ArticleThe Journal of biological chemistry2011
Activation kinetics of RAF protein in the ternary complex of RAF, RAS-GTP, and kinase on the plasma membrane of living cells: single-molecule imaging analysis.
Article in The Journal of biological chemistry, 2011. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 32 papers.
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Who cites it
32 citing papers in PubMed, 47 citations in OpenAlex.
- Targeted degradation of KRAS and induction of bystander effects by a modular bioPROTAC.Molecular therapy. Oncology · 2025Article
- Targeting c-Myc enhances immunotherapy efficacy in combination with Ras inhibitor in triple-negative breast cancer.Clinical and translational medicine · 2025Article
- Kinase signaling cascades: an updated mechanistic landscape.Chemical science · 2025Review
- Cell cycle dependence of ERK activation dynamics is regulated by PI3K and PAK1 signaling.Scientific reports · 2025Article
- Evaluation of information flows in the RAS-MAPK system using transfer entropy measurements.eLife · 2025Article
- MYC and KRAS cooperation: from historical challenges to therapeutic opportunities in cancer.Signal transduction and targeted therapy · 2024Review
- Regulation of RAF family kinases: new insights from recent structural and biochemical studies.Biochemical Society transactions · 2024Review
- Regulation of Ras p21 and RalA GTPases activity by quinine in mammary epithelial cells.Molecular and cellular biochemistry · 2024Article
- Electric field modulation of ERK dynamics shows dependency on waveform and timing.Scientific reports · 2024Article
- Application of single-molecule analysis to singularity phenomenon of cells.Biophysics and physicobiology · 2024Article
- Multiplexed imaging for probing RAS-RAF interactions in living cells.Biochimica et biophysica acta. Biomembranes · 2023Article
- p52Shc regulates the sustainability of ERK activation in a RAF-independent manner.Molecular biology of the cell · 2021Article
- Workflows of the Single-Molecule Imaging Analysis in Living Cells: Tutorial Guidance to the Measurement of the Drug Effects on a GPCR.Methods in molecular biology (Clifton, N.J.) · 2021Article
- In-Cell Single-Molecule Analysis of Molecular State and Reaction Kinetics Coupling.Advances in experimental medicine and biology · 2021Article
- Biphasic spatiotemporal regulation of GRB2 dynamics by p52SHC for transient RAS activation.Biophysics and physicobiology · 2021Article
- K-Ras prenylation as a potential anticancer target.Cancer metastasis reviews · 2020Review
- Biology, pathology, and therapeutic targeting of RAS.Advances in cancer research · 2020Review
- Localization dynamics of endogenous fluorescently labeled RAF1 in EGF-stimulated cells.Molecular biology of the cell · 2019Article
- Complement Receptor 3-Mediated Inhibition of Inflammasome Priming by Ras GTPase-Activating Protein DuringFrontiers in immunology · 2018Article
- Mutation-Specific Mechanisms of Hyperactivation of Noonan Syndrome SOS Molecules Detected with Single-molecule Imaging in Living Cells.Scientific reports · 2017Article
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Authors and funding
4 authors at 3 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
RAS is an important cell signaling molecule, regulating the activities of various effector proteins, including the kinase c-RAF (RAF). Despite the critical function of RAS signaling, the activation kinetics have not been analyzed experimentally in living cells for any of the RAS effectors. Here, we analyzed the kinetics of RAF activation on the plasma membrane in living HeLa cells after stimulation with EGF to activate RAS. RAF is recruited by the active form of RAS (RAS-GTP) from the cytoplasm to the plasma membrane through two RAS-binding sites (the RAS-binding domain and the cysteine-rich domain (CRD)) and is activated by its phosphorylation by still undetermined kinases on the plasma membrane. Using single-molecule imaging, we measured the dissociation time courses of GFP-tagged molecules of wild type RAF and fragments or mutants of RAF containing one or two of the three functional domains (the RAS-binding domain, the CRD, and the catalytic domain) to determine their interaction with membrane components. Each molecule showed a unique dissociation time course, indicating that both its interaction with RAS-GTP and its phosphorylation by the kinases are rate-limiting steps in RAF activation. Based on our experimental results, we propose a kinetic model for the activation of RAF. The model suggests the importance of the interaction between RAS-GTP and CRD for the effective activation of RAF, which is triggered by rapid RAS-GTP-induced conformational changes in RAF and the subsequent presentation of RAF to the kinase. The model also suggests necessary properties of the kinases that activate RAF.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.