Dissertation
Dissertation > Medicine, health > Basic Medical > Pathology > Pathophysiology

Effective of the Interaction between NDP52 and TRAF6 and TRAFs on NF-κB Signal Pathway

Author WangMin
Tutor WangSiYing;ZhengHong;YangXiaoMing
School Anhui Medical University,
Course Pathology and Pathophysiology
Keywords NDP52 TRAF6 NF-κB protein-protein interaction
CLC R363
Type Master's thesis
Year 2011
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Nuclear factor-κB (NF-κB) is a ubiquitously expressed family of transcription factors,plays a centrol role in regulating the expression of a wide variety of genes involved in diverse physiological and pathological processes,including cell growth/death,immune/inflammatory responses,stress responses,and oncogenesis. A key event leading connecting extracellular stimuli to NF-κB activation is the regulation of TRAF6 (tumor necrosis factor receptor associated factor 6) activity.Therefore,we identified NDP52(nuclear dot protein 52) as TRAF6-interacting partner by yeast two-hybrid screen of fetal liver cDNA library using TRAF6 as a bait. NDP52 (nuclear dot protein 52),a cytoplasmic protein,was localized in Nuclear dots (NDs) originally.NDP52 is primarily expressed in Hella cells. NDP52 functions as a receptor that recognizes ubiquitinated bacteria and coordinates their destruction by the autophagy pathway.,and NDP52 can participate antibacterial autophagy directly. At present, we have known a little about the molecular mechanism of NDP52. This study focuses on the interaction proteins of NDP52 and makes an approach to the molecular mechanism of NDP52. We intensively investigate the interactions between NDP52 and TRAF6 and its biological significance.This paper tested the unknown interaction between NDP52 and TRAF6. Firstly, we confirmed the physiological interaction between NDP52 and TRAF6 by extragenous co-immunoprecipitation (Co-IP). The interactions of NDP52 to TRAF1,TRAF3,TRAF5,TRAF6 were confirmed by co-immunoprecipitation respectively. By mating, we identify the interaction between NDP52 and TRAF3,4,6. ,and mapped NDP52-TRAF6 interacting domains by mating, we found TRAF6(1-350aa)interacted with NDP52,and NDP52(224-446aa) interacted with TRAF6 by Co-IP.Then,to test a possible involvement of NDP52 in the NF-κB activation pathway,in reporter gene assay,we found overexpressed NDP52 inhibition basal NF-κB activation,and also TNFα-induced NF-κB activity as well as its basal transcriptional activity. Subsequently,we discovered that overexpressed NDP52 inhibited TRAFs-induced NF-κB activation as well as its basal transcriptional activity.Finally,to explore the mechanism by which the interactions of NDP52 with TRAF6 down-regulate NF-κB signaling,we adopted methods of De-ubiquitinating Assays and In Vivo Ubiquitination ,and we found NDP52 inhibits NF-κB by De-ubiquitinating TRAF6.In conclusion,the interaction between NDP52 and TRAF6 and TRAFs play an important role in the NF-κB signaling,and these findings may help us to explore a new regulatory mechanism in NF-κB pathway.

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