Phosphoinositide 3-kinase is activated by MUC1 but not responsible for MUC1-induced suppression of Toll-like receptor 5 signaling

2007 ◽  
Vol 293 (3) ◽  
pp. L686-L692 ◽  
Author(s):  
Kosuke Kato ◽  
Wenju Lu ◽  
Hirofumi Kai ◽  
K. Chul Kim
Keyword(s):  
Cross Talk ◽  
Negative Regulator ◽  
Regulatory Subunit ◽  
Binding Motif ◽  
Toll Like Receptor ◽  
Akt Phosphorylation ◽  
Phosphoinositide 3 Kinase ◽  
Toll Like Receptor 5

MUC1 is a membrane-tethered mucin-like glycoprotein expressed on the surface of various mucosal epithelial cells as well as hematopoietic cells. Recently, we showed that MUC1 suppresses flagellin-induced Toll-like receptor (TLR) 5 signaling both in vivo and in vitro through cross talk with TLR5. In this study, we determined whether phosphoinositide 3-kinase (PI3K), a negative regulator of TLR5 signaling, is involved in the cross talk between MUC1 and TLR5 using various genetically modified epithelial cell lines. Our results showed 1) activation of MUC1 induced recruitment of the PI3K regulatory subunit p85 to the MUC1 cytoplasmic tail (CT) as well as Akt phosphorylation, 2) MUC1-induced Akt phosphorylation required the presence of Tyr20 within the PI3K binding motif of the MUC1 CT, and 3) mutation of Tyr20 or pharmacological inhibition of PI3K activation failed to block MUC1-induced suppression of TLR5 signaling. We conclude that whereas PI3K is downstream of MUC1 activation and negatively regulates TLR5 signaling, it is not responsible for MUC1-induced suppression of TLR5 signaling.

scholarly journals MiR-1246 regulates the PI3K/AKT signaling pathway by targeting PIK3AP1 and inhibits thyroid cancer cell proliferation and tumor growth

2021 ◽  
Author(s):  
Jingyan Li ◽  
Zhanlei Zhang ◽  
Jieting Hu ◽  
Xiaoting Wan ◽  
Wei Huang ◽  
...  
Keyword(s):  
Thyroid Cancer ◽  
Tumor Growth ◽  
Signaling Pathway ◽  
Gene Expression Omnibus ◽  
Akt Signaling ◽  
Akt Phosphorylation ◽  
Apoptotic Death ◽  
Phosphoinositide 3 Kinase

AbstractOne of the most prevalent forms of endocrine malignancies is thyroid cancer. Herein, we explored the mechanisms whereby miR-1246 is involved in thyroid cancer. Phosphoinositide 3-kinase adapter protein 1 (PIK3AP1) was identified as a potential miR-1246 target, with the online Gene Expression Omnibus (GEO) database. The binding between miR-1246 and PIK3AP1 and the dynamic role of these two molecules in downstream PI3K/AKT signaling were evaluated. Analysis of GEO data demonstrated significant miR-1246 downregulation in thyroid cancer, and we confirmed that overexpression of miR-1246 can inhibit migratory, invasive, and proliferative activity in vitro and tumor growth in vivo. Subsequent studies indicated that miR-1246 overexpression decreased the protein level of PIK3AP1 and the phosphorylation of PI3K and AKT, which were reversed by PIK3AP1 overexpression. At the same time, overexpression of PIK3AP1 also reversed the miR-1246 mimics-induced inhibition proliferative, migratory, and invasive activity, while promoting increases in apoptotic death, confirming that miR-1246 function was negatively correlated with that of PIK3AP1. Subsequently, we found that the miR-1246 mimics-induced inhibition of PI3K/AKT phosphorylation was reversed by the PI3K/AKT activator IGF-1. miR-1246 mimics inhibited proliferative, migratory, and invasive activity while promoting increases in apoptotic death, which were reversed by IGF-1. Furthermore, miR-1246 agomir can inhibit tumor growth in vivo. We confirmed that miR-1246 affects the signaling pathway of PI3K/AKT via targeting PIK3AP1 and inhibits the development of thyroid cancer. Thus, miR-1246 is a new therapeutic target for thyroid cancer.

