The interferon signalling system elicits a robust cytokine response against a wide range of environmental pathogenic and internal pathological signals, leading to induction of a subset of interferon-induced proteins. We applied DSS (disuccinimidyl suberate) mediated cross-linking mass spectrometry (CLMS) to capture novel protein-protein interactions within the realm of interferon induced proteins. In addition to the expected interferon-induced proteins, we identified novel inter- and intra-molecular cross-linked adducts for the canonical interferon induced proteins, such as MX1, USP18, OAS3, and STAT1. We focused on orthogonal validation of a cohort of novel interferon-induced protein networks formed by the HLA-A protein (H2BFS-HLA-A-HMGA1) using co-immunoprecipitation assay, and further investigated them by molecular dynamics simulation. Conformational dynamics of the simulated protein complexes revealed several interaction sites that mirrored the interactions identified in the CLMS findings. Together, we showcase a proof-of-principle CLMS study to identify novel interferon-induced signaling complexes and anticipate broader use of CLMS to identify novel protein interaction dynamics within the tumour microenvironment.

• MeSH
• HLA antigeny MeSH
• HLA-A antigeny MeSH
• hmotnostní spektrometrie metody   MeSH
• interferony * MeSH
• lidé MeSH
• proteiny * chemie   MeSH
• reagencia zkříženě vázaná chemie   MeSH
• thiolesterasa ubikvitinu MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• HLA antigeny MeSH
• HLA-A antigeny MeSH
• interferony * MeSH
• proteiny * MeSH
• reagencia zkříženě vázaná MeSH
• thiolesterasa ubikvitinu MeSH
• USP18 protein, human MeSH Prohlížeč

The p53 tumor suppressor is a transcription factor with roles in cell development, apoptosis, oncogenesis, aging, and homeostasis in response to stresses and infections. p53 is tightly regulated by the MDM2 E3 ubiquitin ligase. The p53-MDM2 pathway has coevolved, with MDM2 remaining largely conserved, whereas the TP53 gene morphed into various isoforms. Studies on prevertebrate ancestral homologs revealed the transition from an environmentally induced mechanism activating p53 to a tightly regulated system involving cell signaling. The evolution of this mechanism depends on structural changes in the interacting protein motifs. Elephants such as Loxodonta africana constitute ideal models to investigate this coevolution as they are large and long-living as well as having 20 copies of TP53 isoformic sequences expressing a variety of BOX-I MDM2-binding motifs. Collectively, these isoforms would enhance sensitivity to cellular stresses, such as DNA damage, presumably accounting for strong cancer defenses and other adaptations favoring healthy aging. Here we investigate the molecular evolution of the p53-MDM2 system by combining in silico modeling and in vitro assays to explore structural and functional aspects of p53 isoforms retaining the MDM2 interaction, whereas forming distinct pools of cell signaling. The methodology used demonstrates, for the first time that in silico docking simulations can be used to explore functional aspects of elephant p53 isoforms. Our observations elucidate structural and mechanistic aspects of p53 regulation, facilitate understanding of complex cell signaling, and suggest testable hypotheses of p53 evolution referencing Peto's Paradox.

• Klíčová slova
• Loxodonta africana, Peto’s Paradox, intrinsic specificity, lifespan, model, molecular evolution, p53 retrogenes, structural variations,
• MeSH
• geny p53 MeSH
• nádorový supresorový protein p53 genetika   metabolismus   MeSH
• nádory * genetika   MeSH
• protein - isoformy genetika   metabolismus   MeSH
• protoonkogenní proteiny c-mdm2 genetika   metabolismus   MeSH
• sloni * genetika   metabolismus   MeSH
• ubikvitinace MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• nádorový supresorový protein p53 MeSH
• protein - isoformy MeSH
• protoonkogenní proteiny c-mdm2 MeSH