DOCSLIB.ORG

SSH3 Promotes Malignant Progression of HCC by Activating FGF1-Mediated FGF/FGFR Pathway

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
SSH3 Promotes Malignant Progression of HCC by Activating FGF1-Mediated FGF/FGFR Pathway European Review for Medical and Pharmacological Sciences 2020; 24: 11561-11568 SSH3 promotes malignant progression of HCC by activating FGF1-mediated FGF/FGFR pathway Q.-S. SHI, Y.-H. ZHANG, J. LONG, Z.-L. QIAN, C.-X. HU Department of Oncology Minimally Invasive Interventional Radiology, Beijing Youan Hospital, Capital Medical University, Beijing, China Abstract. – OBJECTIVE: To investigate the Introduction impact of silencing SSH3 on the expression of FGF/FGFR pathway-related genes FGF1, FG- As the third largest cancer killer in the world, FR1, and FGFR2 in hepatocellular carcinoma the mortality and morbidity of hepatocellular car- (HCC) cell line, so as to further understand the role of SSH3 in proliferation and apoptosis of cinoma (HCC) are increasing year by year around 1,2 HCC cells. the world . In China, it has also become one of PATIENTS AND METHODS: TWe first de- the most common tumors with the highest degree tected SSH3 expression in 51 pairs of tumor of malignancy, and more than 300,000 patients tissue specimens and adjacent tissues collect- die of this cancer every year, which is related to ed from HCC patients through quantitative Re- its biological characteristics of prone to recur- al Time-Polymerase Chain Reaction (qRT-PCR) 3,4 and analyzed the interplay between SSH3 ex- rence and metastasis . The development of HCC pression and clinical characteristics of HCC is a complex process involving multiple genes in patients. In vitro, after SSH3-silenced human vivo and coordinated by multiple steps5,6. At pres- HCC cell line was constructed by lentiviral trans- ent, surgery is one of the most effective methods fection, Cell Counting Kit-8 (CCK-8), cell cloning for HCC treatment, however, the high incidence assay, and flow apoptosis methods were con- of intrahepatic metastasis and vascular invasion ducted to explore the HCC cell functions. Final- and the lack of effective non-surgical treatment, ly, whether SSH3 exerts its biological character- 7,8 istics through the FGF/FGFR pathway and the result in poor prognosis of most HCC patients . mutual regulation mechanism between SSH3 Therefore, it is urgent to carry out in-depth re- and FGF1 were further uncovered. search on the mechanism of HCC progression so RESULTS: It was found that SSH3 expression as to provide basis for identifying new molecular was remarkably higher in tumor tissues of HCC targets and developing new targeted drugs9,10. patients than that in normal tissues. Meanwhile, The Slingshot family includes SSH1, SSH2, in comparison to patients with low expression of 11,12 13 SSH3, patients with high expression of SSH3 had and SSH3 . Lu et al have found that both SSH1 higher pathological grade and larger tumor size. and SSH2 have phosphatase activity in vitro, In addition, after silencing SSH3, HCC cell prolif- which can dephosphorylate phosphorylation, while eration ability was attenuated while the apoptosis SSH3 does not show this activity. In addition, lit- ability was enhanced in comparison to the con- erature has reported that that both SSH1 and SSH2 trol group. Moreover, the protein levels of FGF1/ can be co-located with micro-filaggrin, while the FGFR pathway-related genes FGF1, FGFR1, and localization relationship between SSH3 and mi- FGFR2 were markedly inhibited by the downreg- 14 ulation of SSH3. Meanwhile, cell recovery exper- cro-filaggrin is not clear . Currently, there are still iment demonstrated that the overexpression of very few studies on mechanisms whereby SSH3 FGF1 reversed the impact of SSH3 silencing on is engaged in tumor development, and only one the proliferation and apoptosis of HCC cells. article reported that SSH3 can accelerate metasta- CONCLUSIONS: In summary, SSH3 is capa- sis of colorectal tumor cells via affecting LIMK1/ ble of accelerating the malignant progression Rac1 signal transduction pathway15. Therefore, this of HCC by activating FGF1-mediated FGF/FGFR pathway, thus becoming a new molecular target study intends to specify the impact of SSH3 on for HCC therapy. HCC cell functions by constructing a recombinant adenovirus of SSH3 and using an in vitro SSH3- Key Words: knocked down cell model, so as to further clarify SSH3, FGF1, Hepatocellular carcinoma, Malignant the molecular pathogenic mechanism of SSH3 par- progression. ticipating in the occurrence of HCC. Corresponding Author: Jiang Long, MD; e-mail: [email protected] 11561 Q.