Functions of Human Papillomavirus E5 oncogene in epithelial cells and in the onset of cervical cancer
نویسندگان
چکیده
..................................................................................................................................8 2. REVIEW OF THE LITERATURE ...................................................................................... 13 2.1. HUMAN PAPILLOMAVIRUSES (HPVS) ........................................................................... 13 2.1.1. Genome and life cycle of HPVs......................................................................... 13 2.1.2. Role of HPV proteins ........................................................................................... 15 2.1.2.1. HPV proteins involved in replication and transcription ............................. 15 2.1.2.2. HPV oncogenes................................................................................................. 16 2.2. HPVS AND DISEASE MECHANISMS ................................................................................ 20 2.2.1. HPV-related diseases .......................................................................................... 20 2.2.2. Cellular transformation by HPV ......................................................................... 21 2.2.3. HPV and cellular microRNAs ............................................................................. 23 2.2.4. HPV and cancer .................................................................................................... 24 2.2.5. Cervical cancer screening .................................................................................. 26 2.2.6. Vaccines against HPV infection ........................................................................ 27 2.3. ALTERATIONS IN CELL ADHESION AND CELL MOTILITY IN CANCER ............................ 28 2.4. ROLE OF EZRIN IN CELL ADHESION, CELL MOTILITY AND CARCINOGENESIS............ 30 2.5. MOLECULAR PROFILING BY ARRAY TECHNOLOGIES ................................................... 31 3. AIMS OF THE STUDY ........................................................................................................ 35 4. MATERIALS AND METHODS .......................................................................................... 36 4.1. CELL CULTURE (I-III)....................................................................................................... 36 4.2. RNA EXTRACTION (I, II) ................................................................................................. 37 4.3. MICROARRAYS (I, II)....................................................................................................... 37 4.4. QUANTITATIVE RT-PCR (I, II)....................................................................................... 38 4.5. WESTERN BLOTTING (I, II) ............................................................................................. 38 4.6. WOUND HEALING ASSAY (I) ........................................................................................... 39 4.7. IMMUNOHISTOCHEMISTRY (II)........................................................................................ 40 4.8. PLASMIDS AND TRANSFECTIONS (III)............................................................................ 40 4.9. IMMUNOFLUORESCENCE (III) .........................................................................................41 4.10. STATISTICAL METHODS (I, II) .......................................................................................42 4.11. MICRORNA TARGET PREDICTION (II) .........................................................................43 5. RESULTS AND DISCUSSION ..........................................................................................44 5.1. EFFECT OF HPV-16 E5 PROTEIN ON CELLULAR GENE EXPRESSION (I, II) ..............44 5.1.1. HPV E5 alters expression of extracellular matrix genes ..............................46 5.1.2. HPV E5-induced modifications in the PI3K/Akt pathway and immune response .............................................................................................................................47 5.2. EPITHELIAL CELL ADHESION AND CELL MOTILITY (I III).............................................49 5.2.1. Alterations in expression of cell adhesion and motility-related genes due to HPV 16 E5 expression ................................................................................................49 5.2.2. Regulation of ezrin localization and function in epithelial cells (III)............52 5.2.3. HPV 16 E5 enhances cell motility in vivo (I) ...................................................53 5.3. MICRORNA PROFILING IN EPITHELIAL CELLS EXPRESSING HPV-16 E5 ONCOGENE (II) ..............................................................................................................................................54 5.3.1. HPV E5 is involved in pathogenesis of HPV infection by repressing miR203 (II) .................................................................................................................................57 6. CONCLUDING REMARKS.................................................................................................58 7. ACKNOWLEDGEMENTS...................................................................................................59 8. REFERENCES.......................................................................................................................61
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