Activation of the tumor suppressor merlin by myosin, moesin phosphatase
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| Format: | Abschlussarbeit Buch |
| Sprache: | English |
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Karlsruhe
FZKA
2004
|
| Ausgabe: | Als Ms. gedr. |
| Schriftenreihe: | Wissenschaftliche Berichte / Forschungszentrum Karlsruhe Technik und Umwelt
7017 |
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| 100 | 1 | |a Jin, Hongchuan |d 1974- |e Verfasser |0 (DE-588)129756334 |4 aut | |
| 245 | 1 | 0 | |a Activation of the tumor suppressor merlin by myosin, moesin phosphatase |c Hongchuan Jin. Forschungszentrum Karlsruhe GmbH, Karlsruhe |
| 246 | 1 | 3 | |a FZKA-7017 |
| 250 | |a Als Ms. gedr. | ||
| 264 | 1 | |a Karlsruhe |b FZKA |c 2004 | |
| 300 | |a XI, 105 S. |b Ill., graph. Darst. |c 30 cm | ||
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| 490 | 1 | |a Wissenschaftliche Berichte / Forschungszentrum Karlsruhe Technik und Umwelt |v 7017 | |
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| 943 | 1 | |a oai:aleph.bib-bvb.de:BVB01-013125728 | |
Datensatz im Suchindex
| DE-BY-TUM_call_number | 0001 2005 B 466 |
|---|---|
| DE-BY-TUM_katkey | 1505561 |
| DE-BY-TUM_location | Mag |
| DE-BY-TUM_media_number | 040005947228 |
| _version_ | 1820814482328256512 |
| adam_text | Abstract i
Zusammenfassung ii
Acknowledgements iii
Table of contents v
Abbreviation ix
1. Introduction 1
1.1 Cancer and carcinogenesis 1
1.1.1 Cancer and cancer cells 1
1.1.2 Epigenetic mechanism in carcinogenesis 2
1.1.3 Genetic mechanism in carcinogenesis 3
1.1.3.1 Oncogene 3
1.1.3.2 Tumor suppressor gene 4
1.2 The tumor suppressor gene nf 2 5
1.2.1 NF 2, the disease 6
1.2.2 nf 2, the gene 6
1.2.2.1 The structure of the nf 2 gene 6
1.2.2.2 The regulation of the nf 2 gene 7
1.2.2.3 Mutations of the nf 2 gene 8
1.2.3 Merlin, the protein encoded by the nf 2 gene 9
1.2.3.1 The structure of merlin 9
1.2.3.2 The distribution of merlin 11
1.2.3.3 The interaction partners of merlin 13
1.2.3.4 The function of merlin 16
1.2.3.5 The regulation of merlin 17
1.3 Aims of my project 19
2. Materials and methods 2 •
2.1 Materials 2 x
2.1.1 General Chemicals 21
2.1.2 Primers 22
2.1.2.1 Standard Primers 22
2.1.2.2 Mutagenesis Primers 22
2.1.3 Plasmids 23
2.1.4 Enzymes 23
2.1.5 Cell culture regents 24
2.1.6 Cell lines 24
2.1.7 Antibodies 24
2.1.7.1 Primary antibodies 24
2.1.7.2 Secondary antibodies 25
2.2 Methods 25
2.2.1 Preparation of competent bacteria 25
2.2.1.1 Preparation of chemically competent E.Coli. 25
2.2.1.2 Preparation of electrocompetent E.Coli. 26
2.2.2 Transformation of E. Coli. 26
2.2.2.1 chemical transformation 26
2.2.2.2 Electroporation transformation 26
2.2.3 Plasmid preparation 27
2.2.3.1 Small scale plasmid preparation (mimipreparation) 27
2.2.3.2 Large scale plasmid preparation (maxipreparation) 27
2.2.4 Determination of nucleic acid concentration 28
2.2.5 Restriction endonuclease digestion of DNA 28
2.2.6 Nucleic acid analysis by agarose gel electrophoresis 29
2.2.7 Isolation/purification of DNA from agarose gels 29
2.2.8 Phenol/chloroform extraction of nucleic acid 29
2.2.9 Ligation 30
2.2.10 Precipitation of nucleic acid 30
2.2.11 Polymerase Chain Reaction (PCR) 30
2.2.12 Site directed mutagenesis 30
2.2.13 TOPO TA cloning 31
2.2.14 Cell culture 31
2.2.15 Transfection 32
2.2.16 Cell extracts preparation for Western blot analysis 32
2.2.17 SDS polyacrylamide gel electrophoresis (SDS PAGE) 33
2.2.18 Staining of gels 33
2.2.19 Western blotting 33
2.2.20 Probing 34
2.2.21 Stripping Western blot membrane 34
2.2.22 Immunoprecipitation 34
2.2.22.1 Cell lysis and pre clearing 34
2.2.22.2 Immunoprecipitation 35
2.2.23 In vitro dephosphorylation assay 35
2.2.24 Preparation of GST and GST fusion protein (large scale) 35
2.2.25 GST pull down assay 36
