結(jié)核分枝桿菌PPE10（Rv0442c）通過線性泛素鏈組裝復合物HOIP-NF-κB信號傳導軸改變宿主細胞凋亡和細胞因子
發(fā)布時間：2024-06-10 23:49
結(jié)核分枝桿菌(Mycobacterium tuberculosis,簡稱結(jié)核桿菌)感染引起的結(jié)核病仍然是全世界十大高致死率疾病之一。結(jié)核桿菌成功之處在于能夠主動改變宿主信號轉(zhuǎn)導。結(jié)核桿菌基因組包含大量編碼毒力因子的基因。PE/PPE家族蛋白廣泛參與細菌-宿主互作,與結(jié)核桿菌毒力和抗原變異有關,在調(diào)控宿主細胞信號轉(zhuǎn)導、幫助細菌逃避宿主免疫殺傷過程中發(fā)揮重要作用。結(jié)核分枝桿菌PPE10(Rv0442c)屬于PPE家族,高度保守。PPE10能夠干擾細胞壁完整性,增強海分枝桿菌的毒力并減少宿主細胞泛素的募集。BCG PPE10缺失菌株抑制吞噬體酸化的能力減弱。PPE10 BCG免疫小鼠可以激發(fā)強烈的T細胞反應。為了探索PPE10在細菌-宿主相互作用中的功能,我們構(gòu)建了異源表達PPE10編碼基因Rv0442c的非致病性恥垢分枝桿菌MsPPE10。Rv0442c過表達改變了恥垢分枝桿菌MsPPE10的單菌落形態(tài),增強恥垢分枝桿菌MsPPE10生物膜形成能力。與僅攜帶空載的恥垢分枝桿菌Msvec相比,PPE10提...
【文章頁數(shù)】：84 頁
【學位級別】：博士
【文章目錄】：
Abbreviation
摘要
ABSTRACT
CHAPTER ONE LITERATURE REVIEW
1.1 HISTORY OF TUBERCULOSIS
1.2 THE BURDEN OF TUBERCULOSIS
1.3 BIOLOGICAL CHARACTERISTIC OF M.TUBERCULOSIS
1.4 CLINICAL MANIFESTATIONS OF M.TUBERCULOSIS
1.5 THE PATHOGENICITY OF M.TUBERCULOSIS
1.6 THE VIRULENCE FACTORS OF M.TUBERCULOSIS
1.7 THE IMMUNE-MODULATORY EFFECTS OF M.TUBERCULOSIS
1.8 DIAGNOSIS,TREATMENT,AND VACCINE OF TUBERCULOSIS
1.8.1 Diagnosis of tuberculosis
1.8.2 Treatment of tuberculosis
1.8.3 Vaccines of tuberculosis
1.9 M.TUBERCULOSIS PE/PPE GENES FAMILY
1.10 THE SECRETION SYSTEM AND LOCATION OF PE/PPE PROTEINS FAMILY
1.11 THE ROLE OF PE/PPE PROTEINS IN PATHOGEN-HOST INTERACTIONS
1.12 THE ROLE OF PE/PPE GENES FAMILY IN IMMUNE EVASION
1.13 INTERFERING OF PE/PPE PROTEINS IN HOST CELL DEATH FATE
1.14 THE IMPLICATION OF M.TUBERCULOSIS PE/PPE GENES FAMILY IN VACCINE
CHAPTER TWO INTRODUCTION
AIMS AND HYPOTHESES
CHAPTER THREE MATERIAL AND METHODS
3.1 MATERIAL
3.1.1 Bacterial strains,plasmids,and culture conditions
3.1.2 Culture medium and growth conditions
3.1.3 Chemicals,Reagents,Kits,Media,other
3.1.3.1 Chemicals
3.1.3.2 Reagents and Kits
3.1.3.3 Media and others
3.1.4 SDS-PAGE and Western blotting Reagents
3.1.5 Antibodies used in this study
3.1.6 Instruments and Apparatus
3.2 METHODS
3.2.1 Construction of recombinant M.smegmatis strains
3.2.2 Expression of PPE10 in M.smegmatis
3.2.3 The survival of recombinant M.smegmatis under stresses
3.2.4 Biofilm formation assay
3.2.5 Ethidium bromide accumulation/efflux assay and Nile red uptake assays
3.2.6 Infection of macrophages
3.2.7 RT-PCR assay
3.2.8 Protein analysis by Western Blotting
3.2.9 Apoptosis analysis by fluorescence microscope
3.2.10 Cell signaling pathway inhibition
3.2.11 Flow cytometry
3.2.12 Statistical analysis
CHAPTER FOUR RESULTS
4.1 M.TUBERCULOSIS RV0442C(PPE10)SUCCESSFULLY EXPRESSED IN M.SMEGMATIS
4.2 PPE10 PROMOTES THE RESISTANCE OF M.SMEGMATIS TO STRESSES
4.3 PPE10 ALTERS COLONY MORPHOLOGY AND CELL WALL PERMEABILITY
4.4 PPE10 PROMOTES THE SURVIVAL OF RECOMBINANT M.SMEGMATIS WITHIN MACROPHAGES AND MODULATES THE TRANSCRIPTION LEVELS OF CYTOKINES
4.5 PPE10 DECREASES CASPASES EXPRESSION AND ALTERS APOPTOSIS IN INFECTED MACROPHAGE
4.6 PPE10 ALTERS CYTOKINE EXPRESSION AND CASPASES VIA NF-ΚB SIGNALING PATHWAY
