Identification of novel interaction partners of ZNRF3/RNF43 and analysis of the functional consequence in Wnt signaling pathway: = Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg
Gespeichert in:
| 1. Verfasser: | |
|---|---|
| Format: | Abschlussarbeit Buch |
| Sprache: | English |
| Veröffentlicht: |
Erlangen ; Nürnberg
[2019]
|
| Schlagworte: | |
| Online-Zugang: | Inhaltsverzeichnis Inhaltsverzeichnis |
| Beschreibung: | 85 Seiten Illustrationen, Diagramme 21 cm |
Internformat
MARC
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| 001 | BV046225628 | ||
| 003 | DE-604 | ||
| 005 | 20191121 | ||
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| 008 | 191031s2019 gw a||| m||| 00||| eng d | ||
| 016 | 7 | |a 1197761160 |2 DE-101 | |
| 035 | |a (OCoLC)1126565009 | ||
| 035 | |a (DE-599)DNB1197761160 | ||
| 040 | |a DE-604 |b ger |e rda | ||
| 041 | 0 | |a eng | |
| 044 | |a gw |c XA-DE | ||
| 049 | |a DE-29 |a DE-29T |a DE-355 | ||
| 084 | |a 570 |2 sdnb | ||
| 100 | 1 | |a Gunasekaran, Mekala |e Verfasser |0 (DE-588)1198170654 |4 aut | |
| 245 | 1 | 0 | |a Identification of novel interaction partners of ZNRF3/RNF43 and analysis of the functional consequence in Wnt signaling pathway |b = Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg |c vorgelegt von Mekala Gunasekaran |
| 246 | 1 | 3 | |a Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg |
| 246 | 1 | 1 | |a Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg |
| 264 | 1 | |a Erlangen ; Nürnberg |c [2019] | |
| 300 | |a 85 Seiten |b Illustrationen, Diagramme |c 21 cm | ||
| 336 | |b txt |2 rdacontent | ||
| 337 | |b n |2 rdamedia | ||
| 338 | |b nc |2 rdacarrier | ||
| 502 | |b Dissertation |c Friedrich-Alexander-Universität Erlangen-Nürnberg |d 2019 | ||
| 650 | 0 | 7 | |a Wnt-Proteine |0 (DE-588)7515809-7 |2 gnd |9 rswk-swf |
| 650 | 0 | 7 | |a Signaltransduktion |0 (DE-588)4318717-1 |2 gnd |9 rswk-swf |
| 655 | 7 | |0 (DE-588)4113937-9 |a Hochschulschrift |2 gnd-content | |
| 689 | 0 | 0 | |a Wnt-Proteine |0 (DE-588)7515809-7 |D s |
| 689 | 0 | 1 | |a Signaltransduktion |0 (DE-588)4318717-1 |D s |
| 689 | 0 | |5 DE-604 | |
| 856 | 4 | 2 | |m B:DE-101 |q application/pdf |u https://d-nb.info/1197761160/04 |3 Inhaltsverzeichnis |
| 856 | 4 | 2 | |m DNB Datenaustausch |q application/pdf |u http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=031604217&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |3 Inhaltsverzeichnis |
| 999 | |a oai:aleph.bib-bvb.de:BVB01-031604217 | ||
Datensatz im Suchindex
| _version_ | 1804180631103471616 |
|---|---|
| adam_text | CONTENTS
1.
ZUSAMMENFASSUNG
.......................................................................................................
1
2.
SUMMARY
.............................................................................................................................
2
3.
INTRODUCTION
...................................................................................................................
3
3.1
THE
WNT
SIGNALING
PATHWAY
..............................................................................................
3
3.2
AGONISTS
AND
ANTAGONISTS
OF
WNT
SIGNALING
PATHWAY
................................................
5
3.2.1
AGONISTS
.............................................................................................................................
5
3.2.2
ANTAGONISTS
........................................................................................................................
6
3.3
UBIQUITIN
MECHANISM
.........................................................................................................
6
3.3.1
E3
UBIQUITIN
LIGASES
...........................................................................................................
7
3.4
RNF43
AND
ZNRF3
..............................................................................................................
