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Retinoic Acid-Induced Protein 14 (RAI14) Promotes mTOR-Mediated Inflammation Under Inflammatory Stress and Chemical Hypoxia in a U87 Glioblastoma Cell Line

  • XiaoGang Shen
  • JiaRui Zhang
  • XiaoLong Zhang
  • YiFan Wang
  • YunFeng Hu
  • Jun GuoEmail author
Original Research

Abstract

Retinoic acid-induced 14 is a developmentally regulated gene induced by retinoic acid and is closely associated with NIK/NF-κB signaling. In the present study, we examined the effect of RAI14 on mTOR-mediated glial inflammation in response to inflammatory factors and chemical ischemia. A U87 cell model of LPS- and TNF-α-induced inflammation was used to investigate the role of RAI14 in glial inflammation. U87 cells were treated with siR-RAI14 or everolimus to detect the correlation between mTOR, RAI14, and NF-κB. CoCl2-stimulated U87 cells were used to analyze the effect of RAI14 on mTOR-mediated NF-κB inflammatory signaling under chemical hypoxia. LPS and TNF-α stimulation resulted in the upregulation of RAI14 mRNA and protein levels in a dose- and time-dependent manner. RAI14 knockdown significantly attenuated the level of pro-inflammatory cytokine via inhibiting the IKK/NF-κB pathway. Treatment with an mTOR inhibitor (everolimus) ameliorated NF-κB activity and IKKα/β phosphorylation via RAI14 signaling. Notably, RAI14 also enhanced mTOR-mediated NF-κB activation under conditions of chemical hypoxia. These findings provide significant insight into the role of RAI14 in mTOR-induced glial inflammation, which is closely associated with infection and ischemia stimuli. Thus, RAI14 may be a potential drug target for the treatment of inflammatory diseases.

Keywords

RAI14 Neuroinflmamation mTOR NF-κB Chemical hypoxia 

Abbreviations

4EBP1

Eukaryotic initiation factor 4E binding protein1

CNS

Central nervous system

CoCl2

Cobalt chloride

Co-IP

Co-immunoprecipitation

eIF4E

Eukaryotic translational initiation factor 4 epsilon

Evero

Everolimus

IκB-α

NF-kappa-B inhibitor alpha

IKK

Nuclear factor NF-kappa-B inhibitor kinase

LPS

Lipopolysaccharide

mRNA

Messenger RNA

mTOR

Mammalian target of rapamycin

mTORC

Mammalian target of rapamycin complex

NF-κB

Nuclear factor kappa-light-chain-enhancer of activated B cells

NIK

NF-kappa-B-inducing kinase

PAMPs

Pathogen-associated molecular patterns

PBS

Phosphate-buffered saline

RAI14

Retinoic acid-induced protein 14

Raptor

Regulatory-associated protein of mTOR

RT-qPCR

Quantitative real-time polymerase chain reaction

si-NC

Control siRNA.

siR-RAI14

RAI14 siRNA

siRNA

Small interfering RNA

TLRs

Toll-like receptors

TNF-α

Tumor-necrosis factor-α

Notes

Author Contributions

In this study, JG conceived the general idea. XGS, JRZ, XLZ, YFW, and YFH carried out experiments. XGS, and JG analyzed and interpreted the data and wrote the paper. JG critically reviewed and edited the work. All authors approved the final version of the manuscript.

Funding

This work was supported by Grants from the National Natural Science Foundation of China (81573409) and Natural Science Foundation of Jiangsu Province (BK20161574) and A Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions (Integration of Raditional Chinese and Western Medicine).

Compliance with Ethical Standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical Approval

This article does not contain any studies with human participants or animals performed by any of the authors. The manuscript does not contain clinical studies or patient data.

Supplementary material

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Supplementary material 1 (PDF 186 KB)
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References

