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dc.description.abstractTAp63α, a p63 isoform and member of the p53 tumor suppressor family, is highly expressed exclusively in female germ cells. TAp63 is a critical mediator of the DNA damage response in female germ cells, phosphorylated upon DNA damage and induces oocyte cell death. IR-induced TAp63α phosphorylation leads to TAp63α transcriptional activation after which TAp63α appears to undergo degradation. In an attempt to search for important phosphorylation sites required for TAp63α transcriptional activation and degradation, scanning of p53 and TAp63α protein sequence revealed Ser15 site of p53, known to be important for p53 degradation by Mdm2, is conserved with Ser12 of TAp63α. However, unlike IR-induced phosphorylation of p53 Ser15, which blocks p53 binding to Mdm2 and leads to p53 stabilization, my results suggest IR-induced phosphorylation of Ser12 of TAp63α leads to TAp63α degradation, in the exact opposite manner of p53 regulation. These results interestingly suggest functional divergence of conserved p53 and TAp63α phosphorylation sites. These results led me to study further the mechanism by which TAp63α may be regulated differently from p53 and also to contemplate the physiological necessity for IR-induced TAp63α degradation vs. IR-induced p53 stabilization during oocyte development. During embryonic stages of oocyte development, oocytes generate hundreds of self-inflicted DNA double strand breaks in order to initiate meiotic recombination. I propose during this critical period of meiotic recombination, TAp63α degradation may afford oocytes a brief period of clemency from death so that meiotic recombination may proceed while death is abated. After meiotic recombination, when meiotic DNA DSBs have undergone repair, TAp63α would no longer be degraded allowing TAp63α levels to accumulate in oocytes. Only after oocytes accumulate sufficient TAp63α expression levels may IR induce TAp63α autofeedback phosphorylation and cell death discovered previously to preserve the genomic integrity of the female germline.;TAp63α는 p53 family member protein 인 p63의 isoform 중의 하나입니다. P53의 경우에는 체세포에서 암 억제 유전자로 작용하지만, TAp63Α의 경우에는 여성의 난모세포에서 DNA 손상으로 인한 세포사멸을 유도하는 중요한 매개체 입니다. 본 연구실의 최근 결과에 따르면, TAp63α은 방사선에 의한 세포사멸의 분자적 메커니즘과 더불어 TAp63α의 방사선에 의한 TAp63α의 분해를 통해서 발현량을 조절하는 메커니즘과 관련된 연구를 진행하였습니다. TA 도메인에 존재하는 Ser12 인산화 위치는 p53의 TA 도메인에 존재하는 Ser15 인산화 위치와 보존된 위치입니다. 이 Ser15 인산화 위치는 기존에 낮게 발현하는 p53의 발현량을 증가시켜주는데, TAp63α에서는 단백질 분해라는 반대되는 결과를 얻을 수 있습니다. 저희는 이러한 반대되는 결과가 Mdm2 와 관련이 되어있는지도 살펴보았습니다. Mdm2와의 결합에 필요한 위치인 FWL 위치를 이용해서 실험을 진행하였습니다. 이 결과들을 통해서 Ser12 위치에 인산화가 된 TAp63α는 Mdm2와의 결합이 가능할 수 있다는 가능성을 확인 하였습니다. 위의 결과들은 방사선에 의해 유도된 TAp63α의 Ser12 위치의 인산화를 통해서 난모세포의 성정과정에서 배아기 단계에서는 세포 생존을 조절하고 그 후에는 적절한 세포사멸을 일으킬 수 있게 하는 메커니즘이라고 제시하고 있습니다.-
dc.description.tableofcontentsI. INTRODUCTION 10 II. MATERIALS AND METHODS 27 1. Cell culture 27 2. Chemicals and γ-irradiation treatment 27 3. Gene DNA transfection 28 4. Lentivirus production and viral cell transduction 29 5. Protein Concentration by Brad-ford assay 30 6. SDS PAGE and western blotting 30 7. Immunoprecipitation 31 8. Immunofluorescence 31 III. RESULTS 33 1. IR-induced degradation of the TAp63α involves Ser12 phosphorylation 33 2. IR appears to induce TAp63α phosphorylation pulses 40 3. ATM and CHK2 kinase activity are required for TAp63α degradation 42 4. TAp63α and p53 are phosphorylated by the same upstream kinases but, yield opposite outcomes. 47 5. Transcriptional activity required for TAp63α degradation 51 6. The possibility of the interaction between TAp63α and Mdm2 54 IV. DISCUSSION 56 V. REFERENCES 58 VI. 국문 초록 63-
dc.format.extent2180755 bytes-
dc.publisher이화여자대학교 대학원-
dc.titleDNA damage-induced phosphorylation of TAp63α Ser12 conserved with p53 Ser15 mediates TAp63α degradation rather than stabilization-
dc.typeMaster's Thesis-
dc.format.page64 p.-
dc.identifier.major대학원 바이오융합과학과- 2-
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