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21 pages, 3921 KiB

Open AccessArticle

by Tarana Siddika, Nileeka Balasuriya, Mallory I. Frederick, Peter Rozik, Ilka U. Heinemann and Patrick O’Donoghue

Cells 2022, 11(23), 3834; https://doi.org/10.3390/cells11233834 - 29 Nov 2022

Cited by 4 | Viewed by 2400

Abstract

Protein kinase B (AKT1) is a serine/threonine kinase and central transducer of cell survival pathways. Typical approaches to study AKT1 biology in cells rely on growth factor or insulin stimulation that activates AKT1 via phosphorylation at two key regulatory sites (Thr308, Ser473), yet [...] Read more.

Protein kinase B (AKT1) is a serine/threonine kinase and central transducer of cell survival pathways. Typical approaches to study AKT1 biology in cells rely on growth factor or insulin stimulation that activates AKT1 via phosphorylation at two key regulatory sites (Thr308, Ser473), yet cell stimulation also activates many other kinases. To produce cells with specific AKT1 activity, we developed a novel system to deliver active AKT1 to human cells. We recently established a method to produce AKT1 phospho-variants from Escherichia coli with programmed phosphorylation. Here, we fused AKT1 with an N-terminal cell penetrating peptide tag derived from the human immunodeficiency virus trans-activator of transcription (TAT) protein. The TAT-tag did not alter AKT1 kinase activity and was necessary and sufficient to rapidly deliver AKT1 protein variants that persisted in human cells for 24 h without the need to use transfection reagents. TAT-pAKT1^T308 induced selective phosphorylation of the known AKT1 substrate GSK-3α, but not GSK-3β, and downstream stimulation of the AKT1 pathway as evidenced by phosphorylation of ribosomal protein S6 at Ser240/244. The data demonstrate efficient delivery of AKT1 with programmed phosphorylation to human cells, thus establishing a cell-based model system to investigate signaling that is dependent on AKT1 activity. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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12 pages, 2634 KiB

Open AccessArticle

mTORC1 Activity in Psoriatic Lesions Is Mediated by Aberrant Regulation through the Tuberous Sclerosis Complex

by Antonio Ferreri, Victoria Lang, Roland Kaufmann and Claudia Buerger

Cells 2022, 11(18), 2847; https://doi.org/10.3390/cells11182847 - 13 Sep 2022

Cited by 3 | Viewed by 1705

Abstract

In the basal, proliferative layer of healthy skin, the mTOR complex 1 (mTORC1) is activated, thus regulating proliferation while preventing differentiation. When cells leave the proliferative, basal compartment, mTORC1 signaling is turned off, which allows differentiation. Under inflammatory conditions, this switch is hijacked [...] Read more.

In the basal, proliferative layer of healthy skin, the mTOR complex 1 (mTORC1) is activated, thus regulating proliferation while preventing differentiation. When cells leave the proliferative, basal compartment, mTORC1 signaling is turned off, which allows differentiation. Under inflammatory conditions, this switch is hijacked by cytokines and prevents proper differentiation. It is currently unknown how mTORC1 is regulated to mediate these effects on keratinocyte differentiation. In other tissues, mTORC1 activity is controlled through various pathways via the tuberous sclerosis complex (TSC). Thus, we investigated whether the TS complex is regulated by proinflammatory cytokines and contributes to the pathogenesis of psoriasis. TNF-α as well as IL-1β induced the phosphorylation of TSC2, especially on S939 via the PI3-K/AKT and MAPK pathway. Surprisingly, increased TSC2 phosphorylation could not be detected in psoriasis patients. Instead, TSC2 was strongly downregulated in lesional psoriatic skin compared to non-lesional skin of the same patients or healthy skin. In vitro inflammatory cytokines induced dissociation of TSC2 from the lysosome, followed by destabilization of the TS complex and degradation. Thus, we assume that in psoriasis, inflammatory cytokines induce strong TSC2 phosphorylation, which in turn leads to its degradation. Consequently, chronic mTORC1 activity impairs ordered keratinocyte differentiation and contributes to the phenotypical changes seen in the psoriatic epidermis. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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21 pages, 2798 KiB