7
3.4.1
STRUCTURAL
CHARACTERIZATION
OF
RNF43
AND
ZNRF3
.........................................................
8
3.4.2
ROLE
OF
RNF43
AND
ZNRF3
IN
CANCERS
...........................................................................
9
3.5
LIM
DOMAIN
PROTEINS
........................................................................................................
11
3.6
AJUBA
AND
LIMD1
...............................................................................................................
11
3.6.1.1
AJUBA
AND
LIMD1
IN
WNT
SIGNALING
PATHWAY
..............................................................
12
3.6.1.2
AJUBA
AND
LIMD1
IN
HIPPO
SIGNALING
PATHWAY
...........................................................
12
3.6.1.3
AJUBA
AND
LIMD1
IN
COLORECTAL
CANCER
........................................................................
13
3.7
YEAST
TWO
HYBRID
SYSTEM
................................................................................................
14
3.7.1
MODEL
OF
THE
YEAST
TWO
HYBRID
PRINCIPLE
..........................................................................
14
4.
AIM
.......................................................................................................................................
15
5.
RESULTS
.............................................................................................................................
16
5.1
DESIGNING
ZNRF3-BAIT
.....................................................................................................
16
5.2
AUTO-ACTIVATION
TEST
OF
ZNRF3-BAIT
...............................................................................
17
5.3
LIST
OF
PREY
PROTEINS
FOUND
IN
THE
Y2H
SCREEN
...........................................................
18
5.4
RE-TRANSFORMATION
OF
BAIT
AND
PREY
IN
THE
YEAST
........................................................
20
5.5
SCHEMATIC
DIAGRAM
OF
THE
STRUCTURE
OF
AJUBA
AND
LIMD1
PROTEINS
........................
21
5.6
AMINO
ACID
SEQUENCE
OF
AJUBA
AND
LIMD1
PREY
..........................................................
21
5.7
INTERACTION
OF
AJUBA
AND
ZNRF3-BAIT
IN
MAMMALIAN
CELLS
.....................................
22
5.8
INTERACTION
OF
AJUBA
AND
LIIVID1
WITH
RNF43
IN
MAMMALIAN
CELLS
.........................
23
5.9
INTERACTION
MAPPING
OF
AJUBA
WITH
ZNRF3
..................................................................24
5.10
INTERACTION
OF
ENDOGENOUS
LIMBI
WITH
ZNRF3
..........................................................25
5.11
COLOCALIZATION
ANALYSIS
OF
AJUBA
WITH
ZNRF3
AND
RNF43
..................................... 26
5.11.1
AJUBA+ZNRF3
................................................................................................................
26
5.11.2
AJUBA+RNF43
................................................................................................................
27
5.11.3
EFFECT
OF
THE
PRESENCE
OF
AJUBA
ON
ZNRF3
LOCALIZATION
..............................................27
5.12
COLOCALIZATION
ANALYSIS
OF
LIMBI
WITH
RNF43
..........................................................
29
5.13
INTERACTION
MAPPING
OF
RNF43
WITH
AJUBA
..............................................................
29
5.14
POSSIBILITY
OF
AJUBA
INTERFERING
IN
BVL2
INTERACTION
WITH
ZNRF3
..........................
31
5.15
COMPETITIVE
BINDING
OF
AJUBA
AND
BVL2
TO
ZNRF3
.................................................
31
5.16
FUNCTIONAL
RELEVANCE
OF
AJUBA/LIMB1
INTERACTION
WITH
ZNRF3/RNF43
...............
32
5.16.1
ZNRF3
INHIBITS
WNT
SIGNALING
ACTIVATION
......................................................................
32
5.16.2
EFFECT
OF
AJUBA
AND
LIMBI
ON
WNT
SIGNALING
ACTIVATION
........................................... 33
5.16.3
EFFECT
OF
LIMBI
ON
ZNRF3
MEDIATED
UBIQUITINATION
OF
FZB
....................................
33
5.16.4
GENERATION
OF
STABLE
CELLS
EXPRESSING
GFP-LIMB1
AND
GFP-AJUBA
........................ 34
5.16.5
LOCALIZATION
ANALYSIS
OF
STABLE
CELLS
EXPRESSING
GFP-LIMB1
AND
GFP-AJUBA
.......
