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Regulation of DNA-PKcs-mediated immune signaling by the E3 ligase MARCH5 in mitochondria-damaged cells
| DC Field | Value | Language |
|---|---|---|
| dc.contributor.advisor | 이, 재호 | - |
| dc.contributor.author | 허, 준 | - |
| dc.date.accessioned | 2023-11-16T05:43:57Z | - |
| dc.date.available | 2023-11-16T05:43:57Z | - |
| dc.date.issued | 2023 | - |
| dc.identifier.uri | http://repository.ajou.ac.kr/handle/201003/26852 | - |
| dc.language.iso | en | - |
| dc.title | Regulation of DNA-PKcs-mediated immune signaling by the E3 ligase MARCH5 in mitochondria-damaged cells | - |
| dc.title.alternative | 미토콘드리아 손상 세포에서 E3 연결효소 MARCH5에 의한 DNA-PKcs 매개 면역신호의 조절 | - |
| dc.type | Thesis | - |
| dc.identifier.url | http://dcoll.ajou.ac.kr:9080/dcollection/jsp/common/DcLoOrgPer.jsp?sItemId=000000032854 | - |
| dc.subject.keyword | Mitochondrial DNA damage | - |
| dc.subject.keyword | Pattern recognition receptor | - |
| dc.subject.keyword | Type I interferon response | - |
| dc.subject.keyword | Inflammatory disorder | - |
| dc.subject.keyword | E3 ligase | - |
| dc.subject.keyword | Ubiquitination | - |
| dc.description.degree | Doctor | - |
| dc.contributor.department | 대학원 의생명과학과 | - |
| dc.contributor.affiliatedAuthor | 허, 준 | - |
| dc.date.awarded | 2023 | - |
| dc.type.local | Theses | - |
| dc.citation.date | 2023 | - |
| dc.embargo.liftdate | 9999-12-31 | - |
| dc.embargo.terms | 9999-12-31 | - |
| dc.description.tableOfContents | I. INTRODUCTUON 1
II. MATERIALS AND METHOD 9 A. Plasmids and chemicals 9 B. Cell culture 9 C. Transfection and RNA interference 9 D. Generation of March5 KO RAW264.7 10 E. Western blot analysis 10 F. Mitochondrial fractionation 12 G. Pull-down assay 12 H. Ubiquitination assay 12 I. Immunofluorescence 13 J. Reverse transcription-quantitative polymerase chain reaction 13 K. Mitochondrial DNA copy number assay 15 L. ATP assay 15 M. Statistical analysis 16 III. RESULTS 17 A. Establishment of FokI endonuclease systems to generate mtDNA double-strand breaks 17 B. MTERF1-FokI-mCherry and TFAM-FokI-mCherry localize to mitochondria 19 C. Fluorescence signal of the mtDNA is reduced in an activity-dependent manner at the mt-FokI endonuclease 21 D. Specific targeting of mtDNA by the FokI endonuclease reduces the amount of mtDNA 23 E. Mt-FokIs do not affect nuclear DNA damage response and ATP synthesis upon mtDNA degradation 26 F. A type I interferon response is triggered by mtDSBs even in the absence of cGAS 29 G. Mitochondrial DNA cleavage increases phosphorylation of IRF3 and HSPA8 in a variety of cell lines 32 H. DNA-PKcs mediates phosphorylation of HSPA8 and IRF3 in response to breaks in mtDNA 34 I. Cleaved mtDNA elicits DNA PKcs-mediated type I interferon response via the VDAC macropore 37 J. Mitochondrial E3 ligase MARCH5 degrades DNA-PKcs in a proteasome-dependent manner 40 K. MARCH5 binds and ubiquitylates activated DNA-PKcs 43 L. MARCH5 suppresses DNA-PKcs-mediated type I interferon response 46 M. LPS treatment elicits DNA-PKcs-dependent type I interferon response in murine macrophages 48 N. MARCH5 prevents prolonged type І Interferon response in murine macrophage stimulated by LPS 51 IV. DISCUSSION 54 REFERENCE 57 국문요약 64 | - |
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