Open AccessArticle

AKT1 Transcriptomic Landscape in Breast Cancer Cells

by Bijesh George, Bin Gui, Rajeswari Raguraman, Aswathy Mary Paul, Harikrishna Nakshatri, Madhavan Radhakrishna Pillai and Rakesh Kumar

Cells 2022, 11(15), 2290; https://doi.org/10.3390/cells11152290 - 25 Jul 2022

Cited by 11 | Viewed by 2783

Abstract

Overexpression and hyperactivation of the serine/threonine protein kinase B (AKT) pathway is one of the most common cellular events in breast cancer progression. However, the nature of AKT1-specific genome-wide transcriptomic alterations in breast cancer cells and breast cancer remains unknown to this point. [...] Read more.

Overexpression and hyperactivation of the serine/threonine protein kinase B (AKT) pathway is one of the most common cellular events in breast cancer progression. However, the nature of AKT1-specific genome-wide transcriptomic alterations in breast cancer cells and breast cancer remains unknown to this point. Here, we delineate the impact of selective AKT1 knock down using gene-specific siRNAs or inhibiting the AKT activity with a pan-AKT inhibitor VIII on the nature of transcriptomic changes in breast cancer cells using the genome-wide RNA-sequencing analysis. We found that changes in the cellular levels of AKT1 lead to changes in the levels of a set of differentially expressed genes and, in turn, imply resulting AKT1 cellular functions. In addition to an expected positive relationship between the status of AKT1 and co-expressed cellular genes, our study unexpectedly discovered an inherent role of AKT1 in inhibiting the expression of a subset of genes in both unstimulated and growth factor stimulated breast cancer cells. We found that depletion of AKT1 leads to upregulation of a subset of genes—many of which are also found to be downregulated in breast tumors with elevated high AKT1 as well as upregulated in breast tumors with no detectable AKT expression. Representative experimental validation studies in two breast cancer cell lines showed a reasonable concurrence between the expression data from the RNA-sequencing and qRT-PCR or data from ex vivo inhibition of AKT1 activity in cancer patient-derived cells. In brief, findings presented here provide a resource for further understanding of AKT1-dependent modulation of gene expression in breast cancer cells and broaden the scope and significance of AKT1 targets and their functions. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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14 pages, 12757 KiB

Open AccessArticle

Extracellular Vesicles from BMSCs Prevent Glucocorticoid-Induced BMECs Injury by Regulating Autophagy via the PI3K/Akt/mTOR Pathway

by Jinhui Ma, Mengran Shen, Debo Yue, Weiguo Wang, Fuqiang Gao and Bailiang Wang

Cells 2022, 11(13), 2104; https://doi.org/10.3390/cells11132104 - 03 Jul 2022

Cited by 10 | Viewed by 2209

Abstract

Osteonecrosis of the femoral head (ONFH) is a common clinical disease with a high disability rate. Injury of bone microvascular endothelial cells (BMECs) caused by glucocorticoid administration is one of the important causes of ONFH, and there is currently a lack of effective [...] Read more.

Osteonecrosis of the femoral head (ONFH) is a common clinical disease with a high disability rate. Injury of bone microvascular endothelial cells (BMECs) caused by glucocorticoid administration is one of the important causes of ONFH, and there is currently a lack of effective clinical treatments. Extracellular vesicles derived from bone stem cells (BMSC-EVs) can prevent ONFH by promoting angiogenesis and can inhibit cell apoptosis by regulating autophagy via the PI3K/Akt/mTOR signaling pathway. The present study aimed to investigate the effect of extracellular vesicles derived from bone marrow stem cells (BMSC) on a glucocorticoid-induced injury of BMECs and possible mechanisms. We found that BMSC-EVs attenuated glucocorticoid-induced viability, angiogenesis capacity injury, and the apoptosis of BMECs. BMSC-EVs increased the LC3 level, but decreased p62 (an autophagy protein receptor) expression, suggesting that BMSC-Exos activated autophagy in glucocorticoid-treated BMECs. The protective effects of BMSC-EVs on the glucocorticoid-induced injury of BMECs was mimicked by a known stimulator of autophagy (rapamycin) and could be enhanced by co-treatment with an autophagy inhibitor (LY294002). BMSC-EVs also suppressed the PI3K/Akt/mTOR signaling pathway, which regulates cell autophagy, in glucocorticoid-treated BMECs. In conclusion, the results indicate that BMSC-EVs prevent the glucocorticoid-induced injury of BMECs by regulating autophagy via the PI3K/Akt/mTOR pathway. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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25 pages, 7715 KiB

