Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

ULK1 induces autophagy by phosphorylating Beclin-1 and activating VPS34 lipid kinase

Nature Cell Biology volume 15, pages 741–750 (2013)Cite this article

Subjects

Abstract

Autophagy is the primary cellular catabolic program activated in response to nutrient starvation. Initiation of autophagy, particularly by amino-acid withdrawal, requires the ULK kinases. Despite its pivotal role in autophagy initiation, little is known about the mechanisms by which ULK promotes autophagy. Here we describe a molecular mechanism linking ULK to the pro-autophagic lipid kinase VPS34. Following amino-acid starvation or mTOR inhibition, the activated ULK1 phosphorylates Beclin-1 on Ser 14, thereby enhancing the activity of the ATG14L-containing VPS34 complexes. The Beclin-1 Ser 14 phosphorylation by ULK is required for full autophagic induction in mammals and this requirement is conserved in Caenorhabditis elegans. Our study reveals a molecular link from ULK1 to activation of the autophagy-specific VPS34 complex and autophagy induction.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: ULK is essential for activation of the ATG14L-associated VPS34 following amino-acid starvation.
Figure 2: Beclin-1 Ser 14 is phosphorylated by ULK1 and required for VPS34 activation in response to amino-acid withdrawal.
Figure 3: Beclin-1 is a physiological target of ULK kinase in response to amino-acid withdrawal and mTOR inhibition.
Figure 4: ATG14L stimulates Beclin-1 Ser 14 phosphorylation by promoting association with ULK1.
Figure 5: UVRAG promotes Beclin-1 Ser 14 phosphorylation and association with ULK1.
Figure 6: Beclin-1 Ser 14 phosphorylation plays a critical role in autophagy induction by amino-acid starvation.
Figure 7: The conserved ULK phosphorylation site in C. elegans Bec-1 is required for autophagy.
Figure 8: A working model of VPS34 complex regulation by ULK on amino-acid withdrawal.

Similar content being viewed by others

References

  1. Mizushima, N. & Komatsu, M. Autophagy: renovation of cells and tissues. Cell 147, 728–741 (2011).

    Article  CAS  Google Scholar 

  2. Young, A. R. et al. Starvation and ULK1-dependent cycling of mammalian Atg9 between the TGN and endosomes. J. Cell Sci. 119, 3888–3900 (2006).

    Article  CAS  Google Scholar 

  3. Hara, T. et al. FIP200, a ULK-interacting protein, is required for autophagosome formation in mammalian cells. J. Cell Biol. 181, 497–510 (2008).

    Article  CAS  Google Scholar 

  4. Chan, E. Y., Longatti, A., McKnight, N. C. & Tooze, S. A. Kinase-inactivated ULK proteins inhibit autophagy via their conserved C-terminal domains using an Atg13-independent mechanism. Mol. Cell Biol. 29, 157–171 (2009).

    Article  CAS  Google Scholar 

  5. Backer, J. M. The regulation and function of Class III PI3Ks: novel roles for Vps34. Biochem. J. 410, 1–17 (2008).

    Article  CAS  Google Scholar 

  6. Yan, J. et al. Mouse ULK2, a novel member of the UNC-51-like protein kinases: unique features of functional domains. Oncogene 18, 5850–5859 (1999).

    Article  CAS  Google Scholar 

  7. Ganley, I. G. et al. ULK1.ATG13.FIP200 complex mediates mTOR signaling and is essential for autophagy. J. Biol. Chem. 284, 12297–12305 (2009).

    Article  CAS  Google Scholar 

  8. Hosokawa, N. et al. Nutrient-dependent mTORC1 association with the ULK1-Atg13-FIP200 complex required for autophagy. Mol. Biol. Cell 20, 1981–1991 (2009).

    Article  CAS  Google Scholar 

  9. Jung, C. H. et al. ULK–Atg13–FIP200 complexes mediate mTOR signaling to the autophagy machinery. Mol. Biol. Cell 20, 1992–2003 (2009).

    Article  CAS  Google Scholar 

  10. Kim, J., Kundu, M., Viollet, B. & Guan, K. L. AMPK and mTOR regulate autophagy through direct phosphorylation of Ulk1. Nat. Cell Biol. 13, 132–141 (2011).

    Article  CAS  Google Scholar 

  11. Egan, D. F. et al. Phosphorylation of ULK1 (hATG1) by AMP-activated protein kinase connects energy sensing to mitophagy. Science 331, 456–461 (2011).