-S. Shi, Y.-H. Zhang, J. Long, Z.-L. Qian, C.-X. Hu Patients and Methods tion (Beyotime, Shanghai, China) for 20 min- utes, washed 3 times with phosphate-buffered Patients and HCC Samples saline (PBS), photographed, and counted under a 51 cases of HCC tumor tissue specimens and light-selective environment. paired adjacent ones were obtained from HCC patients aged from 30 to 78 in our hospital. All Flow Cytometry Analysis cases were diagnosed by two senior directors of The method of binding with Annexin V-flu- pathologists. This study was approved by the Eth- orescein isothiocyanate (FITC; Merck, Biller- ics Committee of Beijing Youan Hospital, Capi- ica, MA, USA) and propidium iodide (PI) was tal Medical University. Signed written informed used to be detected by flow cytometry. The cell consents were obtained from all participants be- density was adjusted to about 1×106 cells/mL. fore the study. After the medium was discarded, the cells were washed twice with PBS and gently resuspended Cell Lines and Reagents with 0.5 mL of pre-cooled 1 × binding buffer, Six human HCC cell lines Bel-7402, HepG2, and then, 1.25 Ul Annexin V-FITC was added MHCC88H, SMMC-7221 and Huh7, Hep3B, and for incubation at room temperature and light- one human normal liver cell line LO2 [American proof reaction for 15 min. Subsequently, the cells Type Culture Collection (ATCC; Manassas, VA, were centrifuged at 1000 × g for 5 min at room USA)] used in this study were cultured in Dul- temperature, and the supernatant was removed. becco’s Modified Eagle’s Medium (DMEM) high After gently resuspending the cells with 0.5 mL glucose medium (Life Technologies, Gaithers- of pre-cooled l × binding buffer, 10 Ul PI was burg, MD, USA) supplemented with 10% fetal bo- added, and the sample was placed on ice and vine serum (FBS; Gibco, Rockville, MD, USA), stored in the dark, and then, immediately ana- penicillin (100 U/mL; Thermo Fisher Scientific, lyzed by flow cytometry (BD, Franklin Lakes, Waltham, MA, USA), and streptomycin (100 μg/ NJ, USA). mL; Thermo Fisher Scientific, Waltham, MA, USA) in a 37°C, 5% CO2 incubator. QRT-PCR Assay Total RNA was extracted from HCC cell Transfection lines and tissues using TRIzol reagent (Invit- sh-NC and sh-SSH3 containing the silent rogen, Carlsbad, CA, USA). SSH3 and FGF1 SSH3 lentiviral sequence (Shanghai GenePhar- quantitation were performed by SYBR® Premix ma Company, Shanghai, China) were used in Ex TaqTM, with GaphPad as internal standard the transfection experiments. When cell density (TaKaRa, Otsu, Shiga, Japan). The primers are: reached 30-40%, lentiviral transfection was car- SSH3 F: 5’-TCCAGGTATTGCACCAAGC-3’; ried out according to the manufacturer’s instruc- R: 5’-GCCATAGCCGTCCACTCAT-3’. FGF1 tions. After 48 h, the stale cell lines were harvest- F: 5’-CCCCGTCAGATAATCTGTG-3’; R: ed for quantitative Real Time-Polymerase Chain 5’-CTTGTCAGATACGGGAGG-3’. Glyceral- Reaction (qRT-PCR) analysis and cell functional dehyde 3-phosphate dehydrogenase (GAPDH) assays. F: 5’-GGUGACUAUUCAACCGCAUTT-3’; R: 5’-AUGCGGUUGAAUAGUCACCTT-3’. Cell Proliferation Assay The transfected cells were seeded in 96-well Western Blot plates (2000 cells/well) with 100 μL culture medi- The transfected cells were lysed using cell ly- um. Cell Counting Kit-8 (CCK-8) assay was car- sis buffer, shaken on ice for 30 minutes, and cen- ried out according to the manufacturer’s protocol trifuged at 14,000 × g for 15 minutes at 4°C. The (Dojindo Molecular Technologies, Kumamoto, total protein concentration was calculated