Advertisement

Neurochemical Research

, Volume 43, Issue 8, pp 1539–1548 | Cite as

Ulk1/FUNDC1 Prevents Nerve Cells from Hypoxia-Induced Apoptosis by Promoting Cell Autophagy

  • Li Wang
  • Peng Wang
  • He Dong
  • Shiduan Wang
  • Haichen Chu
  • Wei Yan
  • Xue Zhang
Original Paper
  • 83 Downloads

Abstract

Cell autophagy and cell apoptosis are both observed in the process of hypoxia-induced ischemic cerebral infarction (ICI). Unc-51 like autophagy activating kinase 1 (Ulk1) and FUN14 Domain-containing Protein 1 (FUNDC1) are both involved in the regulation of cell autophagy. This study aimed to investigate the regulatory effects of Ulk1 and FUNDC1 on hypoxia-induced nerve cell autophagy and apoptosis. Cell viability was measured using cell counting kit-8 (CCK-8) assay. Cell apoptosis was detected using Annexin V-PE/7-ADD staining assay. qRT-PCR was used to quantify the mRNA levels of Ulk1 and FUNDC1 in PC-12 cells. Cell transfection was performed to up-regulate the expression of Ulk1. 3-Methyladenine (3-MA) was used as autophagy inhibitor and rapamycin was used as autophagy activator in our experiments. SP600125 was used as c-Jun N-terminal kinase (JNK) inhibitor. Western blotting was performed to analyze the expression levels of key factors that are related to cell autophagy, apoptosis and JNK pathway. We found that hypoxia simultaneously induced apoptosis and autophagy of PC-12 cells. The activation of Ulk1 and FUNDC1 were also found in PC-12 cells after hypoxia induction. Overexpression of Ulk1 promoted the activation of FUNDC1 and prevented PC-12 cells from hypoxia-induced apoptosis. Suppression of Ulk1 had opposite effects. Furthermore, we also found that JNK pathway participated in the effects of Ulk1 overexpression on PC-12 cell apoptosis reduction. To conclude, Ulk1/FUNDC1 played critical regulatory roles in hypoxia-induced nerve cell autophagy and apoptosis. Overexpression of Ulk1 prevented nerve cells from hypoxia-induced apoptosis by promoting cell autophagy.

Keywords

Ischemic cerebral infarction Unc-51 like autophagy activating kinase 1 FUN14 Domain-containing Protein 1 Cell autophagy Cell apoptosis JNK pathway 

Notes

Funding

This work was supported by Youth Research Fund of The Affiliated Hospital of Qingdao University.

Compliance with Ethical Standards

Conflict of interest

The authors declare that they have no conflict of interest.