Abstract 330: Metallothionein Preservation Of Cardiac Akt2 Signaling By Down-regulating Trb3 Prevents Diabetic Cardiomyopathy

2014 ◽  
Vol 115 (suppl_1) ◽  
Author(s):  
YI TAN ◽  
Xiaoqing Yan ◽  
Shanshan Zhou ◽  
Yong Li ◽  
Yan Li ◽  
...  
Keyword(s):  
Diabetic Cardiomyopathy ◽  
Insulin Signaling ◽  
Glycogen Synthase ◽  
Critical Role ◽  
Negative Regulator ◽  
H9c2 Cells ◽  
Akt Phosphorylation ◽  
Gsk 3Β

Cardiac insulin resistance is a key pathogenic factor for diabetic cardiomyopathy, but its mechanism remains largely unclear. Here we demonstrated that diabetes significantly inhibited cardiac Akt phosphorylation from 2 weeks to 2 months in wide-type (WT) mice, but not in cardiac-specific metallothionein-transgenic (MT-TG) mice. Cardiac Akt2 expression and phosphorylation was decreased and insulin-induced cardiac Akt2 and GSK-3β phosphorylation and glycogen synthase dephosphorylation were also decreased in WT, but not MT-TG, diabetic mice. Deletion of the Akt2 gene either in vitro H9c2 cells or in vivo significantly impaired cardiac glucose metabolic signaling. In addition, diabetes significantly increased cardiac Akt negative regulator tribbles (TRB)3 expression only in WT mice, suggesting the possible contribution of MT inhibition of diabetic up-regulation of TRB3 to Akt2 function preservation. Cardiac H9c2 cells with and without forced MT-overexpression (MT-H9c2) were treated with tert-butyl hydroperoxide (tBHP), which significantly reduced Akt2 phosphorylation in both basal and insulin-stimulating conditions only in H9c2 cells. Silencing TRB3 expression with SiRNA completely prevented tBHP’s inhibition of insulin-stimulated Akt2 phosphorylation in H9c2 cells, while overexpression of TRB3 in MT-H9c2 cells completely abolished MT preservation of insulin-stimulated Akt2 phosphorylation. Forced-overexpression of TRB3 by adenovirus-mediated gene delivery in MT-TG hearts also abolished MT’s preservation of cardiac insulin signaling and prevention of diabetic cardiomyopathy. These results suggest that diabetes-attenuated cardiac Akt2 function via up-regulating TRB3 plays a critical role in diabetic inhibition of insulin signaling in the heart. MT preserved cardiac Akt2-mediated insulin signaling by inhibiting TRB3, leading to the prevention of diabetic cardiomyopathy.

scholarly journals PIP4K2A as a negative regulator of PI3K in PTEN-deficient glioblastoma

2019 ◽  
Vol 216 (5) ◽  
pp. 1120-1134 ◽  
Author(s):  
Yong Jae Shin ◽  
Jason K. Sa ◽  
Yeri Lee ◽  
Donggeon Kim ◽  
Nakho Chang ◽  
...  
Keyword(s):  
Tumor Suppressors ◽  
Cellular Proliferation ◽  
Negative Regulator ◽  
Bona Fide ◽  
Xenograft Models ◽  
Phosphoinositide 3 Kinase ◽  
Multiple Signaling ◽  
First Time

Glioblastoma (GBM) is the most malignant brain tumor with profound genomic alterations. Tumor suppressor genes regulate multiple signaling networks that restrict cellular proliferation and present barriers to malignant transformation. While bona fide tumor suppressors such as PTEN and TP53 often undergo inactivation due to mutations, there are several genes for which genomic deletion is the primary route for tumor progression. To functionally identify putative tumor suppressors in GBM, we employed in vivo RNAi screening using patient-derived xenograft models. Here, we identified PIP4K2A, whose functional role and clinical relevance remain unexplored in GBM. We discovered that PIP4K2A negatively regulates phosphoinositide 3-kinase (PI3K) signaling via p85/p110 component degradation in PTEN-deficient GBMs and specifically targets p85 for proteasome-mediated degradation. Overexpression of PIP4K2A suppressed cellular and clonogenic growth in vitro and impeded tumor growth in vivo. Our results unravel a novel tumor-suppressive role of PIP4K2A for the first time and support the feasibility of combining oncogenomics with in vivo RNAi screen.

scholarly journals MiR-1246 regulates the PI3K/AKT signaling pathway by targeting PIK3AP1 and inhibits thyroid cancer cell proliferation and tumor growth

2021 ◽  
Author(s):  
Jingyan Li ◽  
Zhanlei Zhang ◽  
Jieting Hu ◽  
Xiaoting Wan ◽  
Wei Huang ◽  
...  
Keyword(s):  
Thyroid Cancer ◽  
Tumor Growth ◽  
Signaling Pathway ◽  
Gene Expression Omnibus ◽  
Akt Signaling ◽  
Akt Phosphorylation ◽  
Apoptotic Death ◽  
Phosphoinositide 3 Kinase

