Upregulation of CYP1B1 by hypoxia is mediated by ERα activation in breast cancer cells.

Endocrine therapy for breast cancer often leads to drug resistance and tumor recurrence; tumor hypoxia is also associated with mortality and tumor relapse. Cytochrome P450 1B1 (CYP1B1) regulates estrogen metabolism in breast cells and is known to be overexpressed in breast cancer tissue. Although the individual association of hypoxia-induced hypoxia-inducible factor-1-alpha (HIF-1α) and CYP1B1 with tumorigenesis is well known, the association between HIF-1α and CYP1B1 leading to tumorigenesis has not been investigated. Here, we investigated the correlation between hypoxia and CYP1B1 expression in breast cancer cells for tumorigenesis-related mechanisms. Hypoxia was induced in the human breast cancer cell lines MCF-7 (Er-positive) and MDA-MB-231 (triple-negative) and the normal breast epithelial cell line MCF10A; these cell lines were then subjected to immunoblotting, transient transfection, luciferase assays, gene silencing using small interfering RNA, polymerase chain reaction analysis, chromatin immunoprecipitation, co-immunoprecipitation, and mammalian two-hybrid assays. Furthermore, immunofluorescence analysis of the tumor microarrays was performed, and the pub2015 and the Cancer Genome Atlas patient datasets were analyzed. HIF-1α expression in response to hypoxia occurred in both normal and breast cancer cells, whereas CYP1B1 was induced only in estrogen receptor α (ERα)-positive breast cancer cells under hypoxia. HIF-1α activated ERα through direct binding and in a ligand-independent manner to promote CYP1B1 expression. Therefore, we suggested the mechanism by which hypoxia and ER-positivity orchestrate breast cancer relapse.

Pillar-Based Mechanical Induction of an Aggressive Tumorigenic Lung Cancer Cell Model.

Tissue microarchitecture imposes physical constraints to the migration of individual cells. Especially in cancer metastasis, three-dimensional structural barriers within the extracellular matrix are known to affect the migratory behavior of cells, regulating the pathological state of the cells. Here, we employed a culture platform with micropillar arrays of 2 µm diameter and 16 µm pitch (2.16 micropillar) as a mechanical stimulant. Using this platform, we investigated how a long-term culture of A549 human lung carcinoma cells on the (2.16) micropillar-embossed dishes would influence the pathological state of the cell. A549 cells grown on the (2.16) micropillar array with 10 µm height exhibited a significantly elongated morphology and enhanced migration even after the detachment and reattachment, as evidenced in the conventional wound-healing assay, single-cell tracking analysis, and in vivo tumor colonization assays. Moreover, the pillar-induced morphological deformation in nuclei was accompanied by cell-cycle arrest in the S phase, leading to suppressed proliferation. While these marked traits of morphology-migration-proliferation support more aggressive characteristics of metastatic cancer cells, typical indices of epithelial-mesenchymal transition were not found, but instead, remarkable traces of amoeboidal transition were confirmed. Our study also emphasizes the importance of mechanical stimuli from the microenvironment during pathogenesis and how gained traits can be passed onto subsequent generations, ultimately affecting their pathophysiological behavior. Furthermore, this study highlights the potential use of pillar-based mechanical stimuli as an in vitro cell culture strategy to induce more aggressive tumorigenic cancer cell models.

Identification of Nucleolin as a Novel AEG-1-Interacting Protein in Breast Cancer via Interactome Profiling.

Breast cancer is one of the most common malignant diseases worldwide. Astrocyte elevated gene-1 (AEG-1) is upregulated in breast cancer and regulates breast cancer cell proliferation and invasion. However, the molecular mechanisms by which AEG-1 promotes breast cancer have yet to be fully elucidated. In order to delineate the function of AEG-1 in breast cancer development, we mapped the AEG-1 interactome via affinity purification followed by LC-MS/MS. We identified nucleolin (NCL) as a novel AEG-1 interacting protein, and co-immunoprecipitation experiments validated the interaction between AEG-1 and NCL in breast cancer cells. The silencing of NCL markedly reduced not only migration/invasion, but also the proliferation induced by the ectopic expression of AEG-1. Further, we found that the ectopic expression of AEG-1 induced the tyrosine phosphorylation of c-Met, and NCL knockdown markedly reduced this AEG-1 mediated phosphorylation. Taken together, our report identifies NCL as a novel mediator of the oncogenic function of AEG-1, and suggests that c-Met could be associated with the oncogenic function of the AEG-1-NCL complex in the context of breast cancer.