35
5.16.6
CELL
SURFACE
FZB
LEVELS
IN
STABLY
TRANSFECTED
AJUBA/LIMB1
CELLS
.............................
35
5.16.7
SIRNA
MEDIATED
KNOCKDOWN
OF
AJUBA
AND
LIMBI
....................................................
36
5.16.8
CELL
SURFACE
FZB
LEVELS
IN
AJUBA
AND
LIMBI
KNOCKDOWN
CELLS
.................................
37
5.16.9
CELL
SURFACE
FZB
LEVELS
IN
STABLE
AJUBA/LIMB1
CELLS
TRANSFECTED
WITH
ZNRF3
....... 38
5.16.10
EFFECT
OF
RNF43
AND
ZNRF3
ON
CELL
SURFACE
FZB
INTERNALIZATION
........................... 38
5.16.11
EFFECT
OF
ZNRF3
ON
CELL
SURFACE
SNAP-FZ5
IN
STABLE
CELLS
........................................39
5.17
EFFECT
OF
WNT3A
ON
ENDOGENOUS
LIMBI
PROTEIN
LEVELS
.............................................
40
6.
BISCUSSION
......................................................................................................................
41
6.1
IDENTIFICATION
OF
POTENTIAL
INTERACTION
PARTNERS
OF
ZNRF3
USING
YEAST
TWO
HYBRID
SCREEN
........................................................................................................................................
41
6.2
AJUBA
AND
LIMBI
HITS
.......................................................................................................
41
6.3
EFFECT
OF
ZNRF3
AND
AJUBA/LIMB1
ON
WNT
MEDIATED
SIGNALING
...............................
42
6.4
ANALYSIS
OF
AJUBA/LIMB1
AND
ZNRF3/RNF43
LOCALIZATION
.......................................
43
6.5
INTERACTION
MAPPING
OF
AJUBA
AND
ZNRF3/RNF43
.....................................................
43
6.6
AJUBA
AND
DVL2
COMPETES
TO
BIND
TO
ZNRF3
.............................................................
44
6.7
IMPACT
OF
AJUBA/LIMD1
INTERACTION
WITH
ZNRF3/RNF43
ON
WNT
SIGNALING
PATHWAY
WITH
REGARD
TO
WNT
RECEPTOR
LEVEL
.........................................................................................
45
6.7.1
AJUBA/LIMD1
STABILIZES
CELL
SURFACE
FZD
LEVELS
.......................................................
45
6.8
WNT
MEDIATED
DESTABILIZATION
OF
LIMD1
PROTEIN
LEVELS
..............................................
47
6.9
NOVEL
ROLE
OF
AJUBA
AND
LIMD1
IN
WNT/P-CATENIN
SIGNALING
PATHWAY
....................
47
7.
FUTURE
WORK
..................................................................................................................49
8.
MATERIALS
AND
METHODS
...........................................................................................50
8.1
MATERIALS
....................................................................................................................
50
8.1.1
CHEMICALS
.......................................................................................................................
50
8.1.2
ENZYMES
.........................................................................................................................
50
8.1.3
YEAST
GROWTH
MEDIUM
AND
SOLUTIONS
.........................................................................
51
8.1.3.
A
AMINO
ACID
SOLUTIONS
(10X)
..........................................................................................
51
8.1
3
D
AMINO
ACID
DROPOUT
SOLUTIONS
......................................................................................
51
8.1.
3.
C
AMINO
ACID
DROPOUT
PLATES
..........................................................................................
52
8.1.3
D
OTHER
YEAST
SOLUTIONS
AND
BUFFERS
...............................................................................
53
8.1.4
BUFFERS
AND
SOLUTIONS
...................................................................................................
53
8.1.5
BACTERIA
GROWTH
MEDIUM
AND
AGAR
PLATES
.................................................................
55
8.1.6
CELL
LINES
..........................................................................................................................
55
8.1.7
CELL
CULTURE
MEDIUM
......................................................................................................