Abstract One of the most prevalent forms of endocrine malignancies is thyroid cancer. Herein, we explored the mechanisms whereby miR-1246 is involved in thyroid cancer. Phosphoinositide 3-kinase adapter protein 1 (PIK3AP1) was identified as a potential miR-1246 target,with the online Gene Expression Omnibus (GEO) database. The binding between miR-1246 and PIK3AP1 and the dynamic role of these two molecules in downstream PI3K/AKT signaling were evaluated. Analysis of GEO data demonstrated significant miR-1246 downregulation in thyroid cancer, and we confirmed that overexpression of miR-1246 can inhibit migratory, invasive, and proliferative activity in vivo, and tumor growth in vitro. Subsequent studies indicated that miR-1246 overexpression decreased the protein level of PIK3AP1 and the phosphorylation of PI3K and AKT, which were reversed by PIK3AP1 overexpression. At the same time, overexpression of PIK3AP1 also reversed the miR-1246 mimics-induced inhibition proliferative, migratory, and invasive activity while promoting increases in apoptotic death, confirming that miR-1246 function was negatively correlated with that of PIK3AP1. Subsequently, we found that the miR-1246 mimics-induced inhibition of PI3K/AKT phosphorylation, was reversed by the PI3K/AKT activator IGF-1. miR-1246 mimics inhibited proliferative, migratory, and invasive activity while promoting increases in apoptotic death, which were reversed by IGF-1. Furthermore, miR-1246 agomir can inhibit tumor growth in vivo. We confirmed that miR-1246 affects the signaling pathway of PI3K/AKT via targeting PIK3AP1, and inhibits the development of thyroid cancer. Thus, miR-1246 is a new therapeutic target for thyroid cancer.

scholarly journals Formation of Atroviridin by Hypocrea atroviridis Is Conidiation Associated and Positively Regulated by Blue Light and the G Protein GNA3

2007 ◽  
Vol 6 (12) ◽  
pp. 2332-2342 ◽  
Author(s):  
Monika Komon-Zelazowska ◽  
Torsten Neuhof ◽  
Ralf Dieckmann ◽  
Hans von Döhren ◽  
Alfredo Herrera-Estrella ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Blue Light ◽  
Negative Regulator ◽  
Regulatory Subunit ◽  
Loss Of Function ◽  
Cell Wall Degrading Enzymes ◽  
Kinase A

ABSTRACT Species of the mycoparasitic fungal genus Hypocrea/Trichoderma are prominent producers of peptaibols, a class of small linear peptides of fungal origin. Some of these peptaibols have been shown to act synergistically with cell-wall-degrading enzymes in the inhibition of the growth of other fungi in vitro and in vivo. Here we present the structure of the Hypocrea atroviridis peptaibol synthetase gene (pbs1), deduced from the genome sequence of H. atroviridis. It consists of 19 typical peptide synthetase modules with the required additional modifying domains at the N and C termini. Phylogenetic and similarity analyses of the individual amino acid-activating modules is consistent with its ability to synthesize atroviridins. Matrix-assisted laser desorption ionization-time of flight mass spectrometry of surface-grown cultures of H. atroviridis showed that no peptaibols were formed during vegetative growth, but a microheterogenous mixture of atroviridins accumulated when the colonies started to sporulate. This correlation between sporulation and atroviridin formation was shown to be independent of the pathway inducing sporulation (i.e., light, mechanical injury and carbon starvation, respectively). Atroviridin formation was dependent on the function of the two blue light regulators, BLR1 and BLR2, under some but not all conditions of sporulation and was repressed in a pkr1 (regulatory subunit of protein kinase A) antisense strain with constitutively active protein kinase A. Conversely, however, loss of function of the Gα-protein GNA3, which is a negative regulator of sporulation and leads to a hypersporulating phenotype, fully impairs atroviridin formation. Our data show that formation of atroviridin by H. atroviridis occurs in a sporulation-associated manner but is uncoupled from it at the stage of GNA3.

scholarly journals Toll-like receptor 4 restricts retinal progenitor cell proliferation

2008 ◽  
Vol 183 (3) ◽  
pp. 393-400 ◽  
Author(s):  
Ravid Shechter ◽  
Ayal Ronen ◽  
Asya Rolls ◽  
Anat London ◽  
Sharon Bakalash ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Postnatal Period ◽  
Negative Regulator ◽  
Early Postnatal Period ◽  
Toll Like Receptor ◽  
Tlr4 Expression ◽  
Toll Like Receptor 4 ◽  
Retinal Progenitor

