Rare Streptomyces sp. polyketides as modulators of K-Ras localisation

Salim, Angela A.; Xiao, Xue; Cho, Kwang Jin; Piggott, Andrew M.; Lacey, Ernest; Hancock, John F.; Capon, Robert J.

doi:10.1039/c4ob00745j

Cited by 17 publications

(14 citation statements)

References 13 publications

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“…57,58 In addition to PKC, PKG2 can robustly phosphorylate K-Ras on Ser181 as the target of an AMPK regulated signaling pathway. 45 Thus, pharmacological activation of the AMPK, eNOS, sGC, PKG2 pathway at multiple levels, with agents such as DEA-NO, oligomycin, [64][65][66][67] AICAR, sildenafil or metformin all mis-localize K-Ras from the PM and effectively inhibit the proliferation of K-Ras transformed NSCLC cell lines where this PKG2 signaling pathway is intact. 45 K-Ras does not co-localize with PIP 2 in intact PM and K-Ras nanoclustering and PM binding are independent of PIP 2 in lipid addback experiments.…”

Section: Interference With Ptdser Transport Compromises In Vitro Anmentioning

confidence: 99%

Deciphering lipid codes: K‐Ras as a paradigm

Zhou

Hancock

2017

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The cell plasma membrane (PM) is a highly dynamic and heterogeneous lipid environment, driven by complex hydrophobic and electrostatic interactions among the hundreds of types of lipid species. Although the biophysical processes governing lipid lateral segregation in the cell PM have been established in vitro, biological implications of lipid heterogeneity are poorly understood. Of particular interest is how membrane proteins potentially utilize transient spatial clustering of PM lipids to regulate function. This review focuses on a lipid-anchored small GTPase K-Ras as an example to explore how its C-terminal membrane-anchoring domain, consisting of a contiguous hexa-lysine polybasic domain and an adjacent farnesyl anchor, possesses a complex coding mechanism for highly selective lipid sorting on the PM. How this lipid specificity modulates K-Ras signal transmission will also be discussed.

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Section: Interference With Ptdser Transport Compromises In Vitro Anmentioning

confidence: 99%

Deciphering lipid codes: K‐Ras as a paradigm

Zhou

Hancock

2017

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“…We employed high content quantitative confocal imaging to assess the ability of these extracts to mislocalise oncogenic mutant K-Ras (mGFP-K-Ras G12V) from the PM of intact Madin-Darby canine kidney (MDCK cells). 4a In proof of concept studies, we documented staurosporine, 4a oxanthromicins 5 and neoantimycins 6 as promising inhibitors of K-Ras PM localisation. In this report we apply this refined biodiscovery approach to characterise the nM K-Ras mislocalisation properties of the oligomycins, a rare class of polyketide recovered from a soil-derived Streptomyces sp.…”

Section: Introductionmentioning

confidence: 96%

Oligomycins as inhibitors of K-Ras plasma membrane localisation

et al. 2016

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Frequently present in pancreatic, colorectal and non-small cell lung carcinomas, oncogenic mutant K-Ras must be localised to the plasma membrane (PM) to be functional. Inhibitors of K-Ras PM localisation are therefore putative cancer chemotherapeutics. By screening a microbial extract library in a high content cell-based assay we detected the rare oligomycin class of Streptomyces polyketides as inhibitors of K-Ras PM localisation. Cultivation and fractionation of three unique oligomycin producing Streptomyces strains yielded oligomycins A–E (1–5) and 21-hydroxy-oligomycin A (6), together with the new 21-hydroxy-oligomycin C (7) and 40-hydroxy-oligomycin B (8). Structures for 1–8 were assigned by detailed spectroscopic analysis. Cancer cell viability screening confirmed 1–8 were cytotoxic to human colorectal carcinoma cells (IC50 >3 μM), and were inhibitors of the ABC transporter efflux pump P-glycoprotein (P-gp), with 5 being comparable in potency to the positive control verapamil. Significantly, oligomycins 1–8 proved to be exceptionally potent inhibitors of K-Ras PM localisation (Emax 0.67–0.75 with an IC50 ~1.5–14 nM).