Open AccessArticle

PI3K/AKT/mTOR Signaling Pathway Is Required for JCPyV Infection in Primary Astrocytes

by Michael P. Wilczek, Francesca J. Armstrong, Colleen L. Mayberry, Benjamin L. King and Melissa S. Maginnis

Cells 2021, 10(11), 3218; https://doi.org/10.3390/cells10113218 - 18 Nov 2021

Cited by 5 | Viewed by 2298

Abstract

Astrocytes are a main target of JC polyomavirus (JCPyV) in the central nervous system (CNS), where the destruction of these cells, along with oligodendrocytes, leads to the fatal disease progressive multifocal leukoencephalopathy (PML). There is no cure currently available for PML, so it [...] Read more.

Astrocytes are a main target of JC polyomavirus (JCPyV) in the central nervous system (CNS), where the destruction of these cells, along with oligodendrocytes, leads to the fatal disease progressive multifocal leukoencephalopathy (PML). There is no cure currently available for PML, so it is essential to discover antivirals for this aggressive disease. Additionally, the lack of a tractable in vivo models for studying JCPyV infection makes primary cells an accurate alternative for elucidating mechanisms of viral infection in the CNS. This research to better understand the signaling pathways activated in response to JCPyV infection reveals and establishes the importance of the PI3K/AKT/mTOR signaling pathway in JCPyV infection in primary human astrocytes compared to transformed cell lines. Using RNA sequencing and chemical inhibitors to target PI3K, AKT, and mTOR, we have demonstrated the importance of this signaling pathway in JCPyV infection of primary astrocytes not observed in transformed cells. Collectively, these findings illuminate the potential for repurposing drugs that are involved with inhibition of the PI3K/AKT/mTOR signaling pathway and cancer treatment as potential therapeutics for PML, caused by this neuroinvasive virus. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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13 pages, 2238 KiB

Open AccessArticle

GSI Treatment Preserves Protein Synthesis in C2C12 Myotubes

by Joshua R. Huot, Brian Thompson, Charlotte McMullen, Joseph S. Marino and Susan T. Arthur

Cells 2021, 10(7), 1786; https://doi.org/10.3390/cells10071786 - 15 Jul 2021

Cited by 1 | Viewed by 2754

Abstract

It has been demonstrated that inhibiting Notch signaling through γ-secretase inhibitor (GSI) treatment increases myogenesis, AKT/mTOR signaling, and muscle protein synthesis (MPS) in C2C12 myotubes. The purpose of this study was to determine if GSI-mediated effects on myogenesis and MPS are dependent on [...] Read more.

It has been demonstrated that inhibiting Notch signaling through γ-secretase inhibitor (GSI) treatment increases myogenesis, AKT/mTOR signaling, and muscle protein synthesis (MPS) in C2C12 myotubes. The purpose of this study was to determine if GSI-mediated effects on myogenesis and MPS are dependent on AKT/mTOR signaling. C2C12 cells were assessed for indices of myotube formation, anabolic signaling, and MPS following GSI treatment in combination with rapamycin and API-1, inhibitors of mTOR and AKT, respectively. GSI treatment increased several indices of myotube fusion and MPS in C2C12 myotubes. GSI-mediated effects on myotube formation and fusion were completely negated by treatment with rapamycin and API-1. Meanwhile, GSI treatment was able to rescue MPS in C2C12 myotubes exposed to rapamycin or rapamycin combined with API-1. Examination of protein expression revealed that GSI treatment was able to rescue pGSK3β Ser9 despite AKT inhibition by API-1. These findings demonstrate that GSI treatment is able to rescue MPS independent of AKT/mTOR signaling, possibly via GSK3β modulation. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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20 pages, 3554 KiB

