Nucleophosmin and Nucleolin Regulate K-Ras Plasma Membrane Interactions and MAPK Signal Transduction

Inder, Kerry L.; Lau, Chiyan; Loo, Dorothy; Chaudhary, Natasha; Goodall, Andrew; Martin, Sally; Jones, Alun; Hoeven, Dharini van der; Parton, Robert G.; Hill, Michelle M.; Hancock, John F.

doi:10.1074/jbc.m109.001537

Cited by 68 publications

(65 citation statements)

References 42 publications

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“…Nucleolin was reported previously to interact with K-Ras4B but not with H-Ras in the nucleolus (32). In a study published during the final preparation of our manuscript, it was reported that nucleolin from postnuclear supernatants of BHK cells interacts with K-Ras but not with H-Ras (43). The basis for this discrepancy is currently unclear.…”

Section: Discussionmentioning

confidence: 49%

Oncogenic Synergism between ErbB1, Nucleolin, and Mutant Ras

et al. 2011

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Alterations in the ErbB family of growth factor receptors, their signaling components, and mutational activation of Ras proteins are major contributors to malignant transformation. Recently, mutant Ras was shown to be capable of activating ErbB receptors in a ligand-independent manner. Furthermore, it was observed that nucleolin, a transcriptional regulator and ribosome biogenesis factor, can bind both K-Ras and the cytoplasmic tail of ErbB receptors to enhance ErbB receptor activation. However, the functional significance of these interactions to cancer pathogenesis has not been probed. Here, we show that endogenous nucleolin interacts simultaneously in vivo with endogenous Ras and ErbB1 (EGFR) in cancer cells. The C-terminal 212 amino acids of nucleolin were determined to be sufficient to interact with ErbB1 and all Ras protein isoforms (H-, N-, and KRas). Nucleolin partially colocalizes with Ras at the plasma membrane. Moreover, activated but not wild-type Ras facilitates nucleolin interaction with ErbB1 and stabilizes ErbB1 receptor levels. Most importantly, these three oncogenes synergistically facilitate anchorage-independent cell growth in vitro and tumor growth in vivo. Our findings suggest strategies to target nucleolin as a general approach to inhibiting ErbB-and Ras-driven cancers. Cancer Res; 71(6); 2140-51. Ó2011 AACR.

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Section: Discussionmentioning

confidence: 49%

Oncogenic Synergism between ErbB1, Nucleolin, and Mutant Ras

et al. 2011

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“…This interaction may further be modulated through phosphorylation of Ras at the C terminus (31). Additionally, certain membrane proteins, such as galectin (32), nucleophosmin (33), and calmodulin (34) could serve as scaffolds to bring two or more Ras proteins into proximity. Lastly, albeit secondary in driving Ras dimer formation, direct G-domain contacts between the two Ras proteins should not be ruled out based on existing data.…”

Section: Discussionmentioning

confidence: 99%

Ras-GTP dimers activate the Mitogen-Activated Protein Kinase (MAPK) pathway

Nan

Tamgüney

Collisson

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Rat sarcoma (Ras) GTPases regulate cell proliferation and survival through effector pathways including Raf-MAPK, and are the most frequently mutated genes in human cancer. Although it is well established that Ras activity requires binding to both GTP and the membrane, details of how Ras operates on the cell membrane to activate its effectors remain elusive. Efforts to target mutant Ras in human cancers to therapeutic benefit have also been largely unsuccessful. Here we show that Ras-GTP forms dimers to activate MAPK. We used quantitative photoactivated localization microscopy (PALM) to analyze the nanoscale spatial organization of PAmCherry1-tagged KRas 4B (hereafter referred to KRas) on the cell membrane under various signaling conditions. We found that at endogenous expression levels KRas forms dimers, and KRas G12D , a mutant that constitutively binds GTP, activates MAPK. Overexpression of KRas leads to formation of higher order Ras nanoclusters. Conversely, at lower expression levels, KRas G12D is monomeric and activates MAPK only when artificially dimerized. Moreover, dimerization and signaling of KRas are both dependent on an intact CAAX (C, cysteine; A, aliphatic; X, any amino acid) motif that is also known to mediate membrane localization. These results reveal a new, dimerization-dependent signaling mechanism of Ras, and suggest Ras dimers as a potential therapeutic target in mutant Ras-driven tumors.Ras dimer | MAPK signaling | cancer | single molecule imaging | superresolution microscopy T he canonical rat sarcoma (Ras) GTPase family members H-, N-, and K-ras are frequently activated in human cancers (1-4) by recurrent point mutations at codons 12, 13, or 61. These mutations result in constitutive binding of Ras to GTP due to impaired GTP hydrolysis (5). Despite nearly identical G-domains, mammalian Ras isoforms serve nonredundant biological roles and exhibit different mutational spectra in human cancers (1,4,6). These functional differences are in part attributed to distinctions in the membranetethering motif at the C-terminal of Ras known as the hyper-variable region [HVR, which includes the "CAAX" (C, cysteine; A, aliphatic; X, any amino acid) motif] (6, 7). Although mechanisms regulating Ras-GTP levels in cells have been examined extensively, details of how Ras organizes and operates on the cell membrane have been elusive. Efforts on targeting mutant Ras to therapeutic benefits in human cancers by inhibiting membrane localization or GTP binding have not been successful, leaving mutant Ras an intractable drug target (8). Hence, identification of new mechanisms that regulate Ras oncogenesis is crucial to combating mutant Ras-driven cancers.Recent studies using immuno electron microscopy (immuno-EM) have implicated a previously unappreciated spatial mechanism in regulating the biological functions of Ras. In particular, Ras proteins were found to form 5-to 8-membered nanoclusters that serve as signaling scaffolds for recruiting and activating downstream effectors such as Raf and PI3K on the cell membr...

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“…59,60 Moreover, it has been reported the interaction of the nucleocytoplasmic shuttling proteins Nucleophosmin and Nucleolin with KRAS to enhance nanocluster formation and signal gain in the MAPK pathway. 15 Thus, the functional interaction of the cytoplasm confined KRAS with HNRNPA2B1 could be understood in the context of a novel hnRNP cytoplasmic function.…”

Section: Discussionmentioning

confidence: 99%

“…In recent years, emergence of new regulators either by direct interaction or by reversible post-translational modifications, proved to be crucial to fully display KRAS oncogenic phenotype. Interaction of KRAS with Galectin-3, 11 calmodulin, 12 and PDEδ 13,14 or with the initially-defined nuclear protein Nucleophosmin and Nucleolin, 15 proved to be necessary for correct activation of RAF/MEK/ERK signaling pathway.…”

Section: Introductionmentioning

confidence: 99%