56
8.1.80RGANISMS
......................................................................................................................
56
8.1.9
PLASMIDS
.........................................................................................................................
57
8.2.0
ANTIBODIES
.......................................................................................................................
59
8.2.1
OLIGONUCLEOTIDES
...........................................................................................................
60
8.2.2
TECHNICAL
EQUIPMENTS
..................................................................................................
60
8.3
METHODS
.......................................................................................................................
61
8.3.1
YEAST
TECHNIQUES
..........................................................................................................
61
8.3.1.1
GROWING
YEAST
L40
......................................................................................................
61
8.3.1.2CARRIER
DNA
PREPARATION
..................................................................................................
61
8.3.1.3
SMALL
SCALE
TRANSFORMATION
.............................................................................................
61
8.3.1.4
YEAST
TWO
HYBRID
SCREEN
...............................................................................................
62
8.3.1.5
LACZ
ASSAY
.......................................................................................................................
62
8.3.1.6
YEAST
DNA
EXTRACTION
......................................................................................................
62
8.3.1.7
HB101
ELECTROPORATION
..................................................................................................
63
6.3.1.8
PLASMID
DNA
EXTRACTION
FROM
HB101
..........................................................................
63
8.3.1.9
YEAST
PCR
.......................................................................................................................
63
8.3.2
MOLECULAR
BIOLOGY
TECHNIQUES
..................................................................................
64
8.3.2.1
PCR
..................................................................................................................................
64
8.3.2.2
RESTRICTION
DIGESTION,
LIGATION
&
TRANSFORMATION
.......................................................
64
8.3.2.3
MINI
AND
MAXI
PREPARATION
..............................................................................................
65
8.3.2.4
DNA
SEQUENCING
.............................................................................................................
65
8.3.3
CELL
CULTURE
TECHNIQUES
..................................................................................................
65
8.3.3.1
GENERATING
STABLE
CELL
LINES
..........................................................................................
65
8.3.4
BIOCHEMICAL
TECHNIQUES
................................................................................................66
6.3.4.1
IMMUNOPRECIPITATION
.......................................................................................................
66
8.3.4.2
IMMUNOFLUORESCENCE
......................................................................................................
66
6.3.4.3
UBIQUITINATION
ASSAY
.......................................................................................................
67
8.3.4.4
REPORTER
ASSAY
...............................................................................................................
67
8.3.4.5
SIRNA
TRANSFECTION
.........................................................................................................
67
6.3.4.6
WESTERN
BLOTTING
.............................................................................................................
68
6.3.4
7
FACS
..............................................................................................................................
68
8.3.4
6
SNAP
CELL
SURFACE
ASSAY
..............................................................................................
69
9.
LITERATURE
........................................................................................................................
70
10.
SUPPLEMENT
....................................................................................................................
79
10.1
ABBREVIATIONS/UNITS
...........................................................................................................
79
10.2
VECTOR
MAP:
PCDNA3
FLAG-HA-
AJUBA
.............................................................................
81
10.3
AJUBA
AND
LIMD1
PREY
SEQUENCES
FROM
YEAST
TWO
HYBRID
SCREEN
................................
61
10.4
FIGURE
S1
.............................................................................................................................
62
10.5
FIGURE
S2
.............................................................................................................................
62
11.