    Article  CAS  Google Scholar 

  12. Itakura, E. & Mizushima, N. Characterization of autophagosome formation site by a hierarchical analysis of mammalian Atg proteins. Autophagy 6, 764–776 (2010).

    Article  CAS  Google Scholar 

  13. Volinia, S. et al. A human phosphatidylinositol 3-kinase complex related to the yeast Vps34p–Vps15p protein sorting system. EMBO J. 14, 3339–3348 (1995).

    Article  CAS  Google Scholar 

  14. Sun, Q. et al. Identification of Barkor as a mammalian autophagy-specific factor for Beclin 1 and class III phosphatidylinositol 3-kinase. Proc. Natl Acad. Sci. USA 105, 19211–19216 (2008).

    Article  CAS  Google Scholar 

  15. Itakura, E., Kishi, C., Inoue, K. & Mizushima, N. Beclin 1 forms two distinct phosphatidylinositol 3-kinase complexes with mammalian Atg14 and UVRAG. Mol. Biol. Cell 19, 5360–5372 (2008).

    Article  CAS  Google Scholar 

  16. Liang, C. et al. Autophagic and tumour suppressor activity of a novel Beclin1-binding protein UVRAG. Nat. Cell Biol. 8, 688–699 (2006).

    Article  CAS  Google Scholar 

  17. Takahashi, Y. et al. Bif-1 interacts with Beclin 1 through UVRAG and regulates autophagy and tumorigenesis. Nat. Cell Biol. 9, 1142–1151 (2007).

    Article  CAS  Google Scholar 

  18. Di Bartolomeo, S. et al. The dynamic interaction of AMBRA1 with the dynein motor complex regulates mammalian autophagy. J. Cell Biol. 191, 155–168 (2010).

    Article  CAS  Google Scholar 

  19. Pattingre, S. et al. Role of JNK1-dependent Bcl-2 phosphorylation in ceramide-induced macroautophagy. J. Biol. Chem. 284, 2719–2728 (2009).

    Article  CAS  Google Scholar 

  20. Zalckvar, E. et al. DAP-kinase-mediated phosphorylation on the BH3 domain of beclin 1 promotes dissociation of beclin 1 from Bcl-XL and induction of autophagy. EMBO Rep. 10, 285–292 (2009).

    Article  CAS  Google Scholar 

  21. Zhong, Y. et al. Distinct regulation of autophagic activity by Atg14L and Rubicon associated with Beclin 1-phosphatidylinositol-3-kinase complex. Nat. Cell Biol. 11, 468–476 (2009).

    Article  CAS  Google Scholar 

  22. Matsunaga, K. et al. Two Beclin 1-binding proteins, Atg14L and Rubicon, reciprocally regulate autophagy at different stages. Nat. Cell Biol. 11, 385–396 (2009).

    Article  CAS  Google Scholar 

  23. Matsunaga, K. et al. Autophagy requires endoplasmic reticulum targeting of the PI3-kinase complex via Atg14L. J. Cell Biol. 190, 511–521 (2010).

    Article  CAS  Google Scholar 

  24. Kim, J. et al. Differential regulation of distinct Vps34 complexes by AMPK in nutrient stress and autophagy. Cell 152, 290–303 (2013).

    Article  CAS  Google Scholar 

  25. Byfield, M. P., Murray, J. T. & Backer, J. M. hVps34 is a nutrient-regulated lipid kinase required for activation of p70 S6 kinase. J. Biol. Chem. 280, 33076–33082 (2005).

    Article  CAS  Google Scholar 

  26. Gulati, P. et al. Amino acids activate mTOR complex 1 via Ca2+/CaM signaling to hVps34. Cell Metab. 7, 456–465 (2008).

    Article  CAS  Google Scholar 

  27. Nobukuni, T. et al. Amino acids mediate mTOR/raptor signaling through activation of class 3 phosphatidylinositol 3OH-kinase. Proc. Natl Acad. Sci. USA 102, 14238–14243 (2005).

    Article  CAS  Google Scholar 

  28. Wei, Y., Pattingre, S., Sinha, S., Bassik, M. & Levine, B. JNK1-mediated phosphorylation of Bcl-2 regulates starvation-induced autophagy. Mol. Cell 30, 678–688 (2008).

    Article  CAS  Google Scholar 

  29. Lee, E. J. & Tournier, C. The requirement of uncoordinated 51-like kinase 1 (ULK1) and ULK2 in the regulation of autophagy. Autophagy 7, 689–695 (2011).