Open AccessArticle

Dihydroartemisinin Inhibits mTORC1 Signaling by Activating the AMPK Pathway in Rhabdomyosarcoma Tumor Cells

by Jun Luo, Yoshinobu Odaka, Zhu Huang, Bing Cheng, Wang Liu, Lin Li, Chaowei Shang, Chao Zhang, Yang Wu, Yan Luo, Shengyong Yang, Peter J. Houghton, Xiaofeng Guo and Shile Huang

Cells 2021, 10(6), 1363; https://doi.org/10.3390/cells10061363 - 01 Jun 2021

Cited by 6 | Viewed by 3651

Abstract

Dihydroartemisinin (DHA), an anti-malarial drug, has been shown to possess potent anticancer activity, partly by inhibiting the mammalian target of rapamycin (mTOR) complex 1 (mTORC1) signaling. However, how DHA inhibits mTORC1 is still unknown. Here, using rhabdomyosarcoma (RMS) as a model, we found [...] Read more.

Dihydroartemisinin (DHA), an anti-malarial drug, has been shown to possess potent anticancer activity, partly by inhibiting the mammalian target of rapamycin (mTOR) complex 1 (mTORC1) signaling. However, how DHA inhibits mTORC1 is still unknown. Here, using rhabdomyosarcoma (RMS) as a model, we found that DHA reduced cell proliferation and viability in RMS cells, but not those in normal cells, which was associated with inhibition of mTORC1. Mechanistically, DHA did not bind to mTOR or FK506 binding protein 12 (FKBP12). In addition, DHA neither inhibited insulin-like growth factor-1 receptor (IGF-1R), phosphoinositide 3-kinase (PI3K), and extracellular signal-regulated kinase ½ (Erk1/2), nor activated phosphatase and tensin homolog (PTEN) in the cells. Rather, DHA activated AMP-activated protein kinase (AMPK). Pharmacological inhibition of AMPK, ectopic expression dominant negative or kinase-dead AMPK, or knockdown of AMPKa attenuated the inhibitory effect of DHA on mTORC1 in the cells. Additionally, DHA was able to induce dissociation of regulatory-associated protein of mTOR (raptor) from mTOR and inhibit mTORC1 activity. Moreover, treatment with artesunate, a prodrug of DHA, dose-dependently inhibited tumor growth and concurrently activated AMPK and suppressed mTORC1 in RMS xenografts. The results indicated that DHA inhibits mTORC1 by activating AMPK in tumor cells. Our finding supports that DHA or artesunate has a great potential to be repositioned for treatment of RMS. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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17 pages, 4272 KiB

Open AccessArticle

mTORC2 Is Involved in the Induction of RSK Phosphorylation by Serum or Nutrient Starvation

by Po-Chien Chou, Swati Rajput, Xiaoyun Zhao, Chadni Patel, Danielle Albaciete, Won Jun Oh, Heineken Queen Daguplo, Nikhil Patel, Bing Su, Guy Werlen and Estela Jacinto

Cells 2020, 9(7), 1567; https://doi.org/10.3390/cells9071567 - 27 Jun 2020

Cited by 6 | Viewed by 3751

Abstract

Cells adjust to nutrient fluctuations to restore metabolic homeostasis. The mechanistic target of rapamycin (mTOR) complex 2 responds to nutrient levels and growth signals to phosphorylate protein kinases belonging to the AGC (Protein Kinases A,G,C) family such as Akt and PKC. Phosphorylation of [...] Read more.

Cells adjust to nutrient fluctuations to restore metabolic homeostasis. The mechanistic target of rapamycin (mTOR) complex 2 responds to nutrient levels and growth signals to phosphorylate protein kinases belonging to the AGC (Protein Kinases A,G,C) family such as Akt and PKC. Phosphorylation of these AGC kinases at their conserved hydrophobic motif (HM) site by mTORC2 enhances their activation and mediates the functions of mTORC2 in cell growth and metabolism. Another AGC kinase family member that is known to undergo increased phosphorylation at the homologous HM site (Ser380) is the p90 ribosomal S6 kinase (RSK). Phosphorylation at Ser380 is facilitated by the activation of the mitogen-activated protein kinase/extracellular signal regulated kinase (MAPK/ERK) in response to growth factor stimulation. Here, we demonstrate that optimal phosphorylation of RSK at this site requires an intact mTORC2. We also found that RSK is robustly phosphorylated at Ser380 upon nutrient withdrawal or inhibition of glycolysis, conditions that increase mTORC2 activation. However, pharmacological inhibition of mTOR did not abolish RSK phosphorylation at Ser380, indicating that mTOR catalytic activity is not required for this phosphorylation. Since RSK and SIN1β colocalize at the membrane during serum restimulation and acute glutamine withdrawal, mTORC2 could act as a scaffold to enhance RSK HM site phosphorylation. Among the known RSK substrates, the CCTβ subunit of the chaperonin containing TCP-1 (CCT) complex had defective phosphorylation in the absence of mTORC2. Our findings indicate that the mTORC2-mediated phosphorylation of the RSK HM site could confer RSK substrate specificity and reveal that RSK responds to nutrient fluctuations. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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23 pages, 6019 KiB