by bi- Japan). cinchoninic acid (BCA) Protein Assay Kit (Pierce, Rockford, IL, USA). The extracted proteins were Colony Formation Assay separated using a 10% Sodium Dodecyl Sulfate After 48 h of transfection, the cells were cul- Polyacrylamide Gel Electrophoresis (SDS-PAGE) tured with complete medium for 2 weeks, and and subsequently transferred to a polyvinylidene then, fixed in 2 mL of methanol for 20 minutes. difluoride (PVDF) membrane (Millipore, Billeri- After the methanol was aspirated, the cells were ca, MA, USA). Immunoblotting was carried out stained with 0.1% crystal violet staining solu- based on instructions. 11562 SSH3 and hepatocellular carcinoma Dual-Luciferase Reporting I. QPCR analysis revealed that in comparison to Assay the paracancerous normal tissues, the HCC tumor Cells were seeded in 24-well plates and tissues contained a higher SSH3 expression (Fig- co-transfected with SSH3 overexpression and ure 1A), suggesting that SSH3 may serve as an blank vector and pMIR Luciferase reporter plas- oncogene in HCC. Similarly, in vitro, SSH3 also mid (Yeasen, Shanghai, China). The plasmid was showed a higher expression in HCC cell lines Bel- then introduced into the cells using Lipofect- 7402, HepG2, MHCC88H, SMMC-7221, Huh7, amine 2000 (Invitrogen, Carlsbad, CA, USA). and Hep3B, especially in Hub7 and HepG2, than After 48 h, the reporter Luciferase activity was in LO2 cells (Figure 1B). normalized. Additionally, according to the expression of SSH3, we divided the above tumor tissue speci- Statistical Analysis mens into high and low groups to clarify the as- GraphPad Prism 5 V5.01 software (La Jol- sociation between SSH3 levels and some clinical la, CA, USA) was applied to perform statistical indicators, including pathological stage, tumor analysis. The differences between the two groups size, incidence of lymph node or distant metas- were analyzed by using the Student’s t-test. Com- tasis, age, and gender of HCC patients. As a re- parison between multiple groups was done using sult, pathological stage and tumor size (Figure one-way ANOVA test followed by post-hoc test 1C) showed a marked difference between the two (Least Significant Difference).
Load more

Recommended publications
  • A Computational Approach for Defining a Signature of Β-Cell Golgi Stress in Diabetes Mellitus
    Page 1 of 781 Diabetes A Computational Approach for Defining a Signature of β-Cell Golgi Stress in Diabetes Mellitus Robert N. Bone1,6,7, Olufunmilola Oyebamiji2, Sayali Talware2, Sharmila Selvaraj2, Preethi Krishnan3,6, Farooq Syed1,6,7, Huanmei Wu2, Carmella Evans-Molina 1,3,4,5,6,7,8* Departments of 1Pediatrics, 3Medicine, 4Anatomy, Cell Biology & Physiology, 5Biochemistry & Molecular Biology, the 6Center for Diabetes & Metabolic Diseases, and the 7Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202; 2Department of BioHealth Informatics, Indiana University-Purdue University Indianapolis, Indianapolis, IN, 46202; 8Roudebush VA Medical Center, Indianapolis, IN 46202. *Corresponding Author(s): Carmella Evans-Molina, MD, PhD ([email protected]) Indiana University School of Medicine, 635 Barnhill Drive, MS 2031A, Indianapolis, IN 46202, Telephone: (317) 274-4145, Fax (317) 274-4107 Running Title: Golgi Stress Response in Diabetes Word Count: 4358 Number of Figures: 6 Keywords: Golgi apparatus stress, Islets, β cell, Type 1 diabetes, Type 2 diabetes 1 Diabetes Publish Ahead of Print, published online August 20, 2020 Diabetes Page 2 of 781 ABSTRACT The Golgi apparatus (GA) is an important site of insulin processing and granule maturation, but whether GA organelle dysfunction and GA stress are present in the diabetic β-cell has not been tested. We utilized an informatics-based approach to develop a transcriptional signature of β-cell GA stress using existing RNA sequencing and microarray datasets generated using human islets from donors with diabetes and islets where type 1(T1D) and type 2 diabetes (T2D) had been modeled ex vivo. To narrow our results to GA-specific genes, we applied a filter set of 1,030 genes accepted as GA associated.
