Title Page
ACKNOWLEDGEMENT
CONTENTS
ABBREVIATION 7
ABSTRACT 8
I. Introduction 9
II. Materials and Methods 13
II.1. Bioinformatics 13
II.2. Plasmids, Mutant Constructs and Knockdown Constructs 13
II.3. Cell Culture and Transfection 17
II.4. Northern Blot Analysis 17
II.5. In Vitro and In Vivo Deubiquitinating Activity Assays 17
II.6. Immunofluorescence and Immunohistochemistry 18
II.7.Antibody production 19
II.8. Immunoprecipitation and Immunoblotting 19
III. Results 21
III.1. Sequence analysis 21
III.2. Expression analysis of Usp19 26
III.3. Deubiquitinating activity of Usp19 28
III.4. Interaction between Usp19 and Hsp90 30
III.5. Overexpression and Knockdown of Usp19 33
III.6. Localization of Usp19 35
IV. Discussion 39
References 41
Summary in Korean 48
TABLE 1. Primers used for subcloning of Usp19 deletion constructs 16
Figure 1. Ubiquitin-proteasome system and the function of deubiquitinating enzymes 10
Figure 2. The Usp19 deletion constructs 15
Figure.3. The human and mouse Usp19 cDNA 25
Figure.4. Expression analysis of Usp19 27
Figure.5. Deubiquitinating activity of Usp19 29
Figure.6. Interaction between Usp19 and Hsp 90 32
Figure.7. Ablation of Usp19 has no effect on Hsp90 and its clients 34
Figure.8. Co-localization of Usp19 with Hsp90 and localization of endogenous Usp19 37