Molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor:
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| Main Author: | |
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| Format: | Book |
| Language: | English |
| Published: |
Karlsruhe
FZKA
2003
|
| Series: | Wissenschaftliche Berichte / Forschungszentrum Karlsruhe Technik und Umwelt
6953 |
| Subjects: | |
| Online Access: | Inhaltsverzeichnis |
| Item Description: | Zugl.: Karlsruhe, Univ., Diss., 2003. - Mit dt. Zsfassung |
| Physical Description: | 90 S. Ill., graph. Darst. : 30 cm |
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| 100 | 1 | |a Hong, Wei |e Verfasser |4 aut | |
| 245 | 1 | 0 | |a Molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor |c Wei Hong. Forschungszentrum Karlsruhe GmbH |
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| 490 | 1 | |a Wissenschaftliche Berichte / Forschungszentrum Karlsruhe Technik und Umwelt |v 6953 | |
| 500 | |a Zugl.: Karlsruhe, Univ., Diss., 2003. - Mit dt. Zsfassung | ||
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TABLE
OF
CONTENTS
ABSTRACT
ZUSAMMENFASSUNG
TABLE
OF
CONTENTS
1.
INTRODUCTION
.
1
1.1
GLUCOCORTICOID
AND
GLUCOCORTICOID
RECEPTOR
.
1
1.2
REGULATION
OF
GENE
EXPRESSION
BY
GLUCOCORTICOID
.
3
1.2.1
POSITIVE
REGULATION
OF
GENE
EXPRESSION
BY
GLUCOCORTICOID
.
3
1.2.2
NEGATIVE
REGULATION
OF
GENE
EXPRESSION
BY
GLUCOCORTICOID
.
5
1.3
MODULATION
OF
THE
ACTION
OF
GR
BY
MOLECULAR
CHAPERONES
AND
COCHAPERONES
.
6
1.3.1
CHAPERONES
AND
COCHAPERONES
IN
THE
MATURATION
OF
GR
.
6
1.3.2
BAG-1
PROTEINS
.
7
1.3.3
BAG-1
PROTEINS
REGULATE
THE
NUCLEAR
RECEPTORS
.
9
AIM
.
11
2.
MATERIALS
AND
METHODS
.
12
2.1
MATERIALS
.
12
2.1.1
CHEMICALS
.
12
2.1.2
OLIGONUCLEOTIDES
.
14
2.1.2.1
DNA
OLIGONUCLEOTIDES
.
14
2.1.2.2
RNA
OLIGONUCLEOTIDES
.
15
2.1.3
PLASMIDS
AND
CONSTRUCTS
.
15
2.1.4
ENZYMES
.
*
.
16
2.1.5
ANTIBODIES
.
16
2.1.5.1
PRIMARY
ANTIBODIE
.
16
2.1.5.2
SECONDARY
ANTIBODIES
.
17
2.1.6
CELL
LINES
.
17
2.1.7
BACTERIA
.
17
2.1.8
OTHER
MATERIALS
.
18
2.2
METHODS
.
18
2.2.1
SMALL
SCALE
PLASMID
PREPARATION
FROM
BACTERIA
.
19
2.2.2
LARGE
SCALE
PLASMID
PREPARATION
.
19
2.2.3
REATRICATION
ENDONUCLEASE
DIGESTION
OF
DNA
.
19
2.2.4
SIZE
SEPARATION
OF
NUCLEIC
ACID
BY
AGAROSE
GEL
ELECTROPHORESIS
.
19
2.2.5
ISOLATION
AND
PURIFICATION
OF
DNA
FROM
AGAROSE
GEL
.
20
2.2.6
DNA
LIGATION
.
20
2.2.7
SUB-CLONING
.
.
:
.
20
2.2.8
PHENOL/CHLOROFORM
EXTRACTION
OF
NUCLEIC
ACIDS
.
20
2.2.9
PRECIPITATION
OF
NUCLEIC
ACIDS
.
'
.
20
2.2.10
DETERMINATION
OF
NUCLEIC
ACID
CONCENTRATION
.
21
2.2.11
PREPARATION
OF
CHEMICALLY
COMPETENT
BACTERIA
(E.COLI)
.
21
2.2.12
TRANSFORMATION
OF
BACTERIA
(E.COLI)
.
21
2.2.12.1_CHEMICAL
TRANSFORMATION
.
21
2.2.12.2
TRANSFORMATION
BY
ELECTROPORATION
.
22
2.2.13
POLYMERASE
CHAIN
REACTION
(PCR)
.