Open AccessArticle

Mutation-Associated Phenotypic Heterogeneity in Novel and Canonical PIK3CA Helical and Kinase Domain Mutants

by Arman Ali Ghodsinia, J-Ann Marie T. Lego and Reynaldo L. Garcia

Cells 2020, 9(5), 1116; https://doi.org/10.3390/cells9051116 - 30 Apr 2020

Cited by 6 | Viewed by 4683

Abstract

Phosphatidylinositol 3-kinase, catalytic subunit alpha (PIK3CA) is an oncogene often mutated in colorectal cancer (CRC). The contribution of PIK3CA mutations in acquired resistance to anti-epidermal growth factor receptor (EGFR) therapy is well documented, but their prognostic and predictive value remain unclear. Domain- and [...] Read more.

Phosphatidylinositol 3-kinase, catalytic subunit alpha (PIK3CA) is an oncogene often mutated in colorectal cancer (CRC). The contribution of PIK3CA mutations in acquired resistance to anti-epidermal growth factor receptor (EGFR) therapy is well documented, but their prognostic and predictive value remain unclear. Domain- and exon-specific mutations are implicated in either favorable or poor prognoses, but there is paucity in the number of mutations characterized outside of the mutational hotspots. Here, two novel non-hotspot mutants—Q661K in exon 13 and C901R in exon 19—were characterized alongside the canonical exon 9 E545K and exon 20 H1047R mutants in NIH3T3 and HCT116 cells. Q661K and E545K both map to the helical domain, whereas C901R and H1047R map to the kinase domain. Results showed variable effects of Q661K and C901R on morphology, cellular proliferation, apoptosis resistance, and cytoskeletal reorganization, with both not having any effect on cellular migration. In comparison, E545K markedly promoted proliferation, survival, cytoskeletal reorganization, migration, and spheroid formation, whereas H1047R only enhanced the first three. In silico docking suggested these mutations negatively affect binding of the p85 alpha regulatory subunit to PIK3CA, thereby relieving PIK3CA inhibition. Altogether, these findings support intra-domain and mutation-specific variability in oncogenic readouts, with implications in degree of aggressiveness. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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17 pages, 9108 KiB

Open AccessArticle

Topical Ascorbic Acid Ameliorates Oxidative Stress-Induced Corneal Endothelial Damage via Suppression of Apoptosis and Autophagic Flux Blockage

by Yi-Jen Hsueh, Yaa-Jyuhn James Meir, Lung-Kun Yeh, Tze-Kai Wang, Chieh-Cheng Huang, Tsai-Te Lu, Chao-Min Cheng, Wei-Chi Wu and Hung-Chi Chen

Cells 2020, 9(4), 943; https://doi.org/10.3390/cells9040943 - 11 Apr 2020

Cited by 20 | Viewed by 3207

Abstract

Compromised pumping function of the corneal endothelium, due to loss of endothelial cells, results in corneal edema and subsequent visual problems. Clinically and experimentally, oxidative stress may cause corneal endothelial decompensation after phacoemulsification. Additionally, in vitro and animal studies have demonstrated the protective [...] Read more.

