Investigation of the SUMO-interacting motif of p38

Autor: Tse-yu Lin, 林澤宇
Rok vydání: 2015
Druh dokumentu: 學位論文 ; thesis
Popis: 103
SUMOylation is a posttranslational modification by the small ubiquitin-related modifier (SUMO), SUMOylation plays important roles in the regulation of diverse cellular processes. SUMO can interact non-covalently with proteins through the SUMO-interacting motif (SIM). The p38 mitogen-activated protein kinases are a class of mitogen-activated protein kinases that are responsive to stress stimuli. The phosphorylation-dependent nuclear translocation of p38 is a common phenomenon when cells are stimulated with various stresses. Our previous studies demonstrated that overexpressing RFP-SUMO-1/2 caused endogenous p-p38 co-localized with RFP-SUMO-1/2 in nuclear. Our previous studies also demonstrated that p38 cannot be SUMOylated, but it can interact with SUMO-1/2 in yeast two-hybrid. We speculated that p38 has SIM. In this study, we focused on identification of the SIM of p38 and whether the SIM regulates the nuclear translocation of p38 and p38-mediated apoptosis. In immunocytochemistry, we found that wild type p38 and p-p38 co-localized with RFP-SUMO-1/2 in nuclear. Single SIM mutant (SIM-1*, SIM-2* and SIM-3*) p38 and p-p38 also co-localized with RFP-SUMO-1/2 in nuclear. In pull-down assay, results indicated that SIM-3 was essential for SUMO-interacting of p38, SIM-1 and SIM-2 also involved in SUMO-interacting of p38, and the interacting through SIM-3 was stronger. Interacting of SUMO-2 to p38 was stronger than interacting of SUMO-1 to p38. In MTT assay, overexpression of wild type p38 and SUMO-1/2 decreased cell survival rate of AGS cells. SIM-1*, SIM-2* and SIM-3* all inhibited the decreases of the cell survival rate caused by the overexpression of p38 and SUMO-1/2. We found that p38 can interact with SUMO-1/2 through the SIM, and SIM mediates the p38-mediated apoptosis of AGS cells.
Databáze: Networked Digital Library of Theses & Dissertations
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