    [Show full text]
  • SSH3 Antibody Cat
    SSH3 Antibody Cat. No.: 55-994 SSH3 Antibody SSH3 Antibody immunohistochemistry analysis in formalin Flow cytometric analysis of Hela cells (right histogram) fixed and paraffin embedded human breast carcinoma compared to a negative control cell (left histogram).FITC- followed by peroxidase conjugation of the secondary conjugated goat-anti-rabbit secondary antibodies were antibody and DAB staining. used for the analysis. Confocal immunofluorescent analysis of SSH3 Antibody with Hela cell followed by Alexa Fluor 488-conjugated goat anti-rabbit lgG (green). Actin filaments have been labeled with Alexa Fluor555 phalloidin (red). DAPI was used to stain the cell nuclear (blue). Specifications October 1, 2021 1 https://www.prosci-inc.com/ssh3-antibody-55-994.html HOST SPECIES: Rabbit SPECIES REACTIVITY: Human This SSH3 antibody is generated from rabbits immunized with a KLH conjugated synthetic IMMUNOGEN: peptide between 575-602 amino acids from the C-terminal region of human SSH3. TESTED APPLICATIONS: Flow, IF, IHC-P, WB For WB starting dilution is: 1:1000 For IHC-P starting dilution is: 1:50~100 APPLICATIONS: For FACS starting dilution is: 1:10~50 For IF starting dilution is: 1:10~50 PREDICTED MOLECULAR 73 kDa WEIGHT: Properties This antibody is purified through a protein A column, followed by peptide affinity PURIFICATION: purification. CLONALITY: Polyclonal ISOTYPE: Rabbit Ig CONJUGATE: Unconjugated PHYSICAL STATE: Liquid BUFFER: Supplied in PBS with 0.09% (W/V) sodium azide. CONCENTRATION: batch dependent Store at 4˚C for three months and -20˚C, stable for up to one year. As with all antibodies STORAGE CONDITIONS: care should be taken to avoid repeated freeze thaw cycles.
    [Show full text]
  • Pharmacological Targeting of the Mitochondrial Phosphatase PTPMT1 by Dahlia Doughty Shenton Department of Biochemistry Duke
    Pharmacological Targeting of the Mitochondrial Phosphatase PTPMT1 by Dahlia Doughty Shenton Department of Biochemistry Duke University Date: May 1 st 2009 Approved: ___________________________ Dr. Patrick J. Casey, Supervisor ___________________________ Dr. Perry J. Blackshear ___________________________ Dr. Anthony R. Means ___________________________ Dr. Christopher B. Newgard ___________________________ Dr. John D. York Dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Department of Biochemistry in the Graduate School of Duke University 2009 ABSTRACT Pharmacological Targeting of the Mitochondrial Phosphatase PTPMT1 by Dahlia Doughty Shenton Department of Biochemistry Duke University Date: May 1 st 2009 Approved: ___________________________ Dr. Patrick J. Casey, Supervisor ___________________________ Dr. Perry J. Blackshear ___________________________ Dr. Anthony R. Means ___________________________ Dr. Christopher B. Newgard ___________________________ Dr. John D. York An abstract of a dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Department of Biochemistry in the Graduate School of Duke University 2009 Copyright by Dahlia Doughty Shenton 2009 Abstract The dual specificity protein tyrosine phosphatases comprise the largest and most diverse group of protein tyrosine phosphatases and play integral roles in the regulation of cell signaling events. The dual specificity protein tyrosine phosphatases impact multiple
    [Show full text]
  • The Role of GRHL2 and Epigenetic Remodeling in Epithelial–Mesenchymal Plasticity in Ovarian Cancer Cells
    ARTICLE https://doi.org/10.1038/s42003-019-0506-3 OPEN The role of GRHL2 and epigenetic remodeling in epithelial–mesenchymal plasticity in ovarian cancer cells Vin Yee Chung 1, Tuan Zea Tan 1, Jieru Ye1, Rui-Lan Huang2, Hung-Cheng Lai2, Dennis Kappei 1,3, 1234567890():,; Heike Wollmann4, Ernesto Guccione4 & Ruby Yun-Ju Huang1,5 Cancer cells exhibit phenotypic plasticity during epithelial–mesenchymal transition (EMT) and mesenchymal–epithelial transition (MET) involving intermediate states. To study genome-wide epigenetic remodeling associated with EMT plasticity, we integrate the ana- lyses of DNA methylation, ChIP-sequencing of five histone marks (H3K4me1, H3K4me3, H3K27Ac, H3K27me3 and H3K9me3) and transcriptome profiling performed on ovarian cancer cells with different epithelial/mesenchymal states and on a knockdown model of EMT suppressor Grainyhead-like 2 (GRHL2). We have identified differentially methylated CpG sites associated with EMT, found at promoters of epithelial genes and GRHL2 binding sites. GRHL2 knockdown results in CpG methylation gain and nucleosomal remodeling (reduction in permissive marks H3K4me3 and H3K27ac; elevated repressive mark H3K27me3), resembling the changes observed across progressive EMT states. Epigenetic-modifying agents such as 5-azacitidine, GSK126 and mocetinostat further reveal cell state-dependent plasticity upon GRHL2 overexpression. Overall, we demonstrate that epithelial genes are subject to epigenetic control during intermediate phases of EMT/MET involving GRHL2. 1 Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore. 2 Department of Obstetrics and Gynecology, Shuang Ho Hospital, Taipei Medical University, 11031 Taipei, Taiwan. 3 Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117596, Singapore.