References

  1. 1.
    Della-Morte D, Pacifici F, Rundek T (2016) Genetic susceptibility to cerebrovascular disease. Curr Opin Lipidol 27:187–195CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Chen PS, Cheng CL, Kao Yang YH, Li YH (2016) Statin adherence after ischemic stroke or transient ischemic attack is associated with clinical outcome. Circ J 80:731–737CrossRefPubMedGoogle Scholar
  3. 3.
    Yu J, Bao C, Dong Y, Liu X (2015) Activation of autophagy in rat brain cells following focal cerebral ischemia reperfusion through enhanced expression of Atg1/pULK and LC3. Mol Med Rep 12:3339–3344CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Savion N, Izigov N, Morein M, Pri-Chen S, Kotev-Emeth S (2014) S-allylmercapto-N-acetylcysteine (ASSNAC) protects cultured nerve cells from oxidative stress and attenuates experimental autoimmune encephalomyelitis. Neurosci Lett 583:108–113CrossRefPubMedGoogle Scholar
  5. 5.
    Pozo Devoto VM, Bogetti ME, Fiszer de Plazas S (2013) Developmental and hypoxia-induced cell death share common ultrastructural and biochemical apoptotic features in the central nervous system. Neuroscience 252:190–200CrossRefPubMedGoogle Scholar
  6. 6.
    Descloux C, Ginet V, Clarke PG, Puyal J, Truttmann AC (2015) Neuronal death after perinatal cerebral hypoxia-ischemia: focus on autophagy-mediated cell death. Int J Dev Neurosci 45:75–85CrossRefPubMedGoogle Scholar
  7. 7.
    Park JM, Chang HJ, Seo M, Otto NM, Grunwald D, Kim KH, Moriarity B, Kim YM, Starker C, Nho RS (2016) The ULK1 complex mediates MTORC1 signaling to the autophagy initiation machinery via binding and phosphorylating ATG14. Autophagy 12:547–564CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Li R, Zhou P, Guo Y, Lee JS, Zhou B (2017) Tris (1, 3-dichloro-2-propyl) phosphate induces apoptosis and autophagy in SH-SY5Y cells: involvement of ROS-mediated AMPK/mTOR/ULK1 pathways. Food Chem Toxicol 100:183–196CrossRefPubMedGoogle Scholar
  9. 9.
    Yu X, Munoz-Alarcon A, Ajayi A, Webling KE, Steinhof A, Langel U, Strom AL (2013) Inhibition of autophagy via p53-mediated disruption of ULK1 in a SCA7 polyglutamine disease model. J Mol Neurosci 50:586–599CrossRefPubMedGoogle Scholar
  10. 10.
    Li Y, Zhang J, Yang C (2015) UNC-51-like kinase 1 blocks S6k1 phosphorylation contributes to neurodegeneration in Parkinson’s disease model in vitro. Biochem Biophys Res Commun 459:196–200CrossRefPubMedGoogle Scholar
  11. 11.
    Jang WH, Jeong YJ, Choi SH, Urm SH, Seog DH (2017) Interaction of FUN14 domain containing 1, a mitochondrial outer membrane protein, with kinesin light chain 1 via the tetratricopeptide repeat domain. Biomed Rep 6:46–50CrossRefPubMedGoogle Scholar
  12. 12.
    Cheng MY, Cheng YW, Yan J, Hu XQ, Zhang H, Wang ZR, Yin Q, Cheng W (2016) SIRT6 suppresses mitochondrial defects and cell death via the NF-kappaB pathway in myocardial hypoxia/reoxygenation induced injury. Am J Transl Res 8:5005–5015PubMedPubMedCentralGoogle Scholar
  13. 13.
    Wu W, Tian W, Hu Z, Chen G, Huang L, Li W, Zhang X, Xue P, Zhou C, Liu L, Zhu Y, Zhang X, Li L, Zhang L, Sui S, Zhao B, Feng D (2014) ULK1 translocates to mitochondria and phosphorylates FUNDC1 to regulate mitophagy. EMBO Rep 15:566–575CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Zhou J, Li G, Zheng Y, Shen HM, Hu X, Ming QL, Huang C, Li P, Gao N (2015) A novel autophagy/mitophagy inhibitor liensinine sensitizes breast cancer cells to chemotherapy through DNM1L-mediated mitochondrial fission. Autophagy 11:1259–1279CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Qiang L, Wu C, Ming M, Viollet B, He YY (2013) Autophagy controls p38 activation to promote cell survival under genotoxic stress. J Biol Chem 288:1603–1611CrossRefPubMedGoogle Scholar
  16. 16.
    Holmen C, Elsheikh E, Christensson M, Liu J, Johansson AS, Qureshi AR, Jalkanen S, Sumitran-Holgersson S (2007) Anti endothelial cell autoantibodies selectively activate SAPK/JNK signalling in Wegener’s granulomatosis. J Am Soc Nephrol 18:2497–2508CrossRefPubMedGoogle Scholar
  17. 17.