A Novel Protein-Protein Interaction between RSK3 and IκBα and a New Binding Inhibitor That Suppresses Breast Cancer Tumorigenesis.

Multiple cancer-related biological processes are mediated by protein-protein interactions (PPIs). Through interactions with a variety of factors, members of the ribosomal S6 kinase (RSK) family play roles in cell cycle progression and cell proliferation. In particular, RSK3 contributes to cancer viability, but the underlying mechanisms remain unknown. We performed a kinase library screen to find IκBα PPI binding partners and identified RSK3 as a novel IκBα binding partner using a cell-based distribution assay. In addition, we discovered a new PPI inhibitor using mammalian two-hybrid (MTH) analysis. We assessed the antitumor effects of the new inhibitor using cell proliferation and colony formation assays and monitored the rate of cell death by FACS apoptosis assay. IκBα is phosphorylated by the active form of the RSK3 kinase. A small-molecule inhibitor that targets the RSK3/IκBα complex exhibited antitumor activity in breast cancer cells and increased their rate of apoptosis. RSK3 phosphorylation and RSK3/IκBα complex formation might be functionally important in breast tumorigenesis. The RSK3/IκBα-specific binding inhibitor identified in this study represents a lead compound for the development of new anticancer drugs.

Co-relation with novel phosphorylation sites of IκBα and necroptosis in breast cancer cells.

BACKGROUND: Phosphorylation of NF-kappaB inhibitor alpha (IκBα) is key to regulation of NF-κB transcription factor activity in the cell. Several sites of IκBα phosphorylation by members of the IκB kinase family have been identified, but phosphorylation of the protein by other kinases remains poorly understood. We investigated a new phosphorylation site on IκBα and identified its biological function in breast cancer cells. METHODS: Previously, we observed that aurora kinase (AURK) binds IκBα in the cell. To identify the domains of IκBα essential for phosphorylation by AURK, we performed kinase assays with a series of IκBα truncation mutants. AURK significantly promoted activation of IκBα at serine 32 but not serine 36; by contrast, IκB kinase (IKK) family proteins activated both of these residues. We also confirmed phosphorylation of IκBα by matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/TOF MS) and nano-liquid chromatography hybrid quadrupole orbitrap mass spectrometer (nanoLC-MS/MS; Q-Exactive). RESULTS: We identified two novel sites of serine phosphorylation, S63 and S262. Alanine substitution of S63 and S262 (S63A and S262A) of IκBα inhibited proliferation and suppressed p65 transcription activity. In addition, S63A and/or S262A of IκBα regulated apoptotic and necroptotic effects in breast cancer cells. CONCLUSIONS: Phosphorylation of IκBα by AURK at novel sites is related to the apoptosis and necroptosis pathways in breast cancer cells.

Activity-Based Protein Profiling Reveals Potential Dasatinib Targets in Gastric Cancer.

Dasatinib is a multi-target kinase inhibitor, whose targets include BCR-ABL, SRC family kinases, and various cancer kinases. The elevated SRC activity in gastric cancer (GC) has prompted the need for the therapeutic application of dasatinib in GC. We observed that the efficacy of dasatinib varied with the GC cell lines. The differential effect of dasatinib was not correlated with the basal SRC activity of each cell line. Moreover, the GC cell lines showing the strong antitumor effects of dasatinib were refractory to other SRC inhibitors, i.e., bosutinib and saracatinib, suggesting that unexpected dasatinib's targets could exist. To profile the targets of dasatinib in GC, we performed activity-based protein profiling (ABPP) via mass spectrometry using a desthiobiotin-ATP probe. We identified 29 and 18 kinases as potential targets in dasatinib-sensitive (SNU-216, MKN-1) and -resistant (SNU-484, SNU-601) cell lines, respectively. The protein-protein interaction mapping of the differential drug targets in dasatinib-sensitive and -resistant GC using the STRING database suggested that dasatinib could target cellular energy homeostasis in the drug-sensitive GC. RNAi screening for identified targets indicated p90RSK could be a novel dasatinib target, which is important for maintaining the viability and motility of GC cells. Further functional validation of dasatinib off-target actions will provide more effective therapeutic options for GC.