2.2.26 Cleavage of the GST tag from GST fusion protein 36
2.2.27 In vitro binding assay 37
2.2.28 Soft agar assay 37
3. Results 39
3.1 C merlin is regulated similar to full length merlin 39
3.1.1 Phosphorylation of C merlin 39
3.1.2 Regulation of C merlin phosphorylation 40
3.1.3 Identification of the phosphorylation site in C merlin 40
3.2 PP1, but not PP2A, is the catalytic subunit of merlin phosphatase 43
3.2.1 The effect of okadaic acid on merlin phosphorylation 45
3.2.2 In vitro dephosphorylation of C merlin 46
3.2.3 Interaction of merlin with PP1 48
3.3 Myosin/moesin phosphatase is the phosphatase of merlin 49
3.3.1 Interaction between merlin and myosin/moesin phosphatase 51
3.3.2 The relationship between the phosphorylation of merlin 59
and the activity of myosin/moesin phosphatase 3.3.3 The effect of CPI 17 on the phosphorylation and function of merlin.. 62
3.3.4 The effect of CPI 17 on cell growth 65
4. Discussion 67
4.1 Growth control and contact inhibition 67
4.2 The system of RT4 C cell line 68
4.3 The interaction of merlin with myosin/moesin phosphatase 69
4.4 The regulation of the myosin/moesin phosphatase activity 72
4.5 The function merlin 74
4.6 The growth promoting function of CPI 17 75
5. References 79
Appendix 105
|
| any_adam_object | 1 |
| author | Jin, Hongchuan 1974- |
| author_GND | (DE-588)129756334 |
| author_facet | Jin, Hongchuan 1974- |
| author_role | aut |
| author_sort | Jin, Hongchuan 1974- |
| author_variant | h j hj |
| building | Verbundindex |
| bvnumber | BV019800145 |
| ctrlnum | (OCoLC)76672790 (DE-599)BVBBV019800145 |
| discipline | Biologie |
| edition | Als Ms. gedr. |
| format | Thesis Book |
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| genre | (DE-588)4113937-9 Hochschulschrift gnd-content |
| genre_facet | Hochschulschrift |
| id | DE-604.BV019800145 |
| illustrated | Illustrated |
| indexdate | 2024-12-23T18:15:19Z |
| institution | BVB |
| language | English |
| oai_aleph_id | oai:aleph.bib-bvb.de:BVB01-013125728 |
| oclc_num | 76672790 |
| open_access_boolean | |
| owner | DE-91 DE-BY-TUM DE-12 |
| owner_facet | DE-91 DE-BY-TUM DE-12 |
| physical | XI, 105 S. Ill., graph. Darst. 30 cm |
| publishDate | 2004 |
| publishDateSearch | 2004 |
| publishDateSort | 2004 |
| publisher | FZKA |
| record_format | marc |
| series2 | Wissenschaftliche Berichte / Forschungszentrum Karlsruhe Technik und Umwelt |
| spellingShingle | Jin, Hongchuan 1974- Activation of the tumor suppressor merlin by myosin, moesin phosphatase |
| subject_GND | (DE-588)4113937-9 |
| title | Activation of the tumor suppressor merlin by myosin, moesin phosphatase |
| title_alt | FZKA-7017 |
| title_auth | Activation of the tumor suppressor merlin by myosin, moesin phosphatase |
| title_exact_search | Activation of the tumor suppressor merlin by myosin, moesin phosphatase |
| title_full | Activation of the tumor suppressor merlin by myosin, moesin phosphatase Hongchuan Jin. Forschungszentrum Karlsruhe GmbH, Karlsruhe |
| title_fullStr | Activation of the tumor suppressor merlin by myosin, moesin phosphatase Hongchuan Jin. Forschungszentrum Karlsruhe GmbH, Karlsruhe |
| title_full_unstemmed | Activation of the tumor suppressor merlin by myosin, moesin phosphatase Hongchuan Jin. Forschungszentrum Karlsruhe GmbH, Karlsruhe |
| title_short | Activation of the tumor suppressor merlin by myosin, moesin phosphatase |
| title_sort | activation of the tumor suppressor merlin by myosin moesin phosphatase |
| topic_facet | Hochschulschrift |
| url | http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=013125728&sequence=000002&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| volume_link | (DE-604)BV010181833 |
| work_keys_str_mv | AT jinhongchuan activationofthetumorsuppressormerlinbymyosinmoesinphosphatase AT jinhongchuan fzka7017 |