Complex formation of AID and its function in innate immunity
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| Sprache: | English |
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2010
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| LEADER | 00000nam a2200000 c 4500 | ||
|---|---|---|---|
| 001 | BV036966164 | ||
| 003 | DE-604 | ||
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| 007 | t | ||
| 008 | 110119s2010 ad|| m||| 00||| eng d | ||
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| 035 | |a (OCoLC)702667406 | ||
| 035 | |a (DE-599)DNB1009414682 | ||
| 040 | |a DE-604 |b ger |e rakddb | ||
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| 082 | 0 | |a 571.9648 |2 22/ger | |
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| 100 | 1 | |a Metzner, Mirjam |d 1979- |e Verfasser |0 (DE-588)143196456 |4 aut | |
| 245 | 1 | 0 | |a Complex formation of AID and its function in innate immunity |c vorgelegt von Mirjam Metzner |
| 264 | 1 | |c 2010 | |
| 300 | |a V, 128 S. |b Ill., graph. Darst. |c 21 cm | ||
| 336 | |b txt |2 rdacontent | ||
| 337 | |b n |2 rdamedia | ||
| 338 | |b nc |2 rdacarrier | ||
| 502 | |a Erlangen-Nürnberg, Univ., Diss., 2010 | ||
| 538 | |a Langzeitarchivierung Nationalbibliothek gewährleistet | ||
| 655 | 7 | |0 (DE-588)4113937-9 |a Hochschulschrift |2 gnd-content | |
| 776 | 0 | 8 | |i Erscheint auch als |n Online-Ausgabe |o urn:nbn:de:bvb:29-opus-22025 |
| 856 | 4 | |u https://open.fau.de/handle/openfau/1513 |x Verlag |z kostenfrei |3 Volltext | |
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| 912 | |a ebook | ||
| 999 | |a oai:aleph.bib-bvb.de:BVB01-020880964 | ||
Datensatz im Suchindex
| DE-BY-TUM_katkey | 1753370 |
|---|---|
| DE-BY-UBG_katkey | 2753576 |
| _version_ | 1816713564058025985 |
| adam_text | TABLE
OF
CONTENTS
1
ZUSAMMENFASSUNG
1
SUMMARY
2
2
INTRODUCTION
3
2.1
ADAPTIVE
AND
INNATE
IMMUNITY
3
2.2
B
CELL
DEVELOPMENT
AND
ACTIVATION
3
2.3
ACTIVATION-INDUCED
CYTIDINE
DEAMINASE
(AID)
4
2.3.1
AID
IN
ADAPTIVE
IMMUNITY
4
2.3.2
BASIC
FACTS
ABOUT AID
5
2.3.3
APOBEC
FAMILY
OF
CYTIDINE
DEAMINASES
6
2.3.4
AID
COFACTORS
8
2.3.5
AID
MULTIMERS
AND
COMPLEXES
8
2.3.6
AID
AND
RNA
9
2.3.7
AID
DEFICIENCY
AND
AUTOIMMUNE
DISEASE
10
2.3.8
AID
IN
INNATE
IMMUNITY
11
2.3.8.1
AID
PROTECTS
FROM
VIRAL
INFECTION
11
2.3.8.2
AID
INHIBITS
L1
RETROTRANSPOSITION
12
2.4
ENDOGENOUS
RETROELEMENTS
13
2.4.1
L1
ELEMENTS
13
2.4.1.1
THE
L1
RETROTRANSPOSITION
CYCLE
14
2.4.1.2
EXPRESSION
PATTERN
15
2.4.1.3
RESTRICTION
OF
L1
ACTIVITY
15
2.4.2
RETROELEMENTS
AND
AUTOIMMUNE
DISEASE
15
3
SPECIFIC
AIMS
17
4
RESULTS
18
4.1
SET-UP
OF
CELL
SYSTEMS
FOR
ENDOGENOUS
AND
EXOGENOUS
AID
PROTEIN
EXPRESSION
18
4.2
ANTIBODIES
AGAINST
AID
20
4.2.1
COMPARISON
OF
EXISTING
AID
SPECIFIC
ANTIBODIES
21
4.2.2
GENERATION
OF
NEW
MONOCLONAL
AID
SPECIFIC
ANTIBODIES
23
4.3
TWO
FORMS
OF
AID
DIFFERING
IN
THEIR
ABILITY
TO
BIND
AGAROSE
26
4.3.1
TWO
AID
VARIANTS
WITHIN
A
CELL:
ONE
BINDS
TO
AGAROSE,
THE
OTHER
ONE
DOES
NOT
26
4.3.2
AID
BINDING
TO
AGAROSE
IS
NOT
RNASE-SENSITIVE
29
4.3.3
AID
EXPRESSED
IN
E.