Development of a Polo-like Kinase-1 Polo-Box Domain Inhibitor as a Tumor Growth Suppressor in Mice Models.

Polo-like kinase-1 (Plk1) plays a key role in mitosis and has been identified as an attractive anticancer drug target. Plk1 consists of two drug-targeting sites, namely, N-terminal kinase domain (KD) and C-terminal polo-box domain (PBD). As KD-targeting inhibitors are associated with severe side effects, here we report on the pyrazole-based Plk1 PBD inhibitor, KBJK557, which showed a remarkable in vitro anticancer effect by inducing Plk1 delocalization, mitotic arrest, and apoptosis in HeLa cells. Further, in vivo optical imaging analysis and antitumorigenic activities in mouse xenograft models demonstrate that KBJK557 preferentially accumulates in cancer cells and selectively inhibits cancer cell proliferation. Pharmacokinetic profiles and partition coefficients suggest that KBJK557 was exposed in the blood and circulated through the organs with an intermediate level of clearance (t1/2, 7.73 h). The present investigation offers a strategy for specifically targeting cancer using a newly identified small-molecule inhibitor that targets the Plk1 PBD.

Amphiphilic Triazine Polymer Derivatives as Antibacterial And Anti-atopic Agents in Mice Model.

Considering the emergence of bacterial resistance and low proteolytic stability of antimicrobial peptides (AMPs), herein we developed a series of ultra-short triazine based amphipathic polymers (TZP) that are connected with ethylene diamine linkers instead of protease sensitive amide bond. The most potent oligomers, TZP3 and TZP5 not only displayed potent antibacterial action on various drug-resistant pathogens but also exhibited a strong synergic antibacterial activity in combination with chloramphenicol against multidrug-resistant Pseudomonas aeruginosa (MDRPA). Since most of atopic dermatitis (AD) infections are caused by bacterial colonization, we evaluated the potency of TZP3 and TZP5 on AD in vitro and in vivo. In vitro AD analysis of these two polymers showed significant inhibition against the release of ß-hexosaminidase and tumor necrosis factor (TNF-α) from RBL-2H3 cells. In AD-like skin lesions in BALB/c mice model, these two polymers displayed significant potency in suppressing dermal and epidermal thickness, mast cell infiltration and pro-inflammatory cytokines expression. Moreover, these polymers exhibited remarkable efficacy over the allergies caused by the imbalance of Th1/Th2 by regulating total IgE and IgG2a. Finally, the impact of treatment effects of these polymers was examined through analyzing the weights and sizes of spleen and lymph node of AD-induced mice.

Prolonged MEK inhibition leads to acquired resistance and increased invasiveness in KRAS mutant gastric cancer.

Gastric cancer (GC) is one of the most common causes of cancer-associated death. However, traditional therapeutic strategies have failed to significantly improve the survival of patient with advanced GC. While KRAS mutations have been found in some patients with gastric cancer, an effective therapy to treat KRAS-driven gastric cancer has not been established yet. To provide a rationale for clinical application of kinase inhibitors targeting RAS pathways, we first determined the sensitivity of GC cell lines harboring KRAS mutations or amplification to RAS pathway inhibitors. We found that MAPK pathway inhibitors (MEKi and ERKi) were more effective than AKT inhibitor, suggesting that KRAS-driven gastric cancer cells are dependent on MAPK pathway for survival. Further, we established a KRAS mutant GC cell line with acquired resistance to MEK inhibitors in order to mimic clinical situation of kinase inhibitor resistance. A comprehensive analysis of tyrosine phosphorylation in receptor tyrosine kinases in combination with small molecule chemical library screening revealed upregulated c-MET phosphorylation in this resistance cell line with elevated sensitivity to c-MET TKI (crizotinib) and PI3K/mTOR dual inhibitor (BEZ235). We also showed that migration and invasion of resistant cells were promoted, and crizotinib and BEZ235 could inhibit this malignant phenotype. Overall, our results indicate that prolonged MAPK pathway inhibition could result in acquired resistance which is associated with increased malignant phenotype in KRAS mutant GC and pharmacological targeting c-MET and PI3K/mTOR could overcome this problem.

Kahweol inhibits proliferation and induces apoptosis by suppressing fatty acid synthase in HER2-overexpressing cancer cells.

