Global SEC-PCP-SILAC mapping reveals protein complexes mediating NF-κB activation in breast cancer
| Autoři | |
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| Rok publikování | 2022 |
| Druh | Konferenční abstrakty |
| Fakulta / Pracoviště MU | |
| Citace | |
| Popis | NF-?B has essential role in immune response and is associated with lymph node metastasis of luminal A breast tumors [1]. Analysis of protein interactome and its changes in response to NF-?B modulation could uncover pro-metastatic mechanisms related to NF-?B. We apply metabolic isotope labeling SILAC, size exclusion chromatography (SEC) and protein correlation profiling (PCP) [2] to construct a network of interactome rearrangement in response to NF-?B modulation in MCF-7 breast cancer cells. We generated two co-fractionation datasets consisting of 80 fractions from SILAC-labeled and 80 fractions from label-free native MCF-7 lysates with inhibited or native NF-?B activity. LC-MS/MS analysis of SEC fractions using Orbitrap Lumos and Bruker Impact II mass spectrometers quantified 3308 and 5460 protein groups in SILAC and label-free datasets, respectively (FDR = 0.01). Interactome reconstruction using PrinCE [3] detected 7568 interactions among 1520 proteins. Co-elution of subunits of known complexes, such as ribosome, proteasome and MCM, was observed. Modulation of NF-?B was linked to interactome changes of proteins involved in immune response, cell cycle and DNA replication. NF-?B factor RELA interacted with proteins co-eluting with activators of NF-?B and these interactions were modulated by NF-?B inhibition. Our interaction network represents a complex insight into dynamics of MCF-7 protein interactome associated with NF-?B pathway and could serv e as a basis for future studies characterizing NF-?B in breast cancer.This work was supported by Ministry of Health of the Czech Republic (grant No. NU22-08-00230) and by the project National Institute for Cancer Research (Programme EXCELES, ID Project No. LX22NPO5102) - Funded by the European Union - Next Generation EU. |
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