IndraLab
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"Although the K-Ras/Raf-RBD interaction was readily detectable upon co-expression in a single cell line, or following lysis of co-cultured cell lines separately expressing K-Ras and RBD, bearing in mind the limitations of our assay, we were unable to detect the interaction in the intact, co-cultured cell lines or upon treatment of the Raf-RBD-expressing cells with exosomes containing K-Ras."
sparser
"Zhang and others described that K-RAS4A interacts with Raf-1 with higher affinity than K-RAS4B, leading to increased RAF1-1MEK-ERK signalling cascade, and that K-RAS4A showed increased anchorage-independent growth in assays that compared K-RAS4A- and K-RAS4B-transformed NIH 3T3 cells ( xref )."
sparser
"Moreover, significant co-localized signal between K-Ras and Raf1 was found in the cell membrane ( xref ), suggesting that K-Ras affects the cellular localization of Raf1, and the cellular interaction between K-Ras and Raf1 is likely to facilitate the membrane-binding ability of Raf1."
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"Results above show that the BiFC system composed of Raf1 and K-Ras is sensitive for EGF stimulation, and the trend of BiFC fluorescence change is consistent with previous report on the K-Ras response to EGF stimulation [XREF_BIBR], indicating that the activation of K-Ras can promote its complex formation with Raf1, and the Raf1 and K-Ras interaction in BiFC system has not been interfered by the fluorescent protein fragments they connected, and this system is suitable for further studies of Raf1 and K-Ras interaction."
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"However, there are still some unresolved issues regarding their interactions such as where and how the activation of Raf1 and K-Ras occurs in cells, whether K-Ras and Raf1 simply traffic together or are part of a larger multicomponent signaling complexes, as well as whether the ultimate cellular localization of the K-Ras and Raf1 complexes is independent of the original Raf1 and K-Ras locations."
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"Although the K-Ras and Raf-RBD interaction was readily detectable upon co-expression in a single cell line, or following lysis of co-cultured cell lines separately expressing K-Ras and RBD, bearing in mind the limitations of our assay, we were unable to detect the interaction in the intact, co-cultured cell lines or upon treatment of the Raf-RBD-expressing cells with exosomes containing K-Ras."
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"Using this BEVL-BiFC system, we provided direct evidence in individual cells that post-translational modification of K-Ras and its PM localization were not essential for the interaction between K-Ras and Raf-1, while the interaction of Grb2 and K-Ras depended on PM localization of K-Ras."
sparser
"The results also provide direct evidence in individual cells that post-translational modification of K-Ras and its localization at the plasma membrane (PM) were not essential for the interaction of K-Ras and Raf-1, whereas the interaction of Grb2 and K-Ras did depend on the PM localization of K-Ras."
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"In contrast, an apparent cytosolic distribution pattern was observed in cells co-transfected with mcerulean-Raf1 and EGFP-K-RasC185S, suggesting that the membrane localization of K-Ras and Raf1 complexes is not entirely dependent on K-Ras, and that other factors, such as the irreversible conformation formed between K-Ras and Raf1 may play a role."
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"The results also provide direct evidence in individual cells that post-translational modification of K-Ras and its localization at the plasma membrane (PM) were not essential for the interaction of K-Ras and Raf-1, whereas the interaction of Grb2 and K-Ras did depend on the PM localization of K-Ras."
sparser
"To this end, we treated MiaPaCa2 cells with FGTI-2734 as described above, and processed the cells for RAF-1 immunoprecipitation, followed by immunoblotting of KRAS and KSR, a scaffolding protein that binds RAF-1 and mediates RAS activation of RAF-1. xref shows that in the absence or presence of FGTI-2734, RAF-1 was able to bind KRAS, suggesting that both membrane and cytosolic KRAS are able to bind RAF-1."
sparser
"While interaction based on Venus signal between K-Ras4B G12D and C-Raf in membrane was observed at 81.60 ± 13.24 of the cells, interaction between K-Ras4B G12D/K101D/R102E and C-Raf was observed at 28.63 ± 8.97 and interaction between K-Ras4B G12D/R41E/K42D and C-Raf was observed at 5.63 ± 1.43 of the transfected cells. ( xref – xref )."
"Among other effectors, active ras binds and activates the raf kinase, iniziating a kinase cascade involving serine phosporylation of mek1/2 (mapkk) and tyrosine and threonine phosphorylation of erk1/2. the raf family of proteins (raf-1, a-raf, and b-raf) is serine/threonine kinases that bind to the effector region of ras-gtp, thus inducing translocation of the protein to the plasma membrane. Once there, raf proteins are activated and phosphorylated by different protein kinases.. Association of ras with the mapk kinase kinase, raf, initiates the raf mek erk map kinase cascade."
"Among other effectors, active ras binds and activates the raf kinase, iniziating a kinase cascade involving serine phosporylation of mek1/2 (mapkk) and tyrosine and threonine phosphorylation of erk1/2. the raf family of proteins (raf-1, a-raf, and b-raf) is serine/threonine kinases that bind to the effector region of ras-gtp, thus inducing translocation of the protein to the plasma membrane. Once there, raf proteins are activated and phosphorylated by different protein kinases.. Association of ras with the mapk kinase kinase, raf, initiates the raf mek erk map kinase cascade."
sparser
"Our pharmacological and genetic approaches show that cytosolic KRAS is still able to bind one of its major effectors RAF-1 but is not able to activate it, possibly due to inhibition of binding to the scaffolding protein KSR (see xref ) which plays a major role in RAS activation of Raf ( xref )."
sparser
"We also analyzed the potential impact of ubiquitination at K147 on the interaction of KRAS with C-RAF CRD ( xref and xref , xref ). xref and shows that CRD-interacting residues of KRAS do not make direct contact with ubiquitin, suggesting that they are available for interaction with RAF CRD in mUbKRAS 147 ."
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"As shown in XREF_FIG, sub-cellular localization of the K-Ras and Raf1 complexes which was mainly distributed in the cell membrane (XREF_FIG) varied from the K-Ras-C185S and Raf1, which was mainly localized in the cytoplasm (XREF_FIG), and these distinct subcellular localizations were further confirmed in different imaging depth (XREF_FIG), indicating that the C-terminal CAAX site is the determinant for sub-cellular localization of K-Ras and Raf1 complexes, which is consistent with previous reports [XREF_BIBR]."
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"Moreover, significant co-localized signal between K-Ras and Raf1 was found in the cell membrane (XREF_FIG), suggesting that K-Ras affects the cellular localization of Raf1, and the cellular interaction between K-Ras and Raf1 is likely to facilitate the membrane binding ability of Raf1."
sparser
"Co-precipitation of Kras with GST-Raf1-RBD was significantly decreased in cells expressing STAND-Y13-259 when compared to those expressing STAND-A36 or the mock vector control (Fig. xref ), indicating that intracellular STAND-Y13-259 interferes with the interaction of activated Kras and Raf1 by binding to Kras."
sparser
"Using this BEVL-BiFC system, we provided direct evidence in individual cells that post-translational modification of K-Ras and its PM localization were not essential for the interaction between K-Ras and Raf-1, while the interaction of Grb2 and K-Ras depended on PM localization of K-Ras."