Compromised pumping function of the corneal endothelium, due to loss of endothelial cells, results in corneal edema and subsequent visual problems. Clinically and experimentally, oxidative stress may cause corneal endothelial decompensation after phacoemulsification. Additionally, in vitro and animal studies have demonstrated the protective effects of intraoperative infusion of ascorbic acid (AA). Here, we established a paraquat-induced cell damage model, in which paraquat induced reactive oxygen species (ROS) production and apoptosis in the B4G12 and ARPE-19 cell lines. We demonstrate that oxidative stress triggered autophagic flux blockage in corneal endothelial cells and that addition of AA ameliorated such oxidative damage. We also demonstrate the downregulation of Akt phosphorylation in response to oxidative stress. Pretreatment with ascorbic acid reduced the downregulation of Akt phosphorylation, while inhibition of the PI3K/Akt pathway attenuated the protective effects of AA. Further, we establish an in vivo rabbit model of corneal endothelial damage, in which an intracameral infusion of paraquat caused corneal opacity. Administration of AA via topical application increased its concentration in the corneal stroma and reduced oxidative stress in the corneal endothelium, thereby promoting corneal clarity. Our findings indicate a perioperative strategy of topical AA administration to prevent oxidative stress-induced damage, particularly for those with vulnerable corneal endothelia. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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16 pages, 3759 KiB

Open AccessArticle

Galectin-1 Overexpression Activates the FAK/PI3K/AKT/mTOR Pathway and Is Correlated with Upper Urinary Urothelial Carcinoma Progression and Survival

by Yu-Li Su, Hao-Lun Luo, Chun-Chieh Huang, Ting-Ting Liu, Eng-Yen Huang, Ming-Tse Sung, Jen-Jie Lin, Po-Hui Chiang, Yen-Ta Chen, Chih-Hsiung Kang and Yuan-Tso Cheng

Cells 2020, 9(4), 806; https://doi.org/10.3390/cells9040806 - 26 Mar 2020

Cited by 24 | Viewed by 4541

Abstract

Galectin-1 (GAL1) is a β-galactoside-binding protein involved in multiple aspects of tumorigenesis. However, the biological role of GAL1 in upper tract urothelial carcinoma (UTUC) has not been entirely understood. Herein, we investigated the oncological effects of GAL1 expression in tumor specimens and identified [...] Read more.

Galectin-1 (GAL1) is a β-galactoside-binding protein involved in multiple aspects of tumorigenesis. However, the biological role of GAL1 in upper tract urothelial carcinoma (UTUC) has not been entirely understood. Herein, we investigated the oncological effects of GAL1 expression in tumor specimens and identified related gene alterations through molecular analysis of GAL1. Clinical parameter data and tumor specimens were collected from 86 patients with pT3N0M0 UTUC who had undergone radical nephroureterectomy. We analyzed the difference in survival by using Kaplan–Meier analyses and Cox proportional regression models and in GAL1 expression by using immunohistochemical (IHC) methods. Public genomic data from the Cancer Genome Atlas (TCGA) and GSE32894 data sets were analyzed for comparison. Using four urothelial carcinoma (UC) cell lines (BFTC-909, T24, RT4, and J82) as in vitro models, we evaluated the functions of GAL1 in UC cell growth, invasiveness, and migration and its role in downstream signaling pathways. The study population was classified into two groups, GAL1-high (n = 35) and GAL1-low (GAL1 n = 51), according to IHC interpretation. Univariate analysis revealed that high GAL1 expression was significantly associated with poor recurrence-free survival (RFS; p = 0.028) and low cancer-specific survival (CSS; p = 0.025). Multivariate analysis revealed that GAL1-high was an independent predictive factor for RFS (hazard ratio (HR) 2.43; 95% confidence interval (CI) 1.17–5.05, p = 0.018) and CSS (HR 4.04; 95% CI 1.25–13.03, p = 0.019). In vitro studies revealed that GAL1 knockdown significantly reduced migration and invasiveness in UTUC (BFTC-909) and bladder cancer cells (T24). GAL1 knockdown significantly reduced protein levels of matrix metalloproteinase-2 (MMP-2) and MMP-9, which increased tissue inhibitor of metalloproteinase-1 (TIMP-1) and promoted epithelial–mesenchymal transition (EMT). Through gene expression microarray analysis of GAL1 vector and GAL1-KD cells, we identified multiple significant signaling pathways including p53, Forkhead box O (FOXO), and phosphoinositide 3-kinase/protein kinase B (PI3K/AKT). We validated microarray results through immunoblotting, thus proving that downregulation of GAL1 reduced focal adhesion kinase (FAK), p-PI3K, p-AKT, and p-mTOR expression. We concluded that GAL1 expression was highly related to oncological survival in patients with locally advanced UTUC. GAL1 promoted UC invasion and metastasis by activating the FAK/PI3K/AKT/mTOR pathway. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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19 pages, 3005 KiB

