Distinct Binding Preferences between Ras and Raf Family Members and the Impact on Oncogenic Ras Signaling

Terrell, Elizabeth M.; Durrant, David; Ritt, Daniel A.; Sealover, Nancy E.; Sheffels, Erin; Spencer-Smith, Russell; Esposito, Dominic; Zhou, Yong; Hancock, John F.; Kortum, Robert L.; Morrison, Deborah K.

doi:10.1016/j.molcel.2019.09.004

Cited by 89 publications

(84 citation statements)

References 36 publications

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“…The Ras R123-E143 salt bridge remains present throughout the simulations, even as R143 of the Raf-CRD moves to interact with Ras E143 ( Fig (42) . Furthermore, the asymmetry and flexibility that we observe in the dimer may facilitate a given Ras isoform to accommodate heterodimers of Raf, important in signal activation (43), as CRaf residues N161, E174, and H175 are isoform specific and correspond to Q257, Q270, and R271 in BRaf (Fig.…”

Section: Raf-crd Links To the Active Site Through Loop 8 Across The Dmentioning

confidence: 99%

Crystal structure reveals the full Ras:Raf interface and advances mechanistic understanding of Raf activation

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Mattos

2020

Preprint

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The interaction between Ras and Raf-kinase through the Ras-binding (RBD) and cysteine-rich domains (CRD) of Raf is essential for signaling through the mitogen-activated protein kinase (MAPK) pathway, yet the molecular mechanism leading to Raf activation has remained elusive. We present the 2.8 Å crystal structure of the HRas/CRaf-RBD_CRD complex showing the Ras/Raf interface as a continuous surface on Ras. In the Ras dimer, with helices roughly perpendicular to the membrane, the CRD is located between the two Ras protomers and far from the membrane, where its dynamic nature in the Ras binding pocket is expected to accommodate BRaf and CRaf heterodimers. Our structure and its analysis by MD simulations, combined with work in the literature, result in a molecular model in which Ras binding is involved in the release of Raf autoinhibition while the Ras/Raf complex dimerizes to promote a platform for signal amplification, with Raf-CRD poised to have direct and allosteric effects on both the Ras active site and the dimerization interface.

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Section: Raf-crd Links To the Active Site Through Loop 8 Across The Dmentioning

confidence: 99%

Crystal structure reveals the full Ras:Raf interface and advances mechanistic understanding of Raf activation

Cookis

Mattos

2020

Preprint

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“…Given the increased affinity of KRAS Q61H for RAF, and the lack of GTPase activity in KRAS Q61H , which would be expected to allow for stable complex formation, we hypothesized that KRAS Q61H , in particular, may show dependence on the formation of RAF dimers. Prior work showed that the D154Q substitution in KRAS was sufficient to disrupt KRAS-KRAS interactions at the cell membrane that occur via the KRAS a4-a5 interface, but did not alter the baseline biochemical properties of KRAS such as GTPase activity or affinity for RAF (39). The D154Q mutation also abolished the suppressive effects of KRAS WT seen in the genetic background of oncogenic KRAS.…”

Section: Kras Q61h Oligomerization and C-raf Dimerization Are Interdementioning

confidence: 99%

KRASQ61H Preferentially Signals through MAPK in a RAF Dimer-Dependent Manner in Non–Small Cell Lung Cancer

et al. 2020

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Assembly of RAS molecules into complexes at the cell membrane is critical for RAS signaling. We previously showed that oncogenic KRAS codon 61 mutations increase its affinity for RAF, raising the possibility that KRAS Q61H , the most common KRAS mutation at codon 61, upregulates RAS signaling through mechanisms at the level of RAS assemblies. We show here that KRAS Q61H exhibits preferential binding to RAF relative to PI3K in cells, leading to enhanced MAPK signaling in in vitro models and human NSCLC tumors. X-ray crystallography of KRAS Q61H :GTP revealed that a hyperdynamic switch 2 allows for a more stable interaction with switch 1, suggesting that enhanced RAF activity arises from a combination of absent intrinsic GTP hydrolysis activity and increased affinity for RAF. Disruption of KRAS Q61H assemblies by the RAS oligomerdisrupting D154Q mutation impaired RAF dimerization and altered MAPK signaling but had little effect on PI3K signaling. However, KRAS Q61H oligomers but not KRAS G12D oligomers were disrupted by RAF mutations that disrupt RAF-RAF interactions. KRAS Q61H cells show enhanced sensitivity to RAF and MEK inhibitors individually, whereas combined treatment elicited synergistic growth inhibition. Furthermore, KRAS Q61H tumors in mice exhibited high vulnerability to MEK inhibitor, consistent with cooperativity between KRAS Q61H and RAF oligomerization and dependence on MAPK signaling. These findings support the notion that KRAS Q61H and functionally similar mutations may serve as predictive biomarkers for targeted therapies against the MAPK pathway. Significance: These findings show that oncogenic KRAS Q61H forms a cooperative RAS-RAF ternary complex, which renders RAS-driven tumors vulnerable to MEKi and RAFi, thus establishing a framework for evaluating RAS biomarker-driven targeted therapies. Recently, we reported a RAS mutation, D154Q, which neutralizes the suppressive effects of KRAS WT in the context of oncogenic KRAS. This mutation was designed on the basis of a RAS-RAS interaction that occurs via an a4-a5 interface seen in some RAS crystal structures (12). Recent NMR studies of RAS molecules bound to nanodisks also

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“…Isoform-specific signaling is thought to be mediated by differential intracellular localisation that favors preferential coupling to specific effector pathways (4,22,23), and by distinct biochemical properties imparted by allosteric lobe sequence variations between each isoform (24). Recent in vitro analysis revealed distinct binding preferences for Ras-Raf interactions with BRAF binding being highly selective for KRAS whilst CRAF was critical for HRAS-mediated MAP kinase signaling (25). Mutational-specificity is also important for Ras biology (12,(26)(27)(28)(29), and structural and biochemical features underpinning mutational differences in nucleotide cycling, allosteric regulation and GEF, GAP and effector interactions are now being defined (14,25,28,(30)(31)(32).…”

Section: Oncogenic Rasmentioning

confidence: 99%

The Frequency of Ras Mutations in Cancer

2020

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Ras is frequently mutated in cancer; however, there is a lack of consensus in the literature regarding the cancer mutation frequency of Ras, with quoted values varying from 10-30%. This variability is at least in part due to the selective aggregation of data from different databases and the dominant influence of particular cancer types and particular Ras isoforms within these datasets. In order to provide a more definitive figure for Ras mutation frequency in cancer, we cross-referenced the data in all major publicly accessible cancer mutation databases to determine reliable mutation frequency values for each Ras isoform in all major cancer types. These percentages were then applied to current US cancer incidence statistics to estimate the number of new patients each year that have Ras-mutant cancers. We find that ~19% of cancer patients harbor Ras mutations; equivalent to ~3.4 million new cases per year worldwide. We discuss the Ras isoform and mutation-specific trends evident within the datasets that are relevant to current Ras-targeted therapies. Research.

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Distinct Binding Preferences between Ras and Raf Family Members and the Impact on Oncogenic Ras Signaling

Cited by 89 publications

References 36 publications

Crystal structure reveals the full Ras:Raf interface and advances mechanistic understanding of Raf activation

Crystal structure reveals the full Ras:Raf interface and advances mechanistic understanding of Raf activation

KRASQ61H Preferentially Signals through MAPK in a RAF Dimer-Dependent Manner in Non–Small Cell Lung Cancer

The Frequency of Ras Mutations in Cancer

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