    Yang Z, Zhao TZ, Zou YJ, Zhang JH, Feng H (2014) Hypoxia Induces autophagic cell death through hypoxia-inducible factor 1alpha in microglia. PLoS ONE 9:e96509CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Scaiewicz V, Sorin V, Fellig Y, Birman T, Mizrahi A, Galula J, Abu-Lail R, Shneider T, Ohana P, Buscail L, Hochberg A, Czerniak A (2010) Use of H19 gene regulatory sequences in DNA-based therapy for pancreatic cancer. J Oncol 2010:178174CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Wein F, Otto T, Lambertz P, Fandrey J, Hansmann ML, Kuppers R (2015) Potential role of hypoxia in early stages of Hodgkin lymphoma pathogenesis. Haematologica 100:1320–1326CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods 25:402–408CrossRefPubMedGoogle Scholar
  21. 21.
    Yuyama K, Igarashi Y (2016) Physiological and pathological roles of exosomes in the nervous system. Biomol Concepts 7:53–68CrossRefPubMedGoogle Scholar
  22. 22.
    Yousuf S, Atif F, Ahmad M, Hoda N, Ishrat T, Khan B, Islam F (2009) Resveratrol exerts its neuroprotective effect by modulating mitochondrial dysfunctions and associated cell death during cerebral ischemia. Brain Res 1250:242–253CrossRefPubMedGoogle Scholar
  23. 23.
    Wang S, Zhou J, Kang W, Dong Z, Wang H (2016) Tocilizumab inhibits neuronal cell apoptosis and activates STAT3 in cerebral infarction rat model. Bosn J Basic Med Sci 16:145–150PubMedPubMedCentralGoogle Scholar
  24. 24.
    Poluzzi C, Casulli J, Goyal A, Mercer TJ, Neill T, Iozzo RV (2014) Endorepellin evokes autophagy in endothelial cells. J Biol Chem 289:16114–16128CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Maejima Y, Isobe M, Sadoshima J (2016) Regulation of autophagy by Beclin 1 in the heart. J Mol Cell Cardiol 95:19–25CrossRefPubMedGoogle Scholar
  26. 26.
    Tanida I, Ueno T, Kominami E (2008) LC3 and autophagy. Methods Mol Biol 445:77–88CrossRefPubMedGoogle Scholar
  27. 27.
    Schlafli AM, Berezowska S, Adams O, Langer R, Tschan MP (2015) Reliable LC3 and p62 autophagy marker detection in formalin fixed paraffin embedded human tissue by immunohistochemistry. Eur J Histochem 59:2481CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Huang Y, Chen HJ, Zhu JH, Zhao FY, Qu Y, Mu DZ (2016) Effects of PINK1 gene on cell apoptosis and cell autophagy in neonatal mice with hypoxic-ischemic brain damage. J Contemp Pediatr [Zhongguo dang dai er ke za zhi = Chinese] 18:263–269Google Scholar
  29. 29.
    Duan X, Zhang T, Ding S, Wei J, Su C, Liu H, Xu G (2015) microRNA-17-5p modulates bacille calmette-guerin growth in RAW264.7 cells by targeting ULK1. PLoS ONE 10:e0138011CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Zhang XY, Wu XQ, Deng R, Sun T, Feng GK, Zhu XF (2013) Upregulation of sestrin 2 expression via JNK pathway activation contributes to autophagy induction in cancer cells. Cell Signal 25:150–158CrossRefPubMedGoogle Scholar
  31. 31.
    Hansen K, Wagner B, Hamel W, Schweizer M, Haag F, Westphal M, Lamszus K (2007) Autophagic cell death induced by TrkA receptor activation in human glioblastoma cells. J Neurochem 103:259–275CrossRefPubMedGoogle Scholar
  32. 32.
    Zhao D, Wang C, Tang S, Zhang C, Zhang S, Zhou Y, Xiao X (2015) Reactive oxygen species-dependent JNK downregulated olaquindox-induced autophagy in HepG2 cells. J Appl Toxicol 35:709–716CrossRefPubMedGoogle Scholar
  33. 33.
    Chen SY, Maa MC, Chiu LY, Wang JS, Chien CL, Lin WW (2010) WITHDRAWN: zVAD-induced autophagic cell death requires c-Src-dependent ERK and JNK activation and reactive oxygen species generation. Autophagy 6:1CrossRefPubMedGoogle Scholar
  34. 34.
    Sheng Y, Sun B, Guo WT, Zhang YH, Liu X, Xing Y, Dong DL (2013) 3-Methyladenine induces cell death and its interaction with chemotherapeutic drugs is independent of autophagy. Biochem Biophys Res Commun 432:5–9CrossRefPubMedGoogle Scholar

Copyright information

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

Authors and Affiliations

  • Li Wang
    • 1
  • Peng Wang
    • 1
  • He Dong
    • 1
  • Shiduan Wang
    • 1
  • Haichen Chu
    • 1
  • Wei Yan
    • 1
  • Xue Zhang
    • 1
  1. 1.Department of AnesthesiologyThe Affiliated Hospital of Qingdao UniversityQingdaoChina

Personalised recommendations