Investigation of interaction partners of NLRP13

Abstract

NOD-like receptors (NLRs) are the cytoplasmic members of pattern recognition receptor family, broadly with functions in inflammasome formation, signal transduction, transcription activation, cell death, reproduction, and embryonic development. NLRP13 is a member of NLRs containing PYRIN domain as an effector domain. Its expression is conserved in primates and various mammals, but not in rodents. It is a novel protein and there is not a single dedicated article in the literature. Its information can only be achieved from a limited number of sources and the studies of our former lab members. Based on its expression in oocytes, it is suggested that NLRP13 is a maternal-effect gene. We have shown that NLRP13 is involved in inflammasome formation. After LPS/ATP treatment or P. aeruginosa infection, NLRP13 is upregulated in THP-1 monocytes, and proinflammatory cytokine secretion increases in stable THP- 1 macrophages. NLRP13 is cleaved by Caspase-8 activated upon Fas/FasL-mediated FADD recruitment, and the cleaved C-terminal of NLRP13 is partly localized to mitochondria. NLRP13 do not directly involve in pyroptosis via Caspase-8, and PARP-1 cleavage, an apoptosis marker, is decreased in NLRP13-stable THP-1 cells after Caspase-8 is inhibited. In this thesis, the main aim of the study is to identify the novel interaction partners of NLRP13 so that to propose a notion about its role in signaling pathways and its molecular function. To achieve this goal, NLRP13- FLAG-stable THP-1 and Tera-2 cell lines were generated. Co-immunoprecipitation of NLRP13 from stable THP-1 monocytes, LPS/ATP-treated stable THP-1 macrophages and stable Tera-2 cells was performed with FLAG antibody followed by mass spectrometry. Due to various glitches, the results of repeated experiments could not be reached. However, the analysis of the obtained immunoprecipitates continues.