Skip to main content
SpringerLink
Log in

Beclin-1 Deficiency Alters Autophagosome Formation, Lysosome Biogenesis and Enhances Neuronal Vulnerability of HT22 Hippocampal Cells

  • Published:
Molecular Neurobiology Aims and scope Submit manuscript

Abstract

Beclin-1 is assumed to be a critical component participating in autophagosome formation in mammals; however, the exact role of Beclin-1 in autophagy remains controversial. Here (1) we created a HT22-Beclin-1-knockdown cell line using the Q-techBECN1 technique, (2) examined the potential role of Beclin-1 in an autophagic response in hippocampal HT22 neurons challenged with rapamycin, (3) investigated the expression of several gene products involved in the autophagic pathway, and (4) checked the effects of Beclin-1 knockdown on neuronal death induced by AAS. Rapamycin induced and altered the expression of autophagy signature proteins in wild-type cultures as well as in HT22-Beclin-1-knockdown cells. However, among the examined markers, only two factors exhibited dramatic changes when comparing controls to HT22-Beclin-1-knockdown cells. The amount of LC3, an important protein for the initiation of autophagosome formation and LAMP-1, a major constituent of the lysosomal membrane, underwent a dramatic and highly significant increase in control cultures challenged with rapamycin. In contrast, rapamycin was not able to induce any significant changes in LC3 and LAMP-1 levels in HT22-Beclin-1-knockdown cells. In addition, the knockdown of Beclin-1 enhanced neuronal susceptibility to death signals induced by AAS. Our data demonstrate the essential role of Beclin-1 in the formation of autophagosomes and lysosome biogenesis and underline that deletion of this key system is deleterious for cell viability.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. Yoshimori T (2004) Autophagy: a regulated bulk degradation process inside cells. Biochem Biophys Res Commun 313(2):453–8

    Article  CAS  PubMed  Google Scholar 

  2. Kabeya Y, Mizushima N, Ueno T, Yamamoto A, Kirisako T, Noda T, Kominami E, Ohsumi Y et al (2000) LC3, a mammalian homologue of yeast Apg8p, is localized in autophagosome membranes after processing. EMBO J 19:5720–5728

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Klionsky DJ, Emr SD (2000) Autophagy as a regulated pathway of cellular degradation. Science 290:1717–1721

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Klionsky DJ, Abeliovich H, Agostinis P et al (2008) Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes. Autophagy 4:151–175

    Article  CAS  PubMed  Google Scholar 

  5. Liang XH, Kleeman LK, Jiang HH, Gordon G, Goldman JE, Berry G, Herman B, Levine B (1998) Protection against fatal Sindbis virus encephalitis by beclin, a novel Bcl-2-interacting protein. J Virol 72:8586–8596

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Levine B, Klionsky DJ (2004) Development by self-digestion: molecular mechanisms and biological functions of autophagy. Dev Cell 6:463–477

    Article  CAS  PubMed  Google Scholar 

  7. Kihara A, Kabeya Y, Ohsumi Y, Yoshimori T (2001) Beclin-phosphatidylinositol 3-kinase complex functions at the trans-Golgi network. EMBO Rep 2:330–335

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Qu X, Yu J, Bhagat G, Furuya N, Hibshoosh H, Troxel A, Rosen J, Eskelinen EL et al (2003) Promotion of tumorigenesis by heterozygous disruption of the beclin 1 autophagy gene. J Clin Invest 112:1809–1820

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Yue Z, Jin S, Yang C, Levine AJ, Heintz N (2003) Beclin 1, an autophagy gene essential for early embryonic development, is a haploinsufficient tumor suppressor. Proc Natl Acad Sci U S A 100:15077–15082

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Komatsu M, Waguri S, Chiba T, Murata S, Iwata J, Tanida I, Ueno T, Koike M et al (2006) Loss of autophagy in the central nervous system causes neurodegeneration in mice. Nature 441:880–884

    Article  CAS  PubMed  Google Scholar 

  11. Hara T, Nakamura K, Matsui M, Yamamoto A, Nakahara Y, Suzuki-Migishima R, Yokoyama M, Mishima K et al (2006) Suppression of basal autophagy in neural cells causes neurodegenerative disease in mice. Nature 441:885–889

    Article  CAS  PubMed  Google Scholar 

  12. Pickford F, Masliah E, Britschgi M, Lucin K, Narasimhan R, Jaeger PA, Small S, Spencer B et al (2008) The autophagy-related protein beclin 1 shows reduced expression in early Alzheimer disease and regulates amyloid beta accumulation in mice. J Clin Invest 118(6):2190–9

    CAS  PubMed  PubMed Central  Google Scholar 

  13. Spencer B, Potkar R, Trejo M, Rockenstein E, Patrick C, Gindi R, Adame A, Wyss-Coray T et al (2009) Beclin 1 gene transfer activates autophagy and ameliorates the neurodegenerative pathology in alpha-synuclein models of Parkinson’s and Lewy body diseases. J Neurosci 29(43):13578–88

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Nascimento-Ferreira I, Santos-Ferreira T, Sousa-Ferreira L, Auregan G, Onofre I, Alves S, Dufour N, Colomer Gould VF et al (2011) Overexpression of the autophagic beclin-1 protein clears mutant ataxin-3 and alleviates Machado-Joseph disease. Brain 134(Pt 5):1400–15