Kahweol is a coffee-specific diterpene found in the beans of Coffea arabica and has been reported to demonstrate various biological activities, including anti-inflammatory, antioxidant, and apoptotic properties. In the present study, we examined the molecular mechanism of kahweol in human epidermal growth factor receptor-2 (HER2)-overexpressing breast cancer cells. Kahweol preferentially inhibited cell proliferation and induced cell death through the induction of a caspase 3-dependent pathway in HER2-overexpression breast cancer cell lines. Kahweol treatment substantially reduced the levels of HER2 protein, mRNA, and transcriptional activity in SKBR3 cells. Kahweol exerts its potent anticancer efficacy by the upregulation of polyomavirus enhancer activator 3 (PEA3) and downregulation of activator protein 2 (AP-2) to inhibit aberrantly activated HER2 signaling. Fatty acid synthase (FASN) expression and sterol regulatory element-binding protein-1c (SREBP-1c) activity were downregulated by kahweol. In addition, kahweol lowered the levels of phosphorylated Akt and its downstream targets mammalian target of rapamycin (mTOR) and cyclin D1. Furthermore, we found that blocking Akt signaling through kahweol treatment significantly reduced FASN expression and subsequently suppressed cell proliferation in HER2-overexpressing cancer cells. Overall, this study suggests that kahweol could be a useful adjuvant therapeutic agent in the treatment of HER2-overexpressing breast cancer.

A small-molecule inhibitor targeting the AURKC-IκBα interaction decreases transformed growth of MDA-MB-231 breast cancer cells.

The Aurora kinases, Aurora A (AURKA), Aurora B (AURKB), and Aurora C (AURKC), are serine/threonine kinases required for the control of mitosis (AURKA and AURKB) or meiosis (AURKC). Several Aurora kinase inhibitors are being investigated as novel anticancer therapeutics. Recent studies demonstrated that AURKC activation contributes to breast cancer cell transformation. Therefore, AURKC is both a promising marker and therapeutic target for breast cancer; however, its signaling network has not been fully characterized. Using translocation-based cellular assays, we identified IκBα as a binding partner of AURKC, and found that AURKC phosphorylates IκBα at Ser32, thereby activating it. In silico modeling and computational analyses revealed a small-molecule inhibitor (AKCI) that blocked the AURKC-IκBα interaction and exerted antitumor activity in MDA-MB-231 breast cancer cells. Specifically, AKCI induced G2/M cell-cycle arrest through modulation of the p53/p21/CDC2/cyclin B1 pathways. In addition, the drug significantly inhibited MDA-MB-231 cell migration and invasion, as well as decreasing colony formation and tumor growth. Via its interaction with IκBα, AURKC indirectly induced NF-κB activation; accordingly, AKCI decreased PMA-induced activation of NF-κB. Thus, the small-molecule inhibitor AKCI represents a first step towards developing targeted inhibitors of AURKC protein binding, which may lead to further advances in the treatment of breast cancer.

One-Dimensional Supramolecular Nanoplatforms for Theranostics Based on Co-Assembly of Peptide Amphiphiles.

We report a simple and facile strategy for the preparation of multifunctional nanoparticles with programmable properties using self-assembly of precisely designed block amphiphiles in an aqueous solution-state. Versatile, supramolecular nanoplatform for personalized needs, particularly-theranostics, was fabricated by coassembly of peptide amphiphiles (PAs) in aqueous solution, replacing time-consuming and inaccessible chemical synthesis. Fibrils, driven by the assembly of hydrophobic ß-sheet-forming peptide block, were utilized as a nanotemplate for drug loading within their robust core. PAs were tagged with octreotide [somatostatin (SST) analogue] for tumor-targeting or were conjugated with paramagnetic metal ion (Gd3+)-chelating 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) for magnetic resonance (MR) imaging. The two PA types were coassembled to integrate each PA function into original fibrillar nanotemplates. The adoption of a bulky target-specific cyclic octreotide and ß-sheet-forming peptide with enhanced hydrophobicity led to a morphological transition from conventional fibrils to helical fibrils. The resulting one-dimensional nanoaggregates allowed the successful intracellular delivery of doxorubicin (DOX) to MCF-7 cancer cells overexpressing SST receptor (SSTR) and MR imaging by enabling high longitudinal (T1) relaxivity of water protons. Correlation between the structural nature of fibrils formed by PA coassembly and contrast efficacy was elucidated. The coassembly of PAs with desirable functions may thus be a useful strategy for the generation of tailor-made biocompatible nanomaterials.