COLI
FEATURES
SIMILAR
AGAROSE
BINDING
CHARACTERISTICS
30
4.3.4
CONCLUSION
30
4.4
AID
FORMS
HIGH
MOLECULAR
MASS
COMPLEXES
IN
THE
CYTOPLASM
31
4.4.1
SIZE
EXCLUSION
FRACTIONATION
OF
AID-POSITIVE
CELL
LYSATES
31
4.4.2
INHIBITION
OF
NUCLEAR
EXPORT
AND
PROTEASOME
DEGRADATION
OF
AID
33
4.4.3
PROTEIN
DEGRADATION
DURING
FRACTIONATION
AS
A
CONSEQUENCE
OF
RNASE
TREATMENT
34
4.4.4
COMPOSITION
OF
LARGE
AND
MEDIUM
AID
PROTEIN
COMPLEXES
36
4.4.5
CO-IMMUNOPRECIPITATION
WITH
AID
TO
IDENTIFY
AID
COMPLEX
COMPONENTS
38
4.4.6
PURIFICATION
OF
AID
COMPLEXES
VIA
ION
EXCHANGE
CHROMATOGRAPHY
40
4.4.7
AGAROSE
BINDING
OF
HIGH
AND
MEDIUM
MOLECULAR
MASS
AID
COMPLEXES
42
4.5
ANOTHER
FUNCTION
OF
AID
-
INNATE
IMMUNITY
TO
ENDOGENOUS
RETROELEMENTS
43
4.5.1
AID
BINDS
TO
L1
MRNA
BUT
NOT
TO
STERILE
GERM
LINE
TRANSCRIPTS
43
4.5.2
AID
DECREASES
L1
PROTEIN
LEVEL
46
4.5.2.1
ORF1
PROTEIN
LEVEL
IN
PRIMARY
MOUSE
B
CELLS
46
4.5.2.2
ORF1
PROTEIN
LEVEL
IN
HUMAN
CELL
LINES
47
4.5.2.3
ORF1
PROTEIN
LEVEL
IN
B
CELLS
OF
AID-DEFICIENT
PATIENTS
48
4.5.2.4
ORF1
PROTEIN
DEGRADATION
DURING
FPLC
FRACTIONATION
49
4.5.3
AID
BINDS
TO
ORF1
PROTEIN
51
4.5.4
L1
MRNA
IS
NOT
INCREASED
IN
AID-DEFICIENT
B
CELLS
51
4.6
AID
AND
AUTOIMMUNE
DISEASE
52
4.7
HUMAN
APOBEC3
(HA3)
TRANSGENIC
MICE
AND
L1
RETROELEMENTS
55
4.7.1
A3G
FORMS
HMM
AND
LMM
COMPLEXES
IN
HA3
TRANSGENIC
MICE
55
4.7.2
A3G
AND
ORF1
PROTEIN
LEVEL
IN
HA3
