KY1022, a small molecule destabilizing Ras via targeting the Wnt/β-catenin pathway, inhibits development of metastatic colorectal cancer

Cho, Yong Hee; Hyeon, Pu; Kaduwal, Saluja; Park, Jong Chan; Lee, Sang Kyu; Yoon, Jeong Soo; Shin, Wookjin; Kim, Hyun-Tae; Ro, Eun Ji; Koo, Kyung Hwa; Park, Ki‐Sook; Han, Gyoonhee; Choi, Kang Yell

doi:10.18632/oncotarget.13172

Cited by 31 publications

(40 citation statements)

References 39 publications

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“…In addition, and more classically, polyubiquitination can alter RAS function through degradation. Proteasomal degradation of HRAS has been reported to occur in response to activation by the Wnt/␤-catenin signaling pathway (85)(86)(87). This polyubiquitin-dependent degradation occurs upon recruitment of ␤-TrCP-E3 ligase after HRAS phosphorylation by glycogen synthase kinase 3␤.…”

Section: Jbc Reviews: G Protein Ubiquitinationmentioning

confidence: 99%

Regulation of large and small G proteins by ubiquitination

Dohlman

Campbell

2019

Journal of Biological Chemistry

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Edited by Mike Shipston Many sensory and chemical signal inputs are transmitted by intracellular GTP-binding (G) proteins. G proteins make up two major subfamilies: "large" G proteins comprising three subunits and "small" G proteins, such as the proto-oncogene product RAS, which contains a single subunit. Members of both subfamilies are regulated by post-translational modifications, including lipidation, proteolysis, and carboxyl methylation. Emerging studies have shown that these proteins are also modified by ubiquitination. Much of our current understanding of this posttranslational modification comes from investigations of the large G-protein ␣ subunit from yeast (Gpa1) and the three RAS isotypes in humans, NRAS, KRAS, and HRAS. G␣ undergoes both mono-and polyubiquitination, and these modifications have distinct consequences for determining the sites and mechanisms of its degradation. Genetic and biochemical reconstitution studies have revealed the enzymes and binding partners required for addition and removal of ubiquitin, as well as the delivery and destruction of both the mono-and polyubiquitinated forms of the G protein. Complementary studies of RAS have identified multiple ubiquitination sites, each having distinct consequences for binding to regulatory proteins, shuttling to and from the plasma membrane, and degradation. Here, we review what is currently known about these two well-studied examples, Gpa1 and the human RAS proteins, that have revealed additional mechanisms of signal regulation and dysregulation relevant to human physiology. We also compare and contrast the effects of G-protein ubiquitination with other post-translational modifications of these proteins. A variety of sensory and chemical signals are detected by receptors at the cell surface. In many cases, these inputs are transmitted by intracellular GTP-binding proteins (Fig. 1). Generally speaking, large G proteins are activated by seven transmembrane segment receptors, also known as G-proteincoupled receptors (GPCRs). 3 When these receptors bind to an agonist ligand, the G-protein ␣ subunit releases GDP, binds to GTP, and dissociates from the G␤␥ subunit dimer. GTP-bound G␣, G␤␥, or both go on to activate intracellular effectors that transduce the signal. Similarly, many small G proteins are activated by growth factor receptors and their associated guanine nucleotide-exchange factors (GEFs). As with GPCRs, GEFs promote the exchange of GTP for GDP and subsequent activation of downstream effectors. For both large and small G proteins, signaling persists until GTP is converted to GDP, and this inactivation step is accelerated by binding to GTPase-activating proteins (GAPs). Thus, GEFs and GAPs work in opposition to one another to regulate the activity of their cognate G proteins, both large and small. Here, we review newer mechanisms of G-protein regulation by ubiquitination, with a focus on a large G protein from yeast and three small G proteins from humans: NRAS, KRAS, and HRAS. We compare and contrast the effects of ubiquitination with ...

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Section: Jbc Reviews: G Protein Ubiquitinationmentioning

confidence: 99%

Regulation of large and small G proteins by ubiquitination

Dohlman

Campbell

2019

Journal of Biological Chemistry

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“…Aberrant activation of the Wnt/β-catenin and RAS signaling pathways with elevated β-catenin and RAS protein levels has been observed in various human cancers including CRC, gastric cancer, endometrium cancer, and non-small cell lung cancer (NSCLC) [19,[26][27][28][29][30]. Therefore, small molecule-mediated destabilization of both βcatenin and RAS has been shown to suppress the tumor growth in cancers such as CRC, gastric cancer, and NSCLC The effectiveness of small molecules to many different types of cancers is due to the destabilization of β-catenin and RAS proteins which are overexpressed in the different types of cancer [19,[28][29][30], suggesting that stabilization of β-catenin and RAS play critical roles in regulating the activation of Wnt/β-catenin and RAS signaling pathways in many types of cancers as well as the effect of their component gene mutation.…”

Section: Discussionmentioning

confidence: 99%

Small molecule-induced simultaneous destabilization of β-catenin and RAS is an effective molecular strategy to suppress stemness of colorectal cancer cells

