S-Space College of Natural Sciences (자연과학대학) Program in Genetic Engineering (협동과정-유전공학전공) Theses (Ph.D. / Sc.D._협동과정-유전공학전공)
A study on the FADD-mediated necrosis in response to hypoxia-ischemia
저산소-허혈에서 FADD에 의한 Necrosis조절에 관한 연구
- Choi Seon-Guk
- 자연과학대학 협동과정 유전공학전공
- Issue Date
- 서울대학교 대학원
- 학위논문 (박사)-- 서울대학교 대학원 : 협동과정 유전공학전공, 2015. 8. 정용근.
- Fas-associated protein with death domain (FADD) plays a key role in recruiting and activating initiator caspases in the extrinsic apoptosis. However, increasing evidence indicate that FADD is also involved in intrinsic cell death, cell proliferation and necroptosis, indicating multiple functions of FADD in diverse signaling. Here I showed that FADD is dynamically modified by SUMO2 in vitro and in vivo at multiple lysine sites K120/125/149. This SUMOylation was mediated by E3 SUMO ligase PIAS3 and reversed by SENP6 and 7. Notably, FADD SUMOylation occurred during necrosis following the treatment with high dose of calcium ionophore A23187 in HeLa cells. Calcium overload initiated FADD translocation on the mitochondria, which promoted Drp1 translocation on the mitochondria as well. Treatment with A23187 induced Drp1?mediated mitochondrial fragmentation in a FADD SUMOylation-dependent manner. The necrosis triggered by A23187 was blocked in Drp1- or FADD-deficient mouse embryonic fibroblasts and ectopic expression of SUMOylation-defective FADD 3KR mutant. I showed that Drp1 binds to SUMOylated FADD in vitro and in cells. FADD and Drp1 bound to Mff in the mitochondrial. In addition, FADD SUMOylation and its translocation on the mitochondria occurred in culture cells undergoing hypoxia-induced and calcium-dependent necrosis and in ischemic damaged core resulting from middle cerebral artery occlusion (MCAO) in the mouse brain. Strikingly, calcium overload-induced cell death was also blocked by knockdown of caspase-10 but not of caspase-8. However, A23187- and mitochondrial FADD-induced mitochondrial fragmentation drove caspase-independent necrotic death. Interestingly, caspase-10 formed a protein complex with FADD and Drp1 on the mitochondria and potentiates A23187-induced necrosis and Drp1 oligomerization. Furthermore, activity-dead caspase-10 V410I and L285F mutants, which are observed in autoimmune lymphoproliferative syndrome and inhibits TRAIL-induced apoptosis, enhanced Drp1 oligomerization. Together this study reveals a novel role of SUMO modification of FADD in the calcium stress-induced, Drp1- and caspase-10-dependent mitochondrial fragmentation and regulated necrosis, providing insight into the mechanism of hypoxic and ischemia injury in a range of pathologies.