Open AccessArticle

Elevated PDK1 Expression Drives PI3K/AKT/MTOR Signaling Promotes Radiation-Resistant and Dedifferentiated Phenotype of Hepatocellular Carcinoma

by Oluwaseun Adebayo Bamodu, Hang-Lung Chang, Jiann-Ruey Ong, Wei-Hwa Lee, Chi-Tai Yeh and Jo-Ting Tsai

Cells 2020, 9(3), 746; https://doi.org/10.3390/cells9030746 - 18 Mar 2020

Cited by 91 | Viewed by 7395

Abstract

Resistance to radiotherapy (IR), with consequent disease recurrence, continues to limit the efficacy of contemporary anticancer treatment for patients with hepatocellular carcinoma (HCC), especially in late stage. Despite accruing evidence implicating the PI3K/AKT signaling pathway in cancer-promoting hypoxia, cancerous cell proliferation and radiotherapy-resistance, [...] Read more.

Resistance to radiotherapy (IR), with consequent disease recurrence, continues to limit the efficacy of contemporary anticancer treatment for patients with hepatocellular carcinoma (HCC), especially in late stage. Despite accruing evidence implicating the PI3K/AKT signaling pathway in cancer-promoting hypoxia, cancerous cell proliferation and radiotherapy-resistance, it remains unclear which molecular constituent of the pathway facilitates adaptation of aggressive HCC cells to tumoral stress signals and drives their evasion of repeated IR-toxicity. This present study investigated the role of PDK1 signaling in IR-resistance, enhanced DNA damage repair and post-IR relapse, characteristic of aggressive HCC cells, while exploring potential PDK1-targetability to improve radiosensitivity. The study employed bioinformatics analyses of gene expression profile and functional protein–protein interaction, generation of IR-resistant clones, flow cytometry-based ALDH activity and side-population (SP) characterization, siRNA-mediated loss-of-PDK1function, western-blotting, immunohistochemistry and functional assays including cell viability, migration, invasion, clonogenicity and tumorsphere formation assays. We showed that the aberrantly expressed PDK1 characterizes poorly differentiated HCC CVCL_7955, Mahlavu, SK-HEP1 and Hep3B cells, compared to the well-differentiated Huh7 or normal adult liver epithelial THLE-2 cells, and independently activates the PI3K/AKT/mTOR signaling. Molecular ablation of PDK1 function enhanced susceptibility of HCC cells to IR and was associated with deactivated PI3K/AKT/mTOR signaling. Additionally, PDK1-driven IR-resistance positively correlated with activated PI3K signaling, enhanced HCC cell motility and invasiveness, augmented EMT, upregulated stemness markers ALDH1A1, PROM1, SOX2, KLF4 and POU5F1, increased tumorsphere-formation efficiency and suppressed biomarkers of DNA damage—RAD50, MSH3, MLH3 and ERCC2. Furthermore, the acquired IR-resistant phenotype of Huh7 cells was strongly associated with significantly increased ALDH activity, SP-enrichment, and direct ALDH1-PDK1 interaction. Moreover, BX795-mediated pharmacological inhibition of PDK1 synergistically enhances the radiosensitivity of erstwhile resistant cells, increased Bax/Bcl-2 apoptotic ratio, while suppressing oncogenicity and clonogenicity. We provide preclinical evidence implicating PDK1 as an active driver of IR-resistance by activation of the PI3K/AKT/mTOR signaling, up-modulation of cancer stemness signaling and suppression of DNA damage, thus, projecting PDK1-targeting as a putative enhancer of radiosensitivity and a potential new therapeutic approach for patients with IR-resistant HCC. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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Review