PUBLICATION
S3
ACKNOWLEDGEMENTS
|
| any_adam_object | 1 |
| author | Gunasekaran, Mekala |
| author_GND | (DE-588)1198170654 |
| author_facet | Gunasekaran, Mekala |
| author_role | aut |
| author_sort | Gunasekaran, Mekala |
| author_variant | m g mg |
| building | Verbundindex |
| bvnumber | BV046225628 |
| ctrlnum | (OCoLC)1126565009 (DE-599)DNB1197761160 |
| discipline | Biologie |
| format | Thesis Book |
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| genre | (DE-588)4113937-9 Hochschulschrift gnd-content |
| genre_facet | Hochschulschrift |
| id | DE-604.BV046225628 |
| illustrated | Illustrated |
| indexdate | 2024-07-10T08:38:49Z |
| institution | BVB |
| language | English |
| oai_aleph_id | oai:aleph.bib-bvb.de:BVB01-031604217 |
| oclc_num | 1126565009 |
| open_access_boolean | |
| owner | DE-29 DE-29T DE-355 DE-BY-UBR |
| owner_facet | DE-29 DE-29T DE-355 DE-BY-UBR |
| physical | 85 Seiten Illustrationen, Diagramme 21 cm |
| publishDate | 2019 |
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| record_format | marc |
| spelling | Gunasekaran, Mekala Verfasser (DE-588)1198170654 aut Identification of novel interaction partners of ZNRF3/RNF43 and analysis of the functional consequence in Wnt signaling pathway = Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg vorgelegt von Mekala Gunasekaran Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg Erlangen ; Nürnberg [2019] 85 Seiten Illustrationen, Diagramme 21 cm txt rdacontent n rdamedia nc rdacarrier Dissertation Friedrich-Alexander-Universität Erlangen-Nürnberg 2019 Wnt-Proteine (DE-588)7515809-7 gnd rswk-swf Signaltransduktion (DE-588)4318717-1 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Wnt-Proteine (DE-588)7515809-7 s Signaltransduktion (DE-588)4318717-1 s DE-604 B:DE-101 application/pdf https://d-nb.info/1197761160/04 Inhaltsverzeichnis DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=031604217&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Gunasekaran, Mekala Identification of novel interaction partners of ZNRF3/RNF43 and analysis of the functional consequence in Wnt signaling pathway = Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg Wnt-Proteine (DE-588)7515809-7 gnd Signaltransduktion (DE-588)4318717-1 gnd |
| subject_GND | (DE-588)7515809-7 (DE-588)4318717-1 (DE-588)4113937-9 |
| title | Identification of novel interaction partners of ZNRF3/RNF43 and analysis of the functional consequence in Wnt signaling pathway = Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg |
| title_alt | Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg |
| title_auth | Identification of novel interaction partners of ZNRF3/RNF43 and analysis of the functional consequence in Wnt signaling pathway = Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg |
| title_exact_search | Identification of novel interaction partners of ZNRF3/RNF43 and analysis of the functional consequence in Wnt signaling pathway = Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg |
| title_full | Identification of novel interaction partners of ZNRF3/RNF43 and analysis of the functional consequence in Wnt signaling pathway = Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg vorgelegt von Mekala Gunasekaran |
| title_fullStr | Identification of novel interaction partners of ZNRF3/RNF43 and analysis of the functional consequence in Wnt signaling pathway = Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg vorgelegt von Mekala Gunasekaran |
| title_full_unstemmed | Identification of novel interaction partners of ZNRF3/RNF43 and analysis of the functional consequence in Wnt signaling pathway = Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg vorgelegt von Mekala Gunasekaran |
| title_short | Identification of novel interaction partners of ZNRF3/RNF43 and analysis of the functional consequence in Wnt signaling pathway |
| title_sort | identification of novel interaction partners of znrf3 rnf43 and analysis of the functional consequence in wnt signaling pathway identifizierung neuer interaktionspartner von znrf3 rnf43 und analyse der funktionellen konsequenzen im wnt signalweg |
| title_sub | = Identifizierung neuer Interaktionspartner von ZNRF3/RNF43 und Analyse der funktionellen Konsequenzen im Wnt-Signalweg |
| topic | Wnt-Proteine (DE-588)7515809-7 gnd Signaltransduktion (DE-588)4318717-1 gnd |
| topic_facet | Wnt-Proteine Signaltransduktion Hochschulschrift |
| url | https://d-nb.info/1197761160/04 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=031604217&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| work_keys_str_mv | AT gunasekaranmekala identificationofnovelinteractionpartnersofznrf3rnf43andanalysisofthefunctionalconsequenceinwntsignalingpathwayidentifizierungneuerinteraktionspartnervonznrf3rnf43undanalysederfunktionellenkonsequenzenimwntsignalweg AT gunasekaranmekala identifizierungneuerinteraktionspartnervonznrf3rnf43undanalysederfunktionellenkonsequenzenimwntsignalweg |
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Inhaltsverzeichnis