4.7 PPE10 REGULATES NF-ΚB PATHWAY THROUGH LUBAC AND C-IAP1
4.8 PPE10 reduces expression and phosphorylation of RelA/p65 NF-κB
CHAPTER FIVE DISCUSSION AND CONCLUSION
5.1 DISCUSSION
5.2 Conclusion
REFERENCE
Contributions
ACKNOWLEDGEMENT
本文編號：3992081
【文章頁數(shù)】：84 頁
【學位級別】：博士
【文章目錄】：
Abbreviation
摘要
ABSTRACT
CHAPTER ONE LITERATURE REVIEW
1.1 HISTORY OF TUBERCULOSIS
1.2 THE BURDEN OF TUBERCULOSIS
1.3 BIOLOGICAL CHARACTERISTIC OF M.TUBERCULOSIS
1.4 CLINICAL MANIFESTATIONS OF M.TUBERCULOSIS
1.5 THE PATHOGENICITY OF M.TUBERCULOSIS
1.6 THE VIRULENCE FACTORS OF M.TUBERCULOSIS
1.7 THE IMMUNE-MODULATORY EFFECTS OF M.TUBERCULOSIS
1.8 DIAGNOSIS,TREATMENT,AND VACCINE OF TUBERCULOSIS
1.8.1 Diagnosis of tuberculosis
1.8.2 Treatment of tuberculosis
1.8.3 Vaccines of tuberculosis
1.9 M.TUBERCULOSIS PE/PPE GENES FAMILY
1.10 THE SECRETION SYSTEM AND LOCATION OF PE/PPE PROTEINS FAMILY
1.11 THE ROLE OF PE/PPE PROTEINS IN PATHOGEN-HOST INTERACTIONS
1.12 THE ROLE OF PE/PPE GENES FAMILY IN IMMUNE EVASION
1.13 INTERFERING OF PE/PPE PROTEINS IN HOST CELL DEATH FATE
1.14 THE IMPLICATION OF M.TUBERCULOSIS PE/PPE GENES FAMILY IN VACCINE
CHAPTER TWO INTRODUCTION
AIMS AND HYPOTHESES
CHAPTER THREE MATERIAL AND METHODS
3.1 MATERIAL
3.1.1 Bacterial strains,plasmids,and culture conditions
3.1.2 Culture medium and growth conditions
3.1.3 Chemicals,Reagents,Kits,Media,other
3.1.3.1 Chemicals
3.1.3.2 Reagents and Kits
3.1.3.3 Media and others
3.1.4 SDS-PAGE and Western blotting Reagents
3.1.5 Antibodies used in this study
3.1.6 Instruments and Apparatus
3.2 METHODS
3.2.1 Construction of recombinant M.smegmatis strains
3.2.2 Expression of PPE10 in M.smegmatis
3.2.3 The survival of recombinant M.smegmatis under stresses
3.2.4 Biofilm formation assay
3.2.5 Ethidium bromide accumulation/efflux assay and Nile red uptake assays
3.2.6 Infection of macrophages
3.2.7 RT-PCR assay
3.2.8 Protein analysis by Western Blotting
3.2.9 Apoptosis analysis by fluorescence microscope
3.2.10 Cell signaling pathway inhibition
3.2.11 Flow cytometry
3.2.12 Statistical analysis
CHAPTER FOUR RESULTS
4.1 M.TUBERCULOSIS RV0442C(PPE10)SUCCESSFULLY EXPRESSED IN M.SMEGMATIS
4.2 PPE10 PROMOTES THE RESISTANCE OF M.SMEGMATIS TO STRESSES
4.3 PPE10 ALTERS COLONY MORPHOLOGY AND CELL WALL PERMEABILITY
4.4 PPE10 PROMOTES THE SURVIVAL OF RECOMBINANT M.SMEGMATIS WITHIN MACROPHAGES AND MODULATES THE TRANSCRIPTION LEVELS OF CYTOKINES
4.5 PPE10 DECREASES CASPASES EXPRESSION AND ALTERS APOPTOSIS IN INFECTED MACROPHAGE
4.6 PPE10 ALTERS CYTOKINE EXPRESSION AND CASPASES VIA NF-ΚB SIGNALING PATHWAY
4.7 PPE10 REGULATES NF-ΚB PATHWAY THROUGH LUBAC AND C-IAP1
4.8 PPE10 reduces expression and phosphorylation of RelA/p65 NF-κB
CHAPTER FIVE DISCUSSION AND CONCLUSION
5.1 DISCUSSION
5.2 Conclusion
REFERENCE
Contributions
ACKNOWLEDGEMENT
本文編號：3992081
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