Retinal neurogenesis ceases by the early postnatal period, although retinal progenitor cells (RPCs) persist throughout life. In this study, we show that in the mammalian eye, the function of Toll-like receptor 4 (TLR4) extends beyond regulation of the innate immune response; it restricts RPC proliferation. In TLR4-deficient mice, enhanced proliferation of cells reminiscent of RPCs is evident during the early postnatal period. In vitro experiments demonstrate that TLR4 acts as an intrinsic regulator of RPC fate decision. Increased TLR4 expression in the eye correlates with the postnatal cessation of cell proliferation. However, deficient TLR4 expression is not sufficient to extend the proliferative period but rather contributes to resumption of proliferation in combination with growth factors. Proliferation in vivo is inhibited by both MyD88-dependent and -independent pathways, similar to the mechanisms activated by TLR4 in immune cells. Thus, our study attributes a novel role to TLR4 as a negative regulator of RPC proliferation.

scholarly journals Splicing factor SRSF1 promotes breast cancer progression via oncogenic splice switching of PTPMT1

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Jun-Xian Du ◽  
Yi-Hong Luo ◽  
Si-Jia Zhang ◽  
Biao Wang ◽  
Cong Chen ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Progression ◽  
High Risk Group ◽  
Clinical Samples ◽  
Binding Motif ◽  
Ki 67 ◽  
Tissue Samples

Abstract Background Intensive evidence has highlighted the effect of aberrant alternative splicing (AS) events on cancer progression when triggered by dysregulation of the SR protein family. Nonetheless, the underlying mechanism in breast cancer (BRCA) remains elusive. Here we sought to explore the molecular function of SRSF1 and identify the key AS events regulated by SRSF1 in BRCA. Methods We conducted a comprehensive analysis of the expression and clinical correlation of SRSF1 in BRCA based on the TCGA dataset, Metabric database and clinical tissue samples. Functional analysis of SRSF1 in BRCA was conducted in vitro and in vivo. SRSF1-mediated AS events and their binding motifs were identified by RNA-seq, RNA immunoprecipitation-PCR (RIP-PCR) and in vivo crosslinking followed by immunoprecipitation (CLIP), which was further validated by the minigene reporter assay. PTPMT1 exon 3 (E3) AS was identified to partially mediate the oncogenic role of SRSF1 by the P-AKT/C-MYC axis. Finally, the expression and clinical significance of these AS events were validated in clinical samples and using the TCGA database. Results SRSF1 expression was consistently upregulated in BRCA samples, positively associated with tumor grade and the Ki-67 index, and correlated with poor prognosis in a hormone receptor-positive (HR+) cohort, which facilitated proliferation, cell migration and inhibited apoptosis in vitro and in vivo. We identified SRSF1-mediated AS events and discovered the SRSF1 binding motif in the regulation of splice switching of PTPMT1. Furthermore, PTPMT1 splice switching was regulated by SRSF1 by binding directly to its motif in E3 which partially mediated the oncogenic role of SRSF1 by the AKT/C-MYC axis. Additionally, PTPMT1 splice switching was validated in tissue samples of BRCA patients and using the TCGA database. The high-risk group, identified by AS of PTPMT1 and expression of SRSF1, possessed poorer prognosis in the stage I/II TCGA BRCA cohort. Conclusions SRSF1 exerts oncogenic roles in BRCA partially by regulating the AS of PTPMT1, which could be a therapeutic target candidate in BRCA and a prognostic factor in HR+ BRCA patient.

scholarly journals Targeting Toll-Like Receptor 2: Polarization of Porcine Macrophages by a Mycoplasma-Derived Pam2cys Lipopeptide

Vaccines ◽  
2021 ◽  
Vol 9 (7) ◽  
pp. 692
Author(s):  
Giulia Franzoni ◽  
Antonio Anfossi ◽  
Chiara Grazia De Ciucis ◽  
Samanta Mecocci ◽  
Tania Carta ◽  
...  
Keyword(s):  
Mhc Class Ii ◽  
Macrophage Polarization ◽  
Surface Protein ◽  
Antimicrobial Activities ◽  
Toll Like Receptor ◽  
Activation Markers ◽  
Class I Mhc ◽  
Toll Like Receptor 2