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“…Figure 1 shows that the direct AMPK activator aminoimidazole-4-carboxamide riboside (AICAR) very effectively displaces K-RasG12V from the PM. To explore the molecular mechanism, we first examined the cellular distribution of PtdSer, because the PM PtdSer content is critical for K-Ras PM binding (18)(19)(20)(21). We analyzed the localization of the PtdSer probe LactC2 by confocal imaging (4,38).…”

Section: Ampk Activation Triggers Dissociation Of K-ras From the Pmmentioning

confidence: 99%

“…One group of compounds disrupts the cellular phosphatidylserine (PtdSer) distribution or PtdSer levels with a consequent reduction in the PtdSer content of the inner leaflet of the PM (18)(19)(20)(21). Since PtdSer is an anionic phospholipid that provides much of the electrostatic surface potential to the inner PM, a reduced PtdSer PM content leads to K-Ras dissociation.…”

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AMPK and Endothelial Nitric Oxide Synthase Signaling Regulates K-Ras Plasma Membrane Interactions via Cyclic GMP-Dependent Protein Kinase 2

Cho

Casteel

Prakash

et al. 2016

Molecular and Cellular Biology

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f K-Ras must localize to the plasma membrane and be arrayed in nanoclusters for biological activity. We show here that K-Ras is a substrate for cyclic GMP-dependent protein kinases (PKGs). In intact cells, activated PKG2 selectively colocalizes with K-Ras on the plasma membrane and phosphorylates K-Ras at Ser181 in the C-terminal polybasic domain. K-Ras phosphorylation by PKG2 is triggered by activation of AMP-activated protein kinase (AMPK) and requires endothelial nitric oxide synthase and soluble guanylyl cyclase. Phosphorylated K-Ras reorganizes into distinct nanoclusters that retune the signal output. Phosphorylation acutely enhances K-Ras plasma membrane affinity, but phosphorylated K-Ras is progressively lost from the plasma membrane via endocytic recycling. Concordantly, chronic pharmacological activation of AMPK ¡ PKG2 signaling with mitochondrial inhibitors, nitric oxide, or sildenafil inhibits proliferation of K-Ras-positive non-small cell lung cancer cells. The study shows that K-Ras is a target of a metabolic stress-signaling pathway that can be leveraged to inhibit oncogenic K-Ras function. Ras proteins are small GTPases that regulate important cellular signaling cascades to control cell growth, proliferation, and differentiation (1). The three Ras isoforms, H-, N-, and K-Ras4B (here, K-Ras), are ubiquitously expressed in mammalian cells. Ras proteins must be localized to the inner leaflet of the plasma membrane (PM) by a C-terminal membrane anchor for biological activity. In the case of K-Ras, the anchor comprises a posttranslationally attached C-terminal cysteine farnesyl-methyl ester operating in concert with a polybasic motif of 6 lysine residues (2, 3). Electrostatic interactions between the K-Ras C-terminal polybasic domain and the negatively charged inner leaflet of the PM provide membrane affinity (2, 4-9). Maintenance of K-Ras on the PM also requires the chaperone protein PDE␦ (10). Cytosolic PDE␦ binds K-Ras released from the PM as a result of endocytosis and unloads K-Ras in the perinuclear region in response to Arl2/3 binding, whence K-Ras translocates to the recycling endosome (RE) for redelivery to the PM by vesicular transport (11). Ras proteins on the PM are spatially organized into nanodomains, called nanoclusters, that are required for high-fidelity signal transduction by the Ras/mitogen-activated protein kinase (MAPK) pathway (12-14). Ras GTP nanoclusters contain ϳ6 to 7 Ras proteins, are Ͻ20 nm in diameter, and are exclusive platforms for Raf recruitment and MEK/extracellular signal-regulated kinase (ERK) activation. Perturbation of the spatiotemporal dynamics of Ras nanoclustering disrupts cellular signaling (15, 16).We have used a high-content cell-based screen (HCS) to identify multiple chemical compounds that mislocalize K-Ras from the PM and abrogate K-Ras signal transmission (17,18). One group of compounds disrupts the cellular phosphatidylserine (PtdSer) distribution or PtdSer levels with a consequent reduction in the PtdSer content of the inner leaflet of the PM (18-21)....

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Rare Streptomyces sp. polyketides as modulators of K-Ras localisation

Cited by 17 publications

References 13 publications

Deciphering lipid codes: K‐Ras as a paradigm

Deciphering lipid codes: K‐Ras as a paradigm

Oligomycins as inhibitors of K-Ras plasma membrane localisation

AMPK and Endothelial Nitric Oxide Synthase Signaling Regulates K-Ras Plasma Membrane Interactions via Cyclic GMP-Dependent Protein Kinase 2

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