22
2.2.14
CLONING
INTO
PCR2.1-TOPO
TA
VECTOR
.
22
2.2.15
MANUAL
(RADIOACTIVE)
DNA
SEQUENCING
METHOD
.
23
2.2.16
RADIOACTIVE
LABELING
OF
DNA
PROBE
FOR
EMSA
.
23
2.2.17
DETERMINATION
OF
PROTEIN
CONCENTRATION
.
24
2.2.18
SDS-POLYACRYLAMIDE
GEL
ELECTROPHORESIS
(SDS-PAGE)
.
24
2.2.19
COOMASSIE
STAINING
.
25
2.2.20
WESTERN
BLOT
.
25
2.2.21
STRIPPING
WESTERN
BLOT
MEMBRANE
.
26
2.2.22
CELL
CULTURE
.
26
2.2.23
LONG
TERM
STORAGE
OF
CELLS
.
26
2.2.24
TRANSIENT
TRANSFECTION
OF
MAMMALIAN
CELLS
.
27
2.2.25
WHOLE
CELL
EXTRACT
FOR
EMSA
.
27
2.2.26
WHOLE
CELL
EXTRACT
FOR
WESTERN
BLOT
.
27
2.2.27
PREPARATION
OF
GST
AND
GST
FUSION
PROTEINS
.
27
2.2.28
CLEAVAGE
OF
THE
PROTEIN
FUSED
TO
GST
.
28
2.2.29
PREPARATION
OF
RADIOACTIVELY
LABELED
IN
VITRO
TRANSLATED
GR
AND
GR
MUTANTS
.
29
2.2.30
ATP
DEPLETION
OF
COS-7
CELLS
.
29
2.2.31
ISOLATION
OF
TOTAL
RNAFROM
CULTURED
CELLS
.
29
2.2.32
SEPARATION
OF
RNA
BY
AGAROSE
GEL
ELECTROPHORESIS
.
30
2.3
ANALYTICAL
METHODS
.
31
2.3.1
MEASUREMENT
OF
FIREFLY
LUCIFERASE
ACTIVITY
.
31
2.3.2
MEASUREMENT
OF
RENILLA
LUCIFERASE
ACTIVITY
.
31
2.3.3
GST-PULL
DOWN
ASSAY
.
31
2.3.4
ELECTROPHORESIS
MOBILITY
SHIFT
ASSAY
.
32
2.3.5
CHROMATIN
IMMUNOPRECIPITATION
.
32
2.3.6
RNAI
.
34
2.3.7
REAL-TIME
PCR
.
34
3.
RESULTS
.
36
3.1
BAG-1
M
FUNCTIONS
VIA
DISTINCT
AMINO
ACID
SEQUENCES
IN
THE
GR
.
36
3.1.1
LYSINE
496
AND
ISOLEUCINE
497
IN
THE
HINGE
REGION
OF
THE
GR
ARE
ESSENTIAL
FOR
THE
NEGATIVE
REGULATION
OF
BAG-1
M
ON
THE
TRANSA
'
CTIVATION
BY
THE
GR
.
7
.
37
3.1.2
BAG-1
M
DOES
NOT
INHIBIT
THE
DNA
BINDING
BY
THE
K476L/I497G
(GRMT2)
.
7
.
40
3.1.3.
LYSINE
496
AND
ISOLEUCINE
497
ARE
SITES
FOR
THE
INTERACTION
OF
GR
WITH
BAG-1
M
.
42
3.2
DOMAINS
OF
BAG-1
M
INVOLVED
IN
THE
PHYSICAL
CONTACT
WITH
GR
.
43
3.3
ATP
HYDROLYSIS
IS
REQUIRED
FOR
THE
REGULATION
OF
GR
ACTIVITY
BY
BAG-1
M
.
45
3.3.1
ATP
HYDROLYSIS
PROMOTES
BAG-1
M-MEDIATED
REPRESSION
OF
DNA
BINDING
BY
THE
GR
.
45
3.3.2
ATP
HYDROLYSIS
IS
NECESSARY
FOR
BAG-1
M
TO
INHIBIT
DNA
BINDING
BY
GR
.
47
3.4
GR,
BAG-1
,
MOLECULAR
CHAPERONES
ARE
PRESENT
ON
GLUCOCORTICOID
RESPONSE
ELEMENT
.
50
3.5
BAG-1
DOWNREGULATES
DNA
BINDING
BY
THE
GR
.
52
3.6
BAG-1M
IS
NOT
INVOLVED
IN
UBIQUITIN-PROTEASOME
-MEDIATED
DEGRADATION
.
