Title Page
Abstract
Contents
List of Abbreviations 14
Introduction 15
Materials and Methods 19
Results 35
Discussion 86
Conclusion 91
References 93
국문 초록 97
Table 1. PCR primers used in this study 32
Table 2. List of Antibodies 33
Table 3. Comparison of clinical factors between two variants in the rt269 codon (KU cohort) 84
Table 4. Comparison of clinical factors between two variants in the rt269 codon (SNU cohort) 85
Fig. 1. rt269L-infected cells showed improved mitochondrial maintenance with a high rate of functional mitochondria and biogenesis 37
Fig. 2. Confocal microscopy images showing HepG2 cells transfected with mock, rt269L, or rt269I HBV followed TMRM staining indicating mitochondrial outer... 38
Fig. 3. rt269L-infected cells showed improved mitochondrial bioenergetics with a high rate of functional mitochondria and biogenesis 39
Fig. 4. rt269L induced increased mtDNA gene transcription as well as increased PGC-1α protein expression 41
Fig. 5. rt269L activates the PERK-mediated UPR pathway, but an impaired UPR is shown in rt269I-infected cells. 44
Fig. 6. Immunohistochemical analysis of p-eIF2α expression (red arrow) in paraffin-embedded liver tissues 45
Fig. 7. rt269L activates PERK-peIF2α-ATF4 signaling 46
Fig. 8. rt269L genotype C HBV infection led to enhanced autophagy induction 49
Fig. 9. rt269L genotype C HBV infection led to enhanced autophagy induction 51
Fig. 10. EGFP-LC3 assay with flow cytometry 52
Fig. 11. Enhanced autophagy in rt269L HBV infection 53
Fig. 12. Comparison of autophagy/mitophagy induction in genotype A and C HBV infection 55
Fig. 13. HBx in rt269L strongly interact with PI3KC3 (VPS34) 56
Fig. 14. Immunofluorescence for the colocalization of autophagosomes with mitochondria 57
Fig. 15. rt269I induces caspase activation and cell death by releasing cytochrome c 61
Fig. 16. rt269I HBV induces caspase activation and cell death 62
Fig. 17. Detection of intracellular reactive oxygen species in hepatocytes transfected with rt269L or rt269I HBV 63
Fig. 18. rt269I induces caspase activation and cell death 64
Fig. 19. Cell death in liver tissues of mice hydrodynamically injected with the mock, rt269L, or rt269I vector was detected by TUNEL assay 65
Fig. 20. rt269L leads to increased HBx protein stability 67
Fig. 21. rt269L leads to increased HBx protein stability in both genotype 68
Fig. 22. rt269L type HBV infection in both genotype C and A activated phospho-Akt and phospho-PI3K signals 70
Fig. 23. rt269L type HBV infection activated phospho-mTOR and phospho-PI3K signals in HBx protein dependent manner 71
Fig. 24. rt269L attenuates HBx degradation through ubiquitination of HBx 75
Fig. 25. Site-directed mutagenesis to introduce a stop codon downstream of the Polymerase start region 77
Fig. 26. HBV Pol genome and the proteins it encodes are critical for HBx production and ubiquitination 78
Fig. 27. Site-directed mutagenesis to introduce a stop codon upstream of the rt269 region 79
Fig. 28. Elevated hepatic oxidative damage in rt269I HBV infection and its clinical implications 83
Schematic figure 92