    [Show full text]
  • Loss of PTEN Induces Microtentacles Through PI3K-Independent Activation of Cofilin
    Oncogene (2013) 32, 2200–2210 & 2013 Macmillan Publishers Limited All rights reserved 0950-9232/13 www.nature.com/onc ORIGINAL ARTICLE Loss of PTEN induces microtentacles through PI3K-independent activation of cofilin MI Vitolo1, AE Boggs1,2, RA Whipple1, JR Yoon1,2, K Thompson1, MA Matrone3, EH Cho4, EM Balzer5 and SS Martin1,2,6 Loss of PTEN tumor suppressor enhances metastatic risk in breast cancer, although the underlying mechanisms are poorly defined. We report that homozygous deletion of PTEN in mammary epithelial cells induces tubulin-based microtentacles (McTNs) that facilitate cell reattachment and homotypic aggregation. Treatment with contractility-modulating drugs showed that McTNs in PTEN À / À cells are suppressible by controlling the actin cytoskeleton. Because outward microtubule extension is counteracted by actin cortical contraction, increased activity of actin-severing proteins could release constraints on McTN formation in PTEN À / À cells. One such actin-severing protein, cofilin, is activated in detached PTEN À / À cells that could weaken the actin cortex to promote McTNs. Expression of wild-type cofilin, an activated mutant (S3A), and an inactive mutant (S3E) demonstrated that altering cofilin phosphorylation directly affects McTNs formation. Chemical inhibition of PI3K did not reduce McTNs or inactivate cofilin in PTEN À / À cells. Additionally, knock-in expression of the two most common PI3K-activating mutations observed in human cancer patients did not increase McTNs or activate cofilin. PTEN loss and PI3K activation also caused differential activation of the cofilin regulators, LIM-kinase1 (LIMK) and Slingshot-1L (SSH). Furthermore, McTNs were suppressed and cofilin was inactivated by restoration of PTEN in the PTEN À / À cells, indicating that both the elevation of McTNs and the activation of cofilin are specific results arising from PTEN loss.