  1. Caccamo A, Belfiore R, Oddo S (2018) Genetically reducing mTOR signaling rescues central insulin dysregulation in a mouse model of Alzheimer’s disease. Neurobiol Aging 68:59–67.  https://doi.org/10.1016/j.neurobiolaging.2018.03.032 CrossRefPubMedGoogle Scholar
  2. Chen T, Guo Y, Shan J, Zhang J, Shen X, Guo J, Liu XM (2018a) Vector analysis of cytoskeletal structural tension and the mechanisms that underpin spectrin-related forces in pyroptosis. Antioxid Redox Sign.  https://doi.org/10.1089/ars.2017.7366 CrossRefGoogle Scholar
  3. Chen WR, Liu HB, Chen YD, Sha Y, Ma Q, Zhu PJ, Mu Y (2018b) Melatonin attenuates myocardial ischemia/reperfusion injury by inhibiting autophagy via an AMPK/mTOR signaling pathway. Cell Physiol Biochem 47:2067–2076.  https://doi.org/10.1159/000491474 CrossRefPubMedGoogle Scholar
  4. Choi YJ et al (2012) Inhibitory effect of mTOR activator MHY1485 on autophagy: suppression of lysosomal fusion. PLoS ONE 7:e43418.  https://doi.org/10.1371/journal.pone.0043418 CrossRefPubMedPubMedCentralGoogle Scholar
  5. Coffey RT, Shi Y, Long MJC, Marr MTN, Hedstrom L (2016) Ubiquilin-mediated small molecule inhibition of mammalian target of rapamycin complex 1 (mTORC1) signaling. J Biol Chem 291:5221–5233.  https://doi.org/10.1074/jbc.M115.691584 CrossRefPubMedPubMedCentralGoogle Scholar
  6. Collignon E et al (2018) Immunity drives TET1 regulation in cancer through NF-kappaB. Sci Adv 4:7309.  https://doi.org/10.1126/sciadv.aap7309 CrossRefGoogle Scholar
  7. Cosin-Roger J et al (2017) Hypoxia ameliorates intestinal inflammation through NLRP3/mTOR downregulation and autophagy activation. Nat Commun 8:98.  https://doi.org/10.1038/s41467-017-00213-3 CrossRefPubMedPubMedCentralGoogle Scholar
  8. Dan HC, Cooper MJ, Cogswell PC, Duncan JA, Ting JP, Baldwin AS (2008) Akt-dependent regulation of NF-{kappa}B is controlled by mTOR and Raptor in association with IKK. Gene Dev 22:1490–1500.  https://doi.org/10.1101/gad.1662308 CrossRefPubMedGoogle Scholar
  9. Gu L, Huang B, Shen W, Gao L, Ding Z, Wu H, Guo J (2013) Early activation of nSMase2/ceramide pathway in astrocytes is involved in ischemia-associated neuronal damage via inflammation in rat hippocampi. J Neuroinflamm 10:109.  https://doi.org/10.1186/1742-2094-10-109 CrossRefGoogle Scholar
  10. Guo F et al (2013) mTOR regulates DNA damage response through NF-kappaB-mediated FANCD2 pathway in hematopoietic cells. Leukemia 27:2040–2046.  https://doi.org/10.1038/leu.2013.93 CrossRefPubMedPubMedCentralGoogle Scholar
  11. Guo YC, Wang YX, Ge YP, Yu LJ, Guo J (2018) Analysis of subcellular structural tension in axonal growth of neurons. Rev Neurosci 29:125–137.  https://doi.org/10.1515/revneuro-2017-0047 CrossRefPubMedGoogle Scholar
  12. Han R, Gao J, Zhai H, Xiao J, Ding Y, Hao J (2016) RAD001 (everolimus) attenuates experimental autoimmune neuritis by inhibiting the mTOR pathway, elevating Akt activity and polarizing M2 macrophages. Exp Neurol 280:106–114.  https://doi.org/10.1016/j.expneurol.2016.04.005 CrossRefPubMedGoogle Scholar
  13. Hara K et al (1997) Regulation of eIF-4E BP1 phosphorylation by mTOR. J Biol Chem 272:26457–26463CrossRefGoogle Scholar
  14. Hasson SA et al (2013) High-content genome-wide RNAi screens identify regulators of parkin upstream of mitophagy. Nature 504:291–295.  https://doi.org/10.1038/nature12748 CrossRefPubMedPubMedCentralGoogle Scholar
  15. Hay N, Sonenberg N (2004) Upstream and downstream of mTOR. Gene Dev 18:1926–1945CrossRefGoogle Scholar
  16. Hsu Y et al (2013) Genome-wide analysis of three-way interplay among gene expression, cancer cell invasion and anti-cancer compound sensitivity. BMC Med 11:106.  https://doi.org/10.1186/1741-7015-11-106 CrossRefPubMedPubMedCentralGoogle Scholar
  17. Huttlin EL et al (2017) Architecture of the human interactome defines protein communities and disease networks. Nature 545:505–509.  https://doi.org/10.1038/nature22366 CrossRefPubMedPubMedCentralGoogle Scholar
  18. Kielian T (2006) Toll-like receptors in central nervous system glial inflammation and homeostasis. J Neurosci Res 83:711–730CrossRefGoogle Scholar
  19. Kingwell K (2013) Neuro-oncology: everolimus for astrocytoma in tuberous sclerosis complex. Nat Rev Neurol 9:6.  https://doi.org/10.1038/nrneurol.2012.257 CrossRefPubMedGoogle Scholar
  20. Kutty RK et al (2001) Molecular characterization and developmental expression of NORPEG, a novel gene induced by retinoic acid. J Biol Chem 276:2831–2840CrossRefGoogle Scholar