Bakuchiol sensitizes cancer cells to TRAIL through ROS- and JNK-mediated upregulation of death receptors and downregulation of survival proteins.

We investigated whether bakuchiol, an analog of resveratrol enhances the apoptosis ability of tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) in cancer cells. Bakuchiol enhanced expression of cell death receptor (DR) in TRAIL-sensitive and -resistant colon cancer cells in a dose-dependent manner. A combination of bakuchiol with TRAIL significantly inhibited cell growth of TRAIL sensitive HCT116 and TRAIL resistant HT-29 cells. The expression of TRAIL receptors; DR4 and DR5 was significantly increased by treatment of bakuchiol, however, the expression of survival proteins (e.g., cFLIP, survivin, XIAP and Bcl2) was suppressed. Moreover, the expression of apoptosis related proteins such as cleaved caspase-3, -8, -9 and PARP was increased by combination treatment of bakuchiol and TRAIL. Depletion of DR4 or DR5 by small interfering RNA significantly reversed the cell growth inhibitory effects of bakuchiol in HCT116 and HT-29 cells. Pretreatment with the c-Jun N-terminal kinase (JNK) inhibitor SP600125 and the reactive oxygen species (ROS) scavenger N-acetylcysteine reduced the bakuchiol induced cell growth inhibitory effects. The collective results suggest that bakuchiol facilitates TRAIL-induced apoptosis in colon cancer cells through up-regulation of the TRAIL receptors; DR4 and DR5 via ROS/JNK pathway signals.

Proteomic Insights into Sulfur Metabolism in the Hydrogen-Producing Hyperthermophilic Archaeon Thermococcus onnurineus NA1.

The hyperthermophilic archaeon Thermococcus onnurineus NA1 has been shown to produce H2 when using CO, formate, or starch as a growth substrate. This strain can also utilize elemental sulfur as a terminal electron acceptor for heterotrophic growth. To gain insight into sulfur metabolism, the proteome of T. onnurineus NA1 cells grown under sulfur culture conditions was quantified and compared with those grown under H2-evolving substrate culture conditions. Using label-free nano-UPLC-MSE-based comparative proteomic analysis, approximately 38.4% of the total identified proteome (589 proteins) was found to be significantly up-regulated (≥1.5-fold) under sulfur culture conditions. Many of these proteins were functionally associated with carbon fixation, Fe-S cluster biogenesis, ATP synthesis, sulfur reduction, protein glycosylation, protein translocation, and formate oxidation. Based on the abundances of the identified proteins in this and other genomic studies, the pathways associated with reductive sulfur metabolism, H2-metabolism, and oxidative stress defense were proposed. The results also revealed markedly lower expression levels of enzymes involved in the sulfur assimilation pathway, as well as cysteine desulfurase, under sulfur culture condition. The present results provide the first global atlas of proteome changes triggered by sulfur, and may facilitate an understanding of how hyperthermophilic archaea adapt to sulfur-rich, extreme environments.

Genome Sequence of Arthrobacter sp. MWB30, Isolated from a Crude Oil-Contaminated Seashore.

We report here the draft genome sequence of Arthrobacter sp. MWB30 strain, isolated from a crude oil-contaminated seashore in Tae-an, South Korea, which is able to degrade the crude oil and its derivatives. The draft genome sequence of 4,647,008 bp provides a resource for the identification of crude oil-degrading mechanisms in strain MWB30.

Tumor necrosis factor (TNF) receptor-associated factor (TRAF)-interacting protein (TRIP) negatively regulates the TRAF2 ubiquitin-dependent pathway by suppressing the TRAF2-sphingosine 1-phosphate (S1P) interaction.

