Project description:HeLa/16E6-16E7 cells after expression of wild-type E2 compared to an E2 DNA-binding mutant Keywords: Wild-type E2 Infection vs. Mutant E2 Infection
Project description:To get insights into the function of RPAP3, we characterized its partners by performing a proteomic analysis in human cells. We fused the Cterm part of RPAP3 domain to GFP and stably expressed it in HeLa cells using site-specific integration with the Flp-In system. Following differential labeling of GFP-RPAP3-Cter and control cells with isotopically labelled amino-acids (SILAC), whole cell extracts were immuno-precipitated (IP) with anti-GFP antibodies and immunoprecipitates were subjected to quantitative mass-spectrometry analysis. Similar experiments were then performed on two mutated form of RPAP3 that can no longer with the two essential AAA+ ATPases RUVBL1/RUVBL2, important partners of Wild type RPAP3 Cterm. 3 batchs of SILAC analysis K0R0 control Hela H9 vs K4R6 x-FLAG- GFP- RPAP3 Cterm Wild type K0R0 control Hela H9 vs K4R6 x-FLAG- GFP- RPAP3 Cterm mutant 1 R623A-M626A K0R0 control Hela H9 vs K4R6 x-FLAG- GFP- RPAP3 Cterm mutant 2 F630A-S632A