  21. Kutty RK et al (2006) Cell density-dependent nuclear/cytoplasmic localization of NORPEG (RAI14) protein. Biochem Bioph Res Co 345:1333–1341CrossRefGoogle Scholar
  22. Lee D et al (2007) IKK beta suppression of TSC1 links inflammation and tumor angiogenesis via the mTOR pathway. Cell 130:440–455CrossRefGoogle Scholar
  23. Li S et al (2008) Genome-wide coactivation analysis of PGC-1alpha identifies BAF60a as a regulator of hepatic lipid metabolism. Cell Metab 8:105–117.  https://doi.org/10.1016/j.cmet.2008.06.013 CrossRefPubMedPubMedCentralGoogle Scholar
  24. Li J, Tang Y, Cai D (2012) IKKbeta/NF-kappaB disrupts adult hypothalamic neural stem cells to mediate a neurodegenerative mechanism of dietary obesity and pre-diabetes. Nat Cell Biol 14:999–1012.  https://doi.org/10.1038/ncb2562 CrossRefPubMedPubMedCentralGoogle Scholar
  25. Li J et al (2016) Nuclear PKC-theta facilitates rapid transcriptional responses in human memory CD4 + T cells through p65 and H2B phosphorylation. J Cell Sci 129:2448–2461.  https://doi.org/10.1242/jcs.181248 CrossRefPubMedPubMedCentralGoogle Scholar
  26. Liu Y et al (2017) Peripheral immune tolerance alleviates the intracranial lipopolysaccharide injection-induced neuroinflammation and protects the dopaminergic neurons from neuroinflammation-related neurotoxicity. J Neuroinflamm 14:223.  https://doi.org/10.1186/s12974-017-0994-3 CrossRefGoogle Scholar
  27. Peng YF et al (2000) Ankycorbin: a novel actin cytoskeleton-associated protein. Genes Cells 5:1001–1008CrossRefGoogle Scholar
  28. Qian X, Mruk DD, Cheng CY (2013) Rai14 (retinoic acid induced protein 14) is involved in regulating f-actin dynamics at the ectoplasmic specialization in the rat testis*. PLoS ONE 8:e60656.  https://doi.org/10.1371/journal.pone.0060656 CrossRefPubMedPubMedCentralGoogle Scholar
  29. Subhan F et al (2017) Fish scale collagen peptides protect against CoCl2/TNF-alpha-induced cytotoxicity and inflammation via inhibition of ROS, MAPK, and NF-kappaB pathways in HaCaT cells. Oxid Med Cell Longev 2017:9703609.  https://doi.org/10.1155/2017/9703609 CrossRefPubMedPubMedCentralGoogle Scholar
  30. Temiz-Resitoglu M et al (2017) Activation of mTOR/IκB-α/NF-κB pathway contributes to LPS-induced hypotension and inflammation in rats. Eur J Pharmacol 802:7–19.  https://doi.org/10.1016/j.ejphar.2017.02.034 CrossRefPubMedGoogle Scholar
  31. Thoreen CC, Chantranupong L, Keys HR, Wang T, Gray NS, Sabatini DM (2012) A unifying model for mTORC1-mediated regulation of mRNA translation. Nature 485:109–113.  https://doi.org/10.1038/nature11083 CrossRefPubMedPubMedCentralGoogle Scholar
  32. Vitiello D, Neagoe P, Sirois MG, White M (2015) Effect of everolimus on the immunomodulation of the human neutrophil inflammatory response and activation. Cell Mol Immunol 12:40–52.  https://doi.org/10.1038/cmi.2014.24 CrossRefPubMedGoogle Scholar
  33. Wendel H et al (2004) Survival signalling by Akt and eIF4E in oncogenesis and cancer therapy. Nature 428:332–337CrossRefGoogle Scholar
  34. Yang T, Li D, Liu F, Qi L, Yan G, Wang M (2015) Regulation on Beclin-1 expression by mTOR in CoCl2-induced HT22 cell ischemia-reperfusion injury. Brain Res 1614:60–66.  https://doi.org/10.1016/j.brainres.2015.04.016 CrossRefPubMedGoogle Scholar
  35. Yun S et al (2016) 4EBP1/c-MYC/PUMA and NF-kappaB/EGR1/BIM pathways underlie cytotoxicity of mTOR dual inhibitors in malignant lymphoid cells. Blood 127:2711–2722.  https://doi.org/10.1182/blood-2015-02-629485 CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • XiaoGang Shen
    • 1
    • 2
  • JiaRui Zhang
    • 1
    • 2
  • XiaoLong Zhang
    • 1
    • 2
  • YiFan Wang
    • 1
    • 2
  • YunFeng Hu
    • 1
    • 2
  • Jun Guo
    • 1
    • 2
    • 3
    Email author
  1. 1.State Key Laboratory Cultivation Base For TCM Quality and Efficacy, School of Medicine and Life ScienceNanjing University of Chinese MedicineNanjingPeople’s Republic of China
  2. 2.Key Laboratory of Drug Target and Drug for Degenerative DiseaseNanjing University of Chinese MedicineNanjingPeople’s Republic of China
  3. 3.Department of Biochemistry and Molecular Biology, Jiangsu Key Laboratory of Therapeutic Material of Chinese Medicine, School of Medicine and Life ScienceNanjing University of Chinese MedicineNanjingPeople’s Republic of China

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