55
3.7
BAG-1
M-MEDIATED
RECRUITMENT
OF
HISTONE
DEACTYLASES
.
57
3.8
EFFECT
OF
TRICHOSTATIN
A
ON
BAG-1
M-MEDIATED
REPRESSION
OF
GLUCOCORTICOID
RECEPTOR
ACTIVITY
.
61
3.9
COREPRESSORS
AND
HISTONE
DEACETYLASE
1
AND
2
ARE
NOT
ASSEMBLED
WITH
THE
BAG-1
COMPLEX
TARGETED
TO
THE
GRE
OF
HMTIIA
GENE
IN
FTO
CELLS
.
63
4.
DISCUSSION
.
66
4.1
BAG-1
M
DOWNREGULATES
THE
TRANSACTIVATION
ACTIVITY
OF
THE
GR
.
66
4.2
HSP70
IS
INVOLVED
IN
BAG-1
M-MEDIATED
INHIBITION
OF
GR
ACTIVITY
.
68
4.3
BAG-1
M
RECRUITS
HISTONE
DEACETYLASES
IN
COS-7
CELLS
.
70
4.4
DUAL
MECHANISM
FOR
MODULATION
OF
GR
ACTIVITY
BY
BAG-1
M
.
71
4.5
SUMMARY
.
72
ABBREVIATIONS
.
73
REFERENCES
.
76
ACKNOWLEDGEMENTS |
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| indexdate | 2024-08-16T01:01:06Z |
| institution | BVB |
| language | English |
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| owner | DE-12 DE-91 DE-BY-TUM |
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| physical | 90 S. Ill., graph. Darst. : 30 cm |
| publishDate | 2003 |
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| publisher | FZKA |
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| series2 | Wissenschaftliche Berichte / Forschungszentrum Karlsruhe Technik und Umwelt |
| spelling | Hong, Wei Verfasser aut Molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor Wei Hong. Forschungszentrum Karlsruhe GmbH FZKA 6953 Karlsruhe FZKA 2003 90 S. Ill., graph. Darst. : 30 cm txt rdacontent n rdamedia nc rdacarrier Wissenschaftliche Berichte / Forschungszentrum Karlsruhe Technik und Umwelt 6953 Zugl.: Karlsruhe, Univ., Diss., 2003. - Mit dt. Zsfassung Glucocorticosteroidrezeptor (DE-588)4278213-2 gnd rswk-swf Transkriptionsfaktor (DE-588)4303350-7 gnd rswk-swf Chaperonine (DE-588)4338035-9 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Glucocorticosteroidrezeptor (DE-588)4278213-2 s Chaperonine (DE-588)4338035-9 s Transkriptionsfaktor (DE-588)4303350-7 s DE-188 Forschungszentrum Karlsruhe Technik und Umwelt Wissenschaftliche Berichte 6953 (DE-604)BV010181833 6953 DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=010736858&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Hong, Wei Molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor Glucocorticosteroidrezeptor (DE-588)4278213-2 gnd Transkriptionsfaktor (DE-588)4303350-7 gnd Chaperonine (DE-588)4338035-9 gnd |
| subject_GND | (DE-588)4278213-2 (DE-588)4303350-7 (DE-588)4338035-9 (DE-588)4113937-9 |
| title | Molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor |
| title_alt | FZKA 6953 |
| title_auth | Molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor |
| title_exact_search | Molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor |
| title_full | Molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor Wei Hong. Forschungszentrum Karlsruhe GmbH |
| title_fullStr | Molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor Wei Hong. Forschungszentrum Karlsruhe GmbH |
| title_full_unstemmed | Molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor Wei Hong. Forschungszentrum Karlsruhe GmbH |
| title_short | Molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor |
| title_sort | molecular chaperones and cochaperones as regulators of the transcriptional activity of the glucocorticoid receptor |
| topic | Glucocorticosteroidrezeptor (DE-588)4278213-2 gnd Transkriptionsfaktor (DE-588)4303350-7 gnd Chaperonine (DE-588)4338035-9 gnd |
| topic_facet | Glucocorticosteroidrezeptor Transkriptionsfaktor Chaperonine Hochschulschrift |
| url | http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=010736858&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| volume_link | (DE-604)BV010181833 |
| work_keys_str_mv | AT hongwei molecularchaperonesandcochaperonesasregulatorsofthetranscriptionalactivityoftheglucocorticoidreceptor AT hongwei fzka6953 |