Towards understanding the pathogenesis of polyglutamine disorders [Elektronische Ressource] : mRNA and protein expression, proteolytic cleavage and proteasomal degradation / vorgelegt von Katrin S. Lindenberg

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• Publié par: universitat_ulm
• Publié le: 01 janvier 2002
• Nombre de lectures: 37
• Poids de l'ouvrage: 59 Mo

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UNIVERSITÄT ULM
FAKULTÄT FÜR NATURWISSENSCHAFTEN








Towards understanding the pathogenesis of polyglutamine
disorders: mRNA and protein expression, proteolytic
cleavage and proteasomal degradation






Dissertation zur Erlangung des Doktorgrades Dr. rer. nat.
der Fakultät für Naturwissenschaften
der Universität Ulm



vorgelegt von
Katrin S. Lindenberg
geboren in Freiburg im Breisgau



Ulm im November 2002




























Amtierender Dekan der Fakultät für Naturwissenschaften: Prof. Dr. W. Witschel
Erstgutachter: Prof. Dr. K.-D. Spindler
Zweitgutachter:
Tag der Promotion:






























Today is the tomorrow you worried about yesterday.
(Unknown)

































Meinen Geschwistern Robert und Saskia und
meinen Neffen Simon und Johannes gewidmet 1 INTRODUCTION 1
1.1 TRIPLET REPEAT DISORDERS 1
1.2 DISEASES INVOLVING NON-CODING REPEATS 2
1.3 DISEASES INVOLVING CODING REPEATS 4
1.3.1 THE POLYALANINE DISORDERS
1.3.2 THE POLYGLUTAMINE DISORDERS
1.3.2.1 Spinobulbar Muscular Atrophy 6
1.3.2.2 Huntington´s Disease 6
1.3.2.3 The Spinocerebellar Ataxias 7
1.3.2.4 Dentatorubral-Pallidoluysian Atrophy 11
1.4 MOLECULAR PATHOGENESIS OF POLYGLUTAMINE DISORDERS 12
1.4.1 FORMATION OF NEURONAL INTRANUCLEAR INCLUSIONS
1.4.2 CLEAVAGE OF THE MUTANT POLY-Q PROTEINS RESULTS IN TOXIC FRAGMENTS 13
1.4.3 IMPAIRMENT OF PROTEASOMAL DEGRADATION? 14
1.4.4 DO MOLECULAR CHAPERONES PLAY A ROLE IN POLY-Q DISEASES? 15
2 RESULTS 16
2.1 EXPRESSION ANALYSIS OF ATAXIN-7 MRNA AND PROTEIN IN HUMAN BRAIN 16
2.1.1 EXPRESSION OF ATAXIN-7 MRNA IN NORMAL HUMAN BRAIN 16
2.1.2 LOCALIZATION OF ATAXIN-7-IMMUNOREACTIVITY IN NORMAL HUMAN BRAIN 18
2.1.3 IMMUNOLOCALIZATION IN PERIPHERAL ORGANS 20
2.2 TRANSGENIC MOUSE MODELS FOR SCA7 23
2.2.1 MICE EXPRESSING HUMAN MUTANT ATAXIN-7 UNDER THE CONTROL OF PDGF B
DEVELOP A PROGRESSIVE ATAXIC PHENOTYPE 23
2.2.2 MOUSE MODELS FOR SCA7 WITH EXPRESSION OF ATAXIN-7 RESTRICTED TO RETINA
OR PURKINJE CELLS 24
2.2.3 MUTANT ATAXIN-7 ACCUMULATES AND FORMS NUCLEAR INCLUSIONS IN
TRANSGENIC MICE 26
2.2.4 OVEREXPRESSION OF MUTANT ATAXIN-7 INDUCES NEURODEGENERATION OF
TARGETED CELLS 29
2.2.5 MUTANT ATAXIN-7: ACCUMULATION OF N-TERMINAL FRAGMENTS AND LOSS OF C-
TERMINAL IMMUNOREACTIVITIES, PROCESSING AND CELLULAR RESPONSES TO
AGGREGATING N-TERMINAL FRAGMENTS2.2.6 UBIQUITIN AND A DISTINCT SET OF CHAPERONE AND PROTEASOME SUBUNITS ARE
RECRUITED INTO NIS 31
2.3 COMPOSITION OF POLY-Q AGGREGATES I: CLEAVAGE OF MUTANT HUNTINGTIN IN
HUMAN HD BRAIN 32
2.4 COMPOSITION OF POLY-Q AGGREGATES II: PROTEASOMAL SUBUNITS AND
CHAPERONES IN HUMAN SCA3 BRAIN 35
2.4.1 FREQUENCY OF NEURONAL INTRANUCLEAR INCLUSIONS IN PONTINE NEURONS IN
SCA3
2.4.2 ANTIBODIES AGAINST HEAT SHOCK PROTEIN STAIN NEURONAL INTRANUCLEAR
INCLUSIONS IN SCA3 36
2.4.3 A SMALL SUBSET OF NEURONAL INTRANUCLEAR INCLUSIONS IN SCA3 DISPLAY IR
AGAINST THE 20S CATALYTIC CORE, BUT MOST ATAXIN-3 INCLUSIONS EXHIBIT 19S
AND 11S-PROTEASOME IR 37
2.4.4 REDISTRIBUTION OF 26S PROTEASOMAL SUBUNITS IN SCA3 PONS 39
2.5 PROTEASOMAL DEGRADATION IN THE PRESENCE OF MUTANT HTT PROTEIN: STUDIES
IN A CELLULAR MODEL FOR HD 40
2.5.1 GREEN/ RED FLUORESCENT PROTEIN (GFP-/RFP) BASED REPORTER SYSTEMS FOR
QUANTIFYING PROTEASOMAL ACTIVITY IN A CELLULAR MODEL FOR HD 40
2.5.2 ALTERED PROTEASOMAL ACTIVITY IN CELLS EXPRESSING MUTANT HUNTINGTIN 44
3 DISCUSSION 46
3.1 WIDESPREAD EXPRESSION OF ATAXIN-7 DOES NOT ACCOUNT FOR THE FOCAL
PATHOLOGY IN SCA7 46
3.2 MOUSE MODELS IN SCA7: PATHOLOGICAL ASPECTS IN COMPARISON TO OTHER
POLYGLUTAMINE DISEASE 47
3.3 UNFOLDING AND DEGRADATION: THE PROTEASOME PATHWAY AND CHAPERONES IN
POLY-Q-DISORDERS 53
4 SUMMARY 57
5 ZUSAMMENFASSUNG 59
6 REFERENCES 61