TRANSGENIC
MICE
57
5
DISCUSSION
58
5.1
AGAROSE
BINDING
OF
AID
-
CAVEATS
AND
RELEVANCE
58
5.2
AID
COMPLEXES
-
ISOLATION
AND
IDENTIFICATION
OF
THEIR
COMPONENTS
59
II
5.3
AID
SUPPRESSES
ENDOGENOUS
RETROELEMENTS
-
THE
UNDERLYING
MECHANISM
60
5.4
AID
MEDIATES
INNATE
IMMUNITY
61
5.4.1
AID
PROTECTS
FROM
AUTOIMMUNITY
BY
SUPPRESSING
RETROELEMENTS
62
5.4.1.1
CAUSES
OF
AUTOIMMUNITY
62
5.4.1.2
AID,
RETROELEMENTS
AND
AUTOIMMUNITY
63
5.4.1.3
ABSENCE
OF
AID
CAUSES
AUTOIMMUNITY
-
HOW
IT
MAY
WORK
63
5.4.1.4
ABSENCE
OF
AID
CAUSES
AUTOIMMUNITY
-
HOW
TO
PROVE
IT
64
5.4.2
AID
PROTECTS
FROM
TUMORIGENESIS
BY
SUPPRESSING
RETROELEMENTS
65
5.5
AID
COMPLEXES
-
ASSOCIATED
WITH
P
BODIES
AND
STRESS
GRANULES?
65
5.6
SIGNIFICANCE
OF
THIS
STUDY
67
6
MATERIALS
AND
METHODS
69
6.1
MATERIALS
69
6.1.1
OLIGONUCLEOTIDES
69
6.1.1.1
PRIMER
FOR
CLONING
69
6.1.1.2
PRIMER
FOR
SEQUENCING
69
6.1.1.3
PRIMER
FOR
RT-PCR
70
6.1.1.4
PRIMER
FOR
GENOTYPING
PCR
70
6.1.2
PLASMIDS
71
6.1.3
EUKARYOTIC
CELLS
73
6.1.3.1
CELL
LINES
73
6.1.3.2
PRIMARY
CELLS
74
6.1.4
ANTIBODIES
74
6.1.5
MOUSE
STRAINS
76
6.2
METHODS
77
6.2.1
NUCLEIC
ACIDS
77
6.2.1.1
ISOLATION
OF
NUCLEIC
ACIDS
77
A)
RNA
EXTRACTION
77
B)
DNA
EXTRACTION
77
6.2.1.2
CDNA
SYNTHESIS
78
6.2.1.3
POLYMERASE
CHAIN
REACTION
(PCR)
79
A)
PCR
USING
GENOMIC
AND
PLASMID
DNA
79
B)
(SEMIQUANTITATIVE)
RT-PCR
79
C)
QUANTITATIVE
REAL-TIME
RT-PCR
(QRT-PCR)
80
ILL
6.2.