    Article  PubMed  Google Scholar 

  15. Sun T, Li X, Zhang P, Chen WD, Zhang HL, Li DD, Deng R, Qian XJ et al (2015) Acetylation of Beclin 1 inhibits autophagosome maturation and promotes tumour growth. Nat Commun 6:7215

    Article  PubMed  PubMed Central  Google Scholar 

  16. Copetti T, Bertoli C, Dalla E, Demarchi F, Schneider C (2009) p65/RelA modulates BECN1 transcription and autophagy. Mol Cell Biol 29(10):2594–608

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Zhu JH, Horbinski C, Guo F, Watkins S, Uchiyama Y, Chu CT (2007) Regulation of autophagy by extracellular signal-regulated protein kinases during 1-methyl-4-phenylpyridinium-induced cell death. Am J Pathol 170(1):75–86

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Grishchuk Y, Ginet V, Truttmann AC, Clarke PG, Puyal J (2011) Beclin 1-independent autophagy contributes to apoptosis in cortical neurons. Autophagy 7:1115–1131

    Article  CAS  PubMed  Google Scholar 

  19. McCoy F, Hurwitz J, McTavish N, Paul I, Barnes C, O’Hagan B, Odrzywol K, Murray J et al (2010) Obatoclax induces Atg7-dependent autophagy independent of beclin-1 and BAX/BAK. Cell Death Dis 1:e108

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. McKnight NC, Zhong Y, Wold MS, Gong S, Phillips GR, Dou Z, Zhao Y, Heintz N et al (2014) Beclin 1 is required for neuron viability and regulates endosome pathways via the UVRAG-VPS34 complex. PLoS Genet 2:10(10)

    Google Scholar 

  21. Matsunaga K, Saitoh T, Tabata K, Omori H, Satoh T, Kurotori N, Maejima I, Shirahama-Noda K et al (2009) Two Beclin 1-binding proteins, Atg14L and Rubicon, reciprocally regulate autophagy at different stages. Nat Cell Biol 11(4):385–96

    Article  CAS  PubMed  Google Scholar 

  22. Zhong Y, Wang QJ, Yue Z (2009) Atg14L and Rubicon: yin and yang of Beclin 1-mediated autophagy control. Autophagy 5(6):890–1

    Article  PubMed  Google Scholar 

  23. Itakura E, Kishi C, Inoue K, Mizushima N (2008) Beclin 1 forms two distinct phosphatidylinositol 3-kinase complexes with mammalian Atg14 and UVRAG. Mol Biol Cell 19(12):5360–72

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Zhou F, Yang Y, Xing D (2011) Bcl-2 and Bcl-xL play important roles in the crosstalk between autophagy and apoptosis. FEBS J 278:403–413

    Article  CAS  PubMed  Google Scholar 

  25. Kang R, Tang D, Loze MT, Zeh HJ (2011) Apoptosis to autophagy switch triggered by the MHC class III-encoded receptor for advanced glycation endproducts (RAGE). Autophagy 7(1):91–3

    Article  PubMed  Google Scholar 

  26. Kuma A, Hatano M, Matsui M, Yamamoto A, Nakaya H, Yoshimori T, Ohsumi Y, Tokuhisa T (2004) The role of autophagy during the early neonatal starvation period. Nature 432:1032–1036

    Article  CAS  PubMed  Google Scholar 

  27. Rami A, Kögel D (2008) Apoptosis meets autophagy-like cell death in the ischemic penumbra: two sides of the same coin? Autophagy 4(4):422–26

    Article  CAS  PubMed  Google Scholar 

  28. Nikoletopoulou V, Markaki M, Palikaras K, Tavernarakis N (2013) Crosstalk between apoptosis, necrosis and autophagy. Biochim Biophys Acta 1833:3448–3459

    Article  CAS  PubMed  Google Scholar 

  29. Steiger-Barraissoul S, Rami A (2009) Serum deprivation induced autophagy and predominantly an AIF-dependent apoptosis in hippocampal HT22 neurons. Apoptosis 14:1274–88

    Article  CAS  PubMed  Google Scholar 

  30. Rami A (2009) Review: autophagy in neurodegeneration: firefighter and/or incendiarist? Neuropathol Appl Neurobiol 35(5):449–61

    Article  CAS  PubMed  Google Scholar 

  31. Kang R, Zeh HJ, Lotze MT, Tang D (2011) The Beclin 1 network regulates autophagy and apoptosis. Cell Death Differ 18(4):571–80

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgments

This work was supported by the Adolf-Messer-Stiftung (grant to Dr. A. Rami—molecular mechanisms of autophagy). JF was supported partly by the Adolf-Messer-Stiftung and the FPF (Frankfurter Promotionsförderung). We thank J. Stehle for the scientific support of our work.

Author information

Authors and Affiliations

  1. Institut für Zelluläre und Molekulare Anatomie (Anatomie III), Klinikum der Johann Wolfgang von Goethe-Universität, Theodor-Stern-Kai 7, 60590, Frankfurt/Main, Germany

    J. Fekadu & A. Rami

Authors
  1. J. Fekadu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. A. Rami
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to A. Rami.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Fekadu, J., Rami, A. Beclin-1 Deficiency Alters Autophagosome Formation, Lysosome Biogenesis and Enhances Neuronal Vulnerability of HT22 Hippocampal Cells. Mol Neurobiol 53, 5500–5509 (2016). https://doi.org/10.1007/s12035-015-9453-2

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12035-015-9453-2

Keywords

Search

Navigation