    [Show full text]
  • Phosphatases Page 1
    Phosphatases esiRNA ID Gene Name Gene Description Ensembl ID HU-05948-1 ACP1 acid phosphatase 1, soluble ENSG00000143727 HU-01870-1 ACP2 acid phosphatase 2, lysosomal ENSG00000134575 HU-05292-1 ACP5 acid phosphatase 5, tartrate resistant ENSG00000102575 HU-02655-1 ACP6 acid phosphatase 6, lysophosphatidic ENSG00000162836 HU-13465-1 ACPL2 acid phosphatase-like 2 ENSG00000155893 HU-06716-1 ACPP acid phosphatase, prostate ENSG00000014257 HU-15218-1 ACPT acid phosphatase, testicular ENSG00000142513 HU-09496-1 ACYP1 acylphosphatase 1, erythrocyte (common) type ENSG00000119640 HU-04746-1 ALPL alkaline phosphatase, liver ENSG00000162551 HU-14729-1 ALPP alkaline phosphatase, placental ENSG00000163283 HU-14729-1 ALPP alkaline phosphatase, placental ENSG00000163283 HU-14729-1 ALPPL2 alkaline phosphatase, placental-like 2 ENSG00000163286 HU-07767-1 BPGM 2,3-bisphosphoglycerate mutase ENSG00000172331 HU-06476-1 BPNT1 3'(2'), 5'-bisphosphate nucleotidase 1 ENSG00000162813 HU-09086-1 CANT1 calcium activated nucleotidase 1 ENSG00000171302 HU-03115-1 CCDC155 coiled-coil domain containing 155 ENSG00000161609 HU-09022-1 CDC14A CDC14 cell division cycle 14 homolog A (S. cerevisiae) ENSG00000079335 HU-11533-1 CDC14B CDC14 cell division cycle 14 homolog B (S. cerevisiae) ENSG00000081377 HU-06323-1 CDC25A cell division cycle 25 homolog A (S. pombe) ENSG00000164045 HU-07288-1 CDC25B cell division cycle 25 homolog B (S. pombe) ENSG00000101224 HU-06033-1 CDKN3 cyclin-dependent kinase inhibitor 3 ENSG00000100526 HU-02274-1 CTDSP1 CTD (carboxy-terminal domain,
    [Show full text]
  • Dual Specificity Phosphatases from Molecular Mechanisms to Biological Function
    International Journal of Molecular Sciences Dual Specificity Phosphatases From Molecular Mechanisms to Biological Function Edited by Rafael Pulido and Roland Lang Printed Edition of the Special Issue Published in International Journal of Molecular Sciences www.mdpi.com/journal/ijms Dual Specificity Phosphatases Dual Specificity Phosphatases From Molecular Mechanisms to Biological Function Special Issue Editors Rafael Pulido Roland Lang MDPI • Basel • Beijing • Wuhan • Barcelona • Belgrade Special Issue Editors Rafael Pulido Roland Lang Biocruces Health Research Institute University Hospital Erlangen Spain Germany Editorial Office MDPI St. Alban-Anlage 66 4052 Basel, Switzerland This is a reprint of articles from the Special Issue published online in the open access journal International Journal of Molecular Sciences (ISSN 1422-0067) from 2018 to 2019 (available at: https: //www.mdpi.com/journal/ijms/special issues/DUSPs). For citation purposes, cite each article independently as indicated on the article page online and as indicated below: LastName, A.A.; LastName, B.B.; LastName, C.C. Article Title. Journal Name Year, Article Number, Page Range. ISBN 978-3-03921-688-8 (Pbk) ISBN 978-3-03921-689-5 (PDF) c 2019 by the authors. Articles in this book are Open Access and distributed under the Creative Commons Attribution (CC BY) license, which allows users to download, copy and build upon published articles, as long as the author and publisher are properly credited, which ensures maximum dissemination and a wider impact of our publications. The book as a whole is distributed by MDPI under the terms and conditions of the Creative Commons license CC BY-NC-ND. Contents About the Special Issue Editors ....................................
    [Show full text]
  • Dynamics of Dual Specificity Phosphatases and Their Interplay with Protein Kinases in Immune Signaling Yashwanth Subbannayya1,2, Sneha M
    bioRxiv preprint doi: https://doi.org/10.1101/568576; this version posted March 5, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Dynamics of dual specificity phosphatases and their interplay with protein kinases in immune signaling Yashwanth Subbannayya1,2, Sneha M. Pinto1,2, Korbinian Bösl1, T. S. Keshava Prasad2 and Richard K. Kandasamy1,3,* 1Centre of Molecular Inflammation Research (CEMIR), and Department of Clinical and Molecular Medicine (IKOM), Norwegian University of Science and Technology, N-7491 Trondheim, Norway 2Center for Systems Biology and Molecular Medicine, Yenepoya (Deemed to be University), Mangalore 575018, India 3Centre for Molecular Medicine Norway (NCMM), Nordic EMBL Partnership, University of Oslo and Oslo University Hospital, N-0349 Oslo, Norway *Correspondence: Richard K. Kandasamy ([email protected]) Abstract Dual specificity phosphatases (DUSPs) have a well-known role as regulators of the immune response through the modulation of mitogen activated protein kinases (MAPKs). Yet the precise interplay between the various members of the DUSP family with protein kinases is not well understood. Recent multi-omics studies characterizing the transcriptomes and proteomes of immune cells have provided snapshots of molecular mechanisms underlying innate immune response in unprecedented detail. In this study, we focused on deciphering the interplay between members of the DUSP family with protein kinases in immune cells using publicly available omics datasets. Our analysis resulted in the identification of potential DUSP- mediated hub proteins including MAPK7, MAPK8, AURKA, and IGF1R. Furthermore, we analyzed the association of DUSP expression with TLR4 signaling and identified VEGF, FGFR and SCF-KIT pathway modules to be regulated by the activation of TLR4 signaling.