Jump to: Research

35 pages, 2201 KiB

Open AccessReview

Insight into the Role of the PI3K/Akt Pathway in Ischemic Injury and Post-Infarct Left Ventricular Remodeling in Normal and Diabetic Heart

by Bartosz Walkowski, Marcin Kleibert, Miłosz Majka and Małgorzata Wojciechowska

Cells 2022, 11(9), 1553; https://doi.org/10.3390/cells11091553 - 05 May 2022

Cited by 35 | Viewed by 7308

Abstract

Despite the significant decline in mortality, cardiovascular diseases are still the leading cause of death worldwide. Among them, myocardial infarction (MI) seems to be the most important. A further decline in the death rate may be achieved by the introduction of molecularly targeted [...] Read more.

Despite the significant decline in mortality, cardiovascular diseases are still the leading cause of death worldwide. Among them, myocardial infarction (MI) seems to be the most important. A further decline in the death rate may be achieved by the introduction of molecularly targeted drugs. It seems that the components of the PI3K/Akt signaling pathway are good candidates for this. The PI3K/Akt pathway plays a key role in the regulation of the growth and survival of cells, such as cardiomyocytes. In addition, it has been shown that the activation of the PI3K/Akt pathway results in the alleviation of the negative post-infarct changes in the myocardium and is impaired in the state of diabetes. In this article, the role of this pathway was described in each step of ischemia and subsequent left ventricular remodeling. In addition, we point out the most promising substances which need more investigation before introduction into clinical practice. Moreover, we present the impact of diabetes and widely used cardiac and antidiabetic drugs on the PI3K/Akt pathway and discuss the molecular mechanism of its effects on myocardial ischemia and left ventricular remodeling. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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31 pages, 3380 KiB

Open AccessReview

SEA and GATOR 10 Years Later

by Yahir A. Loissell-Baltazar and Svetlana Dokudovskaya

Cells 2021, 10(10), 2689; https://doi.org/10.3390/cells10102689 - 08 Oct 2021

Cited by 5 | Viewed by 3386

Abstract

The SEA complex was described for the first time in yeast Saccharomyces cerevisiae ten years ago, and its human homologue GATOR complex two years later. During the past decade, many advances on the SEA/GATOR biology in different organisms have been made that allowed [...] Read more.

The SEA complex was described for the first time in yeast Saccharomyces cerevisiae ten years ago, and its human homologue GATOR complex two years later. During the past decade, many advances on the SEA/GATOR biology in different organisms have been made that allowed its role as an essential upstream regulator of the mTORC1 pathway to be defined. In this review, we describe these advances in relation to the identification of multiple functions of the SEA/GATOR complex in nutrient response and beyond and highlight the consequence of GATOR mutations in cancer and neurodegenerative diseases. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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16 pages, 9446 KiB

Open AccessReview

TFEB Biology and Agonists at a Glance

by Mingyue Chen, Yashuang Dai, Siyu Liu, Yuxin Fan, Zongxian Ding and Dan Li

Cells 2021, 10(2), 333; https://doi.org/10.3390/cells10020333 - 05 Feb 2021

Cited by 39 | Viewed by 9214

Abstract

Autophagy is a critical regulator of cellular survival, differentiation, development, and homeostasis, dysregulation of which is associated with diverse diseases including cancer and neurodegenerative diseases. Transcription factor EB (TFEB), a master transcriptional regulator of autophagy and lysosome, can enhance autophagic and lysosomal biogenesis [...] Read more.

Autophagy is a critical regulator of cellular survival, differentiation, development, and homeostasis, dysregulation of which is associated with diverse diseases including cancer and neurodegenerative diseases. Transcription factor EB (TFEB), a master transcriptional regulator of autophagy and lysosome, can enhance autophagic and lysosomal biogenesis and function. TFEB has attracted a lot of attention owing to its ability to induce the intracellular clearance of pathogenic factors in a variety of disease models, suggesting that novel therapeutic strategies could be based on the modulation of TFEB activity. Therefore, TFEB agonists are a promising strategy to ameliorate diseases implicated with autophagy dysfunction. Recently, several TFEB agonists have been identified and preclinical or clinical trials are applied. In this review, we present an overview of the latest research on TFEB biology and TFEB agonists. Full article

(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)

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