Cho

Yoon

et al. 2020

Cell Commun Signal

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Background: Cancer stem cells (CSCs), the major driver of tumorigenesis, is a sub-population of tumor cells responsible for poor clinical outcomes. However, molecular mechanism to identify targets for controlling CSCs is poorly understood. Methods: Gene Set Enrichment Analyses (GSEA) of Wnt/β-catenin and RAS signaling pathways in stem-like subtype of colorectal cancer (CRC) patients were performed using two gene expression data set. The therapeutic effects of destabilization of β-catenin and RAS were tested by treatment of small molecule KYA1797K using CRC patient derived cells. Results: Treatment with KYA1797K, a small molecule that destabilizes both β-catenin and RAS via Axin binding, effectively suppresses the stemness of CSCs as shown in CRC spheroids and small intestinal tumors of Apc Min/+ /K-Ras G12D LA2 mice. Moreover, KYA1797K also suppresses the stemness of cells in CRC patient avatar model systems, such as patient-derived tumor organoids (PDTOs) and patient-derived tumor xenograft (PDTX). Conclusion: Our results suggest that destabilization of both β-catenin and RAS is a potential therapeutic strategy for controlling stemness of CRC cells.

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“…Bacterial cells were grown in LB media with ampicillin (100 lg/ml) at 37°C until the optical density at 600 nm reached approximately 0.6, at which time IPTG was added to a concentration of 0.2 or 1 mM, and the cells were incubated for an additional 20 h at 18°C. To prepare uniformly 15 N-and 13 C/ 15 N-labeled proteins for the NMR spectroscopy experiments, the cells were cultured in M9 media that contained 15 NH 4 Cl (Cambridge Isotope Lab) as the source of nitrogen and 13 C-Dglucose. The cell pellets were lysed by sonication in lysis buffer that contained protease inhibitor cocktail (for KRAS-FL, 40 mM of HEPES, 300 mM of NaCl, 5 mM of MgCl 2 , 5 mM of b-mercaptoethanol, pH 8.0; for KRAS-110-165, 25 mM of NaPi, 300 mM of NaCl, 10 mM of BME, pH 6.8; for b-catenin-FL, 20 mM of NaPi, 50 mM of NaCl, 10 mM of b-mercaptoethanol, pH 8.0 with 10% glycerol, 0.1% Triton X-100; for b-catenin-661-781, 25 mM of HEPES, 300 mM of NaCl, pH 7.5).…”

Section: Protein Expression and Purification For Nuclear Magnetic Resmentioning

confidence: 99%

“…All NMR experiments were performed at 298K on a Bruker Avance 600 MHz instrument (Billerica), and 2D 1 H-15 N HSQC experiments were performed with the following conditions: 90% H 2 O/10% 2 H 2 O NMR buffer that contained 50 mM of HEPES, 50 mM of NaCl, 2 mM of MgCl 2 , 2 mM of dithiothreitol (DTT), pH 7.4. For NMR titration experiments, 15 N-labeled KRAS was purified from M9 media that contained 15 NH 4 Cl, and b-catenin was obtained from LB media. The 1 H-15 N HSQC spectra were obtained at various molar ratios of 15 Nlabeled proteins with respect to partner proteins.…”

Section: Nmr Spectroscopy Titrationmentioning

confidence: 99%

β‐Catenin‐ RAS interaction serves as a molecular switch for RAS degradation via GSK 3β

et al. 2018

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RAS proteins play critical roles in various cellular processes, including growth and transformation. RAS proteins are subjected to protein stability regulation via the Wnt/β‐catenin pathway, and glycogen synthase kinase 3 beta (GSK3β) is a key player for the phosphorylation‐dependent RAS degradation through proteasomes. GSK3β‐mediated RAS degradation does not occur in cells that express a nondegradable mutant (MT) β‐catenin. Here, we show that β‐catenin directly interacts with RAS at the α‐interface region that contains the GSK3β phosphorylation sites, threonine 144 and threonine 148 residues. Exposure of these sites by prior β‐catenin degradation is required for RAS degradation. The introduction of a peptide that blocks the β‐catenin‐RAS interaction by binding to β‐catenin rescues the GSK3β‐mediated RAS degradation in colorectal cancer (CRC) cells that express MT β‐catenin. The coregulation of β‐catenin and RAS stabilities by the modulation of their interaction provides a mechanism for Wnt/β‐catenin and RAS‐ERK pathway cross‐talk and the synergistic transformation of CRC by both APC and KRAS mutations.

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KY1022, a small molecule destabilizing Ras via targeting the Wnt/β-catenin pathway, inhibits development of metastatic colorectal cancer

Cited by 31 publications

References 39 publications

Regulation of large and small G proteins by ubiquitination

Regulation of large and small G proteins by ubiquitination

Small molecule-induced simultaneous destabilization of β-catenin and RAS is an effective molecular strategy to suppress stemness of colorectal cancer cells

β‐Catenin‐ RAS interaction serves as a molecular switch for RAS degradation via GSK 3β

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