1.4
DETECTION
OF
NUCLEIC
ACIDS
81
6.2.1.5
CLONING
81
6.2.1.6
SEQUENCING
82
6.2.2
CELLS
83
6.2.2.1
CELL
CULTURE
OF
MAMMALIAN
CELLS
83
6.2.2.2
ISOLATION
OF
SPLENIC
CELLS
84
6.2.2.3
AID
INDUCTION
IN
PRIMARY
B
CELLS
84
6.2.2.4
AID
INDUCTION
IN
STABLE
HELA
AND
18-81
CELL
TRANSFECTANTS
84
6.2.2.5
TRANSFECTION
OF
EUKARYOTIC
CELLS
85
A)
STABLE
TRANSFECTION
85
B)
TRANSIENT
TRANSFECTION
85
6.2.2.6
PRODUCTION
OF
RETROVIRAL
SUPERNATANT
AND
INFECTION
OF
PRIMARY
B
CELLS
86
6.2.3
PROTEIN
86
6.2.3.1
PROTEIN
LYSATES
86
6.2.3.2
SDS
PAA
GEL
ELECTROPHORESIS
(SDS-PAGE)
AND
WESTERN
BLOT
ANALYSIS
87
6.2.3.3
PROTEIN
STAINING
OF
PAA
GELS
89
A)
COOMASSIE
BLUE
STAINING
89
B)
SYPRO
RUBY
STAINING
90
C)
SILVER
STAINING
91
6.2.3.4
AGAROSE
BINDING
ASSAY
92
6.2.3.5
(CO-)
IMMUNOPRECIPITATION
93
6.2.3.6
TANDEM-AFFINITY
PURIFICATION
(TAP)
93
6.2.3.7
RIP
ASSAY
(=
RNA-IP)
97
6.2.3.8
ELISA
98
6.2.3.9
FLOW CYTOMETRY
99
6.2.3.10
IMMUNOFLUORESCENCE
STAINING
100
6.2.3.11
FAST
PROTEIN
LIQUID
CHROMATOGRAPHY
(FPLC)
101
A)
SIZE
EXCLUSION
CHROMATOGRAPHY
(=
GEL
FILTRATION
CHROMATOGRAPHY)
101
B)
ION
EXCHANGE
CHROMATOGRAPHY
103
6.2.3.12
GENERATION
OF
MONOCLONAL
ANTIBODIES
105
7
REFERENCES
106
8
ABBREVIATIONS
121
9
CURRICULUM
VITAE
125
IV
10
PRESENTATIONS
AND
PUBLICATIONS
127
11
ACKNOWLEDGMENTS
128
V
|
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| author | Metzner, Mirjam 1979- |
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| author_role | aut |
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| ctrlnum | (OCoLC)702667406 (DE-599)DNB1009414682 |
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| dewey-hundreds | 500 - Natural sciences and mathematics |
| dewey-ones | 571 - Physiology & related subjects |
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| discipline | Biologie |
| format | Thesis Book |
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| genre_facet | Hochschulschrift |
| id | DE-604.BV036966164 |
| illustrated | Illustrated |
| index_date | 2024-09-19T15:27:00Z |
| indexdate | 2024-11-25T17:37:10Z |
| institution | BVB |
| language | English |
| oai_aleph_id | oai:aleph.bib-bvb.de:BVB01-020880964 |
| oclc_num | 702667406 |
| open_access_boolean | 1 |
| owner | DE-384 DE-473 DE-BY-UBG DE-703 DE-1051 DE-824 DE-29 DE-12 DE-91 DE-BY-TUM DE-19 DE-BY-UBM DE-1049 DE-92 DE-739 DE-898 DE-BY-UBR DE-355 DE-BY-UBR DE-706 DE-20 DE-1102 DE-29T |
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| physical | V, 128 S. Ill., graph. Darst. 21 cm |
| psigel | ebook |
| publishDate | 2010 |
| publishDateSearch | 2010 |
| publishDateSort | 2010 |
| record_format | marc |
| spellingShingle | Metzner, Mirjam 1979- Complex formation of AID and its function in innate immunity |
| subject_GND | (DE-588)4113937-9 |
| title | Complex formation of AID and its function in innate immunity |
| title_auth | Complex formation of AID and its function in innate immunity |
| title_exact_search | Complex formation of AID and its function in innate immunity |
| title_full | Complex formation of AID and its function in innate immunity vorgelegt von Mirjam Metzner |
| title_fullStr | Complex formation of AID and its function in innate immunity vorgelegt von Mirjam Metzner |
| title_full_unstemmed | Complex formation of AID and its function in innate immunity vorgelegt von Mirjam Metzner |
| title_short | Complex formation of AID and its function in innate immunity |
| title_sort | complex formation of aid and its function in innate immunity |
| topic_facet | Hochschulschrift |
| url | https://open.fau.de/handle/openfau/1513 https://nbn-resolving.org/urn:nbn:de:bvb:29-opus-22025 http://d-nb.info/1009649892/34 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=020880964&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
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