    [Show full text]
  • Inhibition of Dual-Specificity Phosphatase 26 by Ethyl-3,4-Dephostatin: Ethyl-3,4-Dephostatin As a Multiphosphatase Inhibitor
    ORIGINAL ARTICLES College of Pharmacy, Chung-Ang University, Seoul, Republic of Korea Inhibition of dual-specificity phosphatase 26 by ethyl-3,4-dephostatin: Ethyl-3,4-dephostatin as a multiphosphatase inhibitor HUIYUN SEO, SAYEON CHO Received September 30, 2015, accepted November 6, 2015 Sayeon Cho, Ph.D., College of Pharmacy, Chung-Ang University, Seoul 156-756, Republic of Korea [email protected] Pharmazie 71: 196–200 (2016) doi: 10.1691/ph.2016.5803 Protein tyrosine phosphatases (PTPs) regulate protein function by dephosphorylating phosphorylated proteins in many signaling cascades and some of them have been targets for drug development against many human diseases. There have been many reports that some chemical inhibitors could regulate particular phosphatases. However, there was no extensive study on specificity of inhibitors towardss phosphatases. We investigated the effects of ethyl-3,4-dephostatin, a potent inhibitor of five PTPs including PTP-1B and Src homology-2-containing protein tyrosine phosphatase-1 (SHP-1), on thirteen other PTPs using in vitro phosphatase assays. Of them, dual-specificity protein phosphatase 26 (DUSP26), which inhibits mitogen-activated protein kinase (MAPK) and p53 tumor suppressor and is known to be overexpressed in anaplastic thyroid carcinoma, was inhibited by ethyl- 3,4-dephostatin in a concentration-dependent manner. Kinetic studies with ethyl-3,4-dephostatin and DUSP26 revealed competitive inhibition, suggesting that ethyl-3,4-dephostatin binds to the catalytic site of DUSP26 like other substrate PTPs. Moreover, ethyl-3,4-dephostatin protects DUSP26-mediated dephosphorylation of p38, a member of the MAPK family, and p53. Taken together, these results suggest that ethyl-3,4-dephostatin functions as a multiphosphatase inhibitor and is useful as a therapeutic agent for cancers overexpressing DUSP26.
    [Show full text]
  • The Human Phosphatase Interactome
    FEBS Letters 586 (2012) 2732–2739 journal homepage: www.FEBSLetters.org Review The human phosphatase interactome: An intricate family portrait ⇑ Francesca Sacco a,1, Livia Perfetto a,1, Luisa Castagnoli a, Gianni Cesareni a,b, a Department of Biology, University of Rome ‘‘Tor Vergata’’, Rome, Italy b Research Institute ‘‘Fondazione Santa Lucia’’, Rome, Italy article info abstract Article history: The concerted activities of kinases and phosphatases modulate the phosphorylation levels of Received 23 March 2012 proteins, lipids and carbohydrates in eukaryotic cells. Despite considerable effort, we are still miss- Revised 8 May 2012 ing a holistic picture representing, at a proteome level, the functional relationships between Accepted 8 May 2012 kinases, phosphatases and their substrates. Here we focus on phosphatases and we review and inte- Available online 21 May 2012 grate the available information that helps to place the members of the protein phosphatase super- Edited by Marius Sudol, Giulio Superti-Furga families into the human protein interaction network. In addition we show how protein interaction and Wilhelm Just domains and motifs, either covalently linked to the phosphatase domain or in regulatory/adaptor subunits, play a prominent role in substrate selection. Keywords: Ó 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved. Human phosphatome Phosphatase family classification Substrate recognition specificity 1. Introduction protein kinases. 428 are known or predicted to phosphorylate ser- ine and threonine residues, while the remaining 90 are members of Phosphorylation is a widespread post-translational modifica- the tyrosine kinase family [3,12]. By contrast, in the human gen- tion governing signal propagation [1].