The signaling pathway downstream of TNF receptor (TNFR) is involved in the induction of a wide range of cellular processes, including cell proliferation, activation, differentiation, and apoptosis. TNFR-associated factor 2 (TRAF2) is a key adaptor molecule in TNFR signaling complexes that promotes downstream signaling cascades, such as nuclear factor-κB (NF-κB) and mitogen-activated protein kinase activation. TRAF-interacting protein (TRIP) is a known cellular binding partner of TRAF2 and inhibits TNF-induced NF-κB activation. Recent findings that TRIP plays a multifunctional role in antiviral response, cell proliferation, apoptosis, and embryonic development have increased our interest in exploring how TRIP can affect the TNFR-signaling pathway on a molecular level. In our current study, we demonstrated that TRIP is negatively involved in the TNF-induced inflammatory response through the down-regulation of proinflammatory cytokine production. Here, we demonstrated that the TRAF2-TRIP interaction inhibits Lys(63)-linked TRAF2 ubiquitination by inhibiting TRAF2 E3 ubiquitin (Ub) ligase activity. The TRAF2-TRIP interaction inhibited the binding of sphingosine 1-phosphate, which is a cofactor of TRAF2 E3 Ub ligase, to the TRAF2 RING domain. Finally, we demonstrated that TRIP functions as a negative regulator of proinflammatory cytokine production by inhibiting TNF-induced NF-κB activation. These results indicate that TRIP is an important cellular regulator of the TNF-induced inflammatory response.

Endosulfan induces COX-2 expression via NADPH oxidase and the ROS, MAPK, and Akt pathways.

Endosulfan (1,4,5,6,7,7-hexachloro-8,9,10-trinorborn-5-en-2,3-ylenebismet-hylene) is correlated with endocrine disruption, reproductive, and immune dysfunctions. Recently, endosulfan was shown to have an effect on inflammatory pathways, but its influence on cyclooxygenase-2(COX-2) expression is unclear. This study investigated the effects of COX-2 and molecular mechanisms by endosulfan in murine macrophage RAW 264.7 cells. Endosulfan significantly induced COX-2 protein and mRNA levels, as well as COX-2 promoter-driven luciferase activity and the production of prostaglandin E2, a major COX-2 metabolite. Transfection experiments with several human COX-2 promoter constructs revealed that endosulfan activated NF-κB, C/EBP, AP-1, and CREB. Moreover, Akt and mitogen-activated protein kinases (MAPK) were significantly activated by endosulfan. Moreover, endosulfan increased production of the ROS and the ROS-producing NAPDH-oxidase (NOX) family oxidases, NOX2, and NOX3. Endosulfan-induced Akt/MAPK pathways and COX-2 expression were attenuated by DPI, a specific NOX inhibitor, and the ROS scavenger N-acetylcysteine. These results demonstrate that endosulfan induces COX-2 expression via NADPH oxidase, ROS, and Akt/MAPK pathways. These findings provide further insight into the signal transduction pathways involved in the inflammatory effects of endosulfan.

Draft Genome Sequence of Sphingopyxis sp. Strain MWB1, a Crude-Oil-Degrading Marine Bacterium.

Sphingopyxis sp. strain MWB1, which is capable of degrading crude oil, diesel, and kerosene, was isolated from crude oil-contaminated seashore in Tae-an, South Korea. Here, we report the draft genome sequence of this strain, which comprises 3,118,428 bp with a G+C content of 62.85 mol%.

Caveolin-1-dependent and -independent uPAR signaling pathways contribute to ganglioside GT1b induced early apoptosis in A549 lung cancer cells.

Urokinase receptor interacts with α5ß1-integrin and enhances cancer cell proliferation and metastasis. Activation of α5ß1-integrin requires caveolin-1 and is regulated by uPAR, which upregulates persistently the activated ERK necessary for tumor growth. In this study, we show that the ganglioside GT1b induces proapoptotic signaling through two uPAR-ERK signaling pathways in A549 lung cancer cells. GT1b downregulated the expression of α5ß1 integrin, caveolin-1, fibronectin, FAK, and ERK, whereas GT1b upregulated the expression of p53 and uPAR, suggesting GT1b mediated depletion of caveolin-1 in uPAR-expressing A549 cells also disrupts uPAR/integrin complexes, resulting in downregulation of fibronectin-α5ß1-integrin-ERK signaling. Following p53 siRNA treatment, FAK and ERK expression was recovered, meaning the presence of reentry uPAR-FAK-ERK signaling pathway. These findings reveal that GT1b is involved in both caveolin-1-dependent uPAR-α5ß1-integrin-ERK signaling and caveolin-1-independent uPAR-FAK-ERK signaling. These results suggest a novel function of GT1b as a dual regulator of ERK by modulating caveolin-1 and p53.