Toll-like receptor 2 (TLR2) ligands are attracting increasing attention as prophylactic and immunotherapeutic agents against pathogens and tumors. We previously observed that a synthetic diacylated lipopeptide based on a surface protein of Mycoplasma agalactiae (Mag-Pam2Cys) strongly activated innate immune cells, including porcine monocyte-derived macrophages (moMΦ). In this study, we utilized confocal microscopy, flow cytometry, multiplex cytokine ELISA, and RT-qPCR to conduct a comprehensive analysis of the effects of scalar doses of Mag-Pam2Cys on porcine moMΦ. We observed enhanced expression of activation markers (MHC class I, MHC class II DR, CD25), increased phagocytotic activity, and release of IL-12 and proinflammatory cytokines. Mag-Pam2Cys also upregulated the gene expression of several IFN-α subtypes, p65, NOS2, and molecules with antimicrobial activities (CD14, beta defensin 1). Overall, our data showed that Mag-Pam2Cys polarized porcine macrophages towards a proinflammatory antimicrobial phenotype. However, Mag-Pam2Cys downregulated the expression of IFN-α3, six TLRs (TLR3, -4, -5, -7, -8, -9), and did not interfere with macrophage polarization induced by the immunosuppressive IL-10, suggesting that the inflammatory activity evoked by Mag-Pam2Cys could be regulated to avoid potentially harmful consequences. We hope that our in vitro results will lay the foundation for the further evaluation of this diacylated lipopeptide as an immunopotentiator in vivo.

scholarly journals 68Ga, 44Sc and 177Lu-labeled AAZTA5-PSMA-617: synthesis, radiolabeling, stability and cell binding compared to DOTA-PSMA-617 analogues

2020 ◽  
Vol 5 (1) ◽  
Author(s):  
Jean-Philippe Sinnes ◽  
Ulrike Bauder-Wüst ◽  
Martin Schäfer ◽  
Euy Sung Moon ◽  
Klaus Kopka ◽  
...  
Keyword(s):  
Human Serum ◽  
Room Temperature ◽  
Solid Phase ◽  
Negative Influence ◽  
Binding Motif ◽  
Lncap Cells ◽  
Cell Binding ◽  
Binding Characteristics

Abstract Background The AAZTA chelator and in particular its bifunctional derivative AAZTA5 was recently investigated to demonstrate unique capabilities to complex diagnostic and therapeutic trivalent radiometals under mild conditions. This study presents a comparison of 68Ga, 44Sc and 177Lu-labeled AAZTA5-PSMA-617 with DOTA-PSMA-617 analogues. We evaluated the radiolabeling characteristics, in vitro stability of the radiolabeled compounds and evaluated their binding affinity and internalization behavior on LNCaP tumor cells in direct comparison to the radiolabeled DOTA-conjugated PSMA-617 analogs. Results AAZTA5 was synthesized in a five-step synthesis and coupled to the PSMA-617 backbone on solid phase. Radiochemical evaluation of AAZTA5-PSMA-617 with 68Ga, 44Sc and 177Lu achieved quantitative radiolabeling of > 99% after less than 5 min at room temperature. Stabilities against human serum, PBS buffer and EDTA and DTPA solutions were analyzed. While there was a small degradation of the 68Ga complex over 2 h in human serum, PBS and EDTA/DTPA, the 44Sc and 177Lu complexes were stable at 2 h and remained stable over 8 h and 1 day. For all three compounds, i.e. [natGa]Ga-AAZTA5-PSMA-617, [natSc]Sc-AAZTA5-PSMA-617 and [natLu]Lu-AAZTA5-PSMA-617, in vitro studies on PSMA-positive LNCaP cells were performed in direct comparison to radiolabeled DOTA-PSMA-617 yielding the corresponding inhibition constants (Ki). Ki values were in the range of 8–31 nM values which correspond with those of [natGa]Ga-DOTA-PSMA-617, [natSc]Sc-DOTA-PSMA-617 and [natLu]Lu-DOTA-PSMA-617, i.e. 5–7 nM, respectively. Internalization studies demonstrated cellular membrane to internalization ratios for the radiolabeled 68Ga, 44Sc and 177Lu-AAZTA5-PSMA-617 tracers (13–20%IA/106 cells) in the same range as the ones of the three radiolabeled DOTA-PSMA-617 tracers (17–20%IA/106 cells) in the same assay. Conclusions The AAZTA5-PSMA-617 structure proved fast and quantitative radiolabeling with all three radiometal complexes at room temperature, excellent stability with 44Sc, very high stability with 177Lu and medium stability with 68Ga in human serum, PBS and EDTA/DTPA solutions. All three AAZTA5-PSMA-617 tracers showed binding affinities and internalization ratios in LNCaP cells comparable with that of radiolabeled DOTA-PSMA-617 analogues. Therefore, the exchange of the chelator DOTA with AAZTA5 within the PSMA-617 binding motif has no negative influence on in vitro LNCaP cell binding characteristics. In combination with the faster and milder radiolabeling features, AAZTA5-PSMA-617 thus demonstrates promising potential for in vivo application for theranostics of prostate cancer.

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