSCRIPT
EXPRESSION 129
4.7.2 PROFILES OF TRANSCRIPTIONAL REGULATION OF WEHI-231 B-LYMPHOMA
CELLS
OVEREXPRESSING NFATCL/AA- AND NFATCL/PC 130
4.7.3 FUNCTIONAL ONTOLOGY: GENES REGULATED IN NFATCL/AA AND NFATCL/PC
OVEREXPRESSING CELLS 131
4.8 CHIP
ANALYSIS OF NFATCL/AA-AVI AND NFATCL/PC-AVI PROTEINS 136
CONTENT
4.8.1 NFATCL/AA-AVI, NFATCL/PC-AVI AND BIRA STREPTAVIDIN CHIP 136
4.8.2 PRDML ANDAICDA BINDING BY NFATCL/AA AND -P/C IN WEHI-231 CELLS 137
4.8.3 CHIP SEQUENCING 138
V. SUMMARY 140
VI. ABBREVIATIONS 143
VII. REFERENCES 147
VIII. PUBLICATIONS 159
IX. VITAE 160
X. ACKNOWLEDEGMENTS 161
5
|
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| author | Rudolf, Ronald 1981- |
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| ctrlnum | (OCoLC)869884426 (DE-599)DNB1044532459 |
| discipline | Biologie Medizin |
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| spelling | Rudolf, Ronald 1981- Verfasser (DE-588)1046848089 aut Transcriptional regulation of and by NFATc1 in Lymphocytes Ronald Rudolf. Betreuer: E. Serfling Transkriptionelle Regulation von und durch NFATc1 in Lymphozyten 2013 161 Blatt Ill., graph. Darst. txt rdacontent n rdamedia nc rdacarrier Zsfassung in dt. und engl. Sprache Würzburg, Univ., Diss., 2013 Langzeitarchivierung gewährleistet, LZA Transkriptionsfaktor (DE-588)4303350-7 gnd rswk-swf Genregulation (DE-588)4122166-7 gnd rswk-swf Lymphozyt (DE-588)4036762-9 gnd rswk-swf Maus (DE-588)4169148-9 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Lymphozyt (DE-588)4036762-9 s Transkriptionsfaktor (DE-588)4303350-7 s Genregulation (DE-588)4122166-7 s Maus (DE-588)4169148-9 s DE-604 Serfling, E. dgs Erscheint auch als Online-Ausgabe urn:nbn:de:bvb:20-opus-83993 https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-83993 Resolving-System kostenfrei Volltext http://d-nb.info/1044532459/34 Langzeitarchivierung Nationalbibliothek DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=027025296&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Rudolf, Ronald 1981- Transcriptional regulation of and by NFATc1 in Lymphocytes Transkriptionsfaktor (DE-588)4303350-7 gnd Genregulation (DE-588)4122166-7 gnd Lymphozyt (DE-588)4036762-9 gnd Maus (DE-588)4169148-9 gnd |
| subject_GND | (DE-588)4303350-7 (DE-588)4122166-7 (DE-588)4036762-9 (DE-588)4169148-9 (DE-588)4113937-9 |
| title | Transcriptional regulation of and by NFATc1 in Lymphocytes |
| title_alt | Transkriptionelle Regulation von und durch NFATc1 in Lymphozyten |
| title_auth | Transcriptional regulation of and by NFATc1 in Lymphocytes |
| title_exact_search | Transcriptional regulation of and by NFATc1 in Lymphocytes |
| title_full | Transcriptional regulation of and by NFATc1 in Lymphocytes Ronald Rudolf. Betreuer: E. Serfling |
| title_fullStr | Transcriptional regulation of and by NFATc1 in Lymphocytes Ronald Rudolf. Betreuer: E. Serfling |
| title_full_unstemmed | Transcriptional regulation of and by NFATc1 in Lymphocytes Ronald Rudolf. Betreuer: E. Serfling |
| title_short | Transcriptional regulation of and by NFATc1 in Lymphocytes |
| title_sort | transcriptional regulation of and by nfatc1 in lymphocytes |
| topic | Transkriptionsfaktor (DE-588)4303350-7 gnd Genregulation (DE-588)4122166-7 gnd Lymphozyt (DE-588)4036762-9 gnd Maus (DE-588)4169148-9 gnd |
| topic_facet | Transkriptionsfaktor Genregulation Lymphozyt Maus Hochschulschrift |
| url | https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-83993 http://d-nb.info/1044532459/34 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=027025296&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| work_keys_str_mv | AT rudolfronald transcriptionalregulationofandbynfatc1inlymphocytes AT serflinge transcriptionalregulationofandbynfatc1inlymphocytes AT rudolfronald transkriptionelleregulationvonunddurchnfatc1inlymphozyten AT serflinge transkriptionelleregulationvonunddurchnfatc1inlymphozyten |