    [Show full text]
  • PDF Download
    SSH3 Polyclonal Antibody Catalog No : YT4428 Reactivity : Human,Mouse,Rat Applications : WB,IHC-p,IF(paraffin section),ELISA Gene Name : SSH3 Protein Name : Protein phosphatase Slingshot homolog 3 Human Gene Id : 54961 Human Swiss Prot Q8TE77 No : Mouse Gene Id : 245857 Mouse Swiss Prot Q8K330 No : Rat Gene Id : 365396 Rat Swiss Prot No : Q5XIS1 Immunogen : The antiserum was produced against synthesized peptide derived from human SSH3. AA range:357-406 Specificity : SSH3 Polyclonal Antibody detects endogenous levels of SSH3 protein. Formulation : Liquid in PBS containing 50% glycerol, 0.5% BSA and 0.02% sodium azide. Source : Rabbit Dilution : Western Blot: 1/500 - 1/2000. Immunohistochemistry: 1/100 - 1/300. ELISA: 1/40000. Not yet tested in other applications. Purification : The antibody was affinity-purified from rabbit antiserum by affinity- chromatography using epitope-specific immunogen. Concentration : 1 mg/ml 1 / 3 Storage Stability : -20°C/1 year Molecularweight : 72996 Observed Band : 74 Cell Pathway : Regulates Actin and Cytoskeleton, Background : slingshot protein phosphatase 3(SSH3) Homo sapiens The ADF (actin- depolymerizing factor)/cofilin family (see MIM 601442) is composed of stimulus- responsive mediators of actin dynamics. ADF/cofilin proteins are inactivated by kinases such as LIM domain kinase-1 (LIMK1; MIM 601329). The SSH family appears to play a role in actin dynamics by reactivating ADF/cofilin proteins in vivo (Niwa et al., 2002 [PubMed 11832213]).[supplied by OMIM, Mar 2008], Function : catalytic activity:A phosphoprotein + H(2)O = a protein + phosphate.,catalytic activity:Protein tyrosine phosphate + H(2)O = protein tyrosine + phosphate.,function:Protein phosphatase which may play a role in the regulation of actin filament dynamics.
    [Show full text]
  • Epigenome-Wide Association Study on Asthma and Chronic Obstructive
    www.nature.com/scientificreports OPEN Epigenome‑wide association study on asthma and chronic obstructive pulmonary disease overlap reveals aberrant DNA methylations related to clinical phenotypes Yung‑Che Chen1,2*, Ying‑Huang Tsai1, Chin‑Chou Wang1,6, Shih‑Feng Liu1,7, Ting‑Wen Chen3,4,5, Wen‑Feng Fang1,6, Chiu‑Ping Lee1, Po‑Yuan Hsu1, Tung‑Ying Chao1, Chao‑Chien Wu1, Yu‑Feng Wei8, Huang‑Chih Chang1, Chia‑Cheng Tsen1, Yu‑Ping Chang1, Meng‑Chih Lin1,2* & Taiwan Clinical Trial Consortium of Respiratory Disease (TCORE) group* We hypothesized that epigenetics is a link between smoking/allergen exposures and the development of Asthma and chronic obstructive pulmonary disease (ACO). A total of 75 of 228 COPD patients were identifed as ACO, which was independently associated with increased exacerbations. Microarray analysis identifed 404 diferentially methylated loci (DML) in ACO patients, and 6575 DML in those with rapid lung function decline in a discovery cohort. In the validation cohort, ACO patients had hypermethylated PDE9A (+ 30,088)/ZNF323 (− 296), and hypomethylated SEPT8 (− 47) genes as compared with either pure COPD patients or healthy non‑smokers. Hypermethylated TIGIT (− 173) gene and hypomethylated CYSLTR1 (+ 348)/CCDC88C (+ 125,722)/ADORA2B (+ 1339) were associated with severe airfow limitation, while hypomethylated IFRD1 (− 515) gene with frequent exacerbation in all the COPD patients. Hypermethylated ZNF323 (− 296) / MPV17L (+ 194) and hypomethylated PTPRN2 (+ 10,000) genes were associated with rapid lung function decline. In vitro cigarette smoke extract and ovalbumin concurrent exposure resulted in specifc DNA methylation changes of the MPV17L / ZNF323 genes, while 5‑aza‑2′‑deoxycytidine treatment reversed promoter hypermethylation‑mediated MPV17L under‑expression accompanied with reduced apoptosis and decreased generation of reactive oxygen species.
    [Show full text]