    Article  CAS  Google Scholar 

  30. Gillooly, D. J. et al. Localization of phosphatidylinositol 3-phosphate in yeast and mammalian cells. EMBO J. 19, 4577–4588 (2000).

    Article  CAS  Google Scholar 

  31. Bach, M., Larance, M., James, D.E. & Ramm, G. The serine/threonine kinase ULK1 is a target of multiple phosphorylation events. Biochem. J. 440, 283–291 (2011).

    Article  CAS  Google Scholar 

  32. Fan, W., Nassiri, A. & Zhong, Q. Autophagosome targeting and membrane curvature sensing by Barkor/Atg14(L). Proc. Natl Acad. Sci. USA 108, 7769–7774 (2011).

    Article  CAS  Google Scholar 

  33. Zhang, Y. et al. SEPA-1 mediates the specific recognition and degradation of P granule components by autophagy in C. elegans. Cell 136, 308–321 (2009).

    Article  CAS  Google Scholar 

  34. Zhao, Y., Tian, E. & Zhang, H. Selective autophagic degradation of maternally-loaded germline P granule components in somatic cells during C. elegans embryogenesis. Autophagy 5, 717–719 (2009).

    Article  CAS  Google Scholar 

  35. Tian, Y. et al. C. elegans screen identifies autophagy genes specific to multicellular organisms. Cell 141, 1042–1055 (2010).

    Article  CAS  Google Scholar 

  36. Petiot, A., Ogier-Denis, E., Blommaart, E. F., Meijer, A. J. & Codogno, P. Distinct classes of phosphatidylinositol 3′-kinases are involved in signaling pathways that control macroautophagy in HT-29 cells. J. Biol. Chem. 275, 992–998 (2000).

    Article  CAS  Google Scholar 

  37. Obara, K. & Ohsumi, Y. PtdIns 3-kinase orchestrates autophagosome formation in yeast. J. Lipids 2011, 498768 (2011).

    Article  Google Scholar 

  38. Suzuki, K., Kubota, Y., Sekito, T. & Ohsumi, Y. Hierarchy of Atg proteins in pre-autophagosomal structure organization.. Genes Cells 12, 209–218 (2007).

    Article  CAS  Google Scholar 

  39. Obara, K., Noda, T., Niimi, K. & Ohsumi, Y. Transport of phosphatidylinositol 3-phosphate into the vacuole via autophagic membranes in Saccharomyces cerevisiae. Genes Cells 13, 537–547 (2008).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We would like to thank H. Zhang for LGG-1 and PGL-1 antibodies; J. Jewell, C. Hansen and K. Tumaneng for critical reading of this manuscript; and M. Farquhar for electron microscopy. Phosphosite identification by mass spectrometry was performed by the Proteomics Facility at the Fred Hutchinson Cancer Research Center. Confocal analysis was performed at the UCSD Neuroscience Microscopy Shared Facility (Grant P30 NS047101). This work was supported by National Institutes of Health (NIH) grants GM51586, GM62694 and CA108941, and the Department of Defense (W81XWH-0901-0279). R.C.R. is supported by a Canadian Institutes of Health Research (CIHR) postdoctoral fellowship.

Author information

Authors and Affiliations

  1. Department of Pharmacology and Moores Cancer Center, University of California San Diego, La Jolla, California 92093, USA

    Ryan C. Russell, Haixin Yuan, Hyun Woo Park, Joungmok Kim, Haerin Kim & Kun-Liang Guan

  2. Department of Molecular and Cell Biology, University of California Berkeley, Berkeley, California 94720, USA

    Ye Tian & Andrew Dillin

  3. Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, Minnesota 55455, USA

    Yu-Yun Chang & Thomas P. Neufeld

  4. Department of Oral Biochemistry, School of Dentistry, Kyung Hee University, Seoul 130-701, Korea

    Joungmok Kim

Authors
  1. Ryan C. Russell
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ye Tian
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Haixin Yuan
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Hyun Woo Park
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Yu-Yun Chang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Joungmok Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Haerin Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Thomas P. Neufeld
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Andrew Dillin
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Kun-Liang Guan
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

R.C.R. planned and performed experiments, Y.T. planned and performed experiments, H.Y., H.W.P., Y-Y.C and H.K. performed experiments, J.K., T.P.N., A.D. and K-L.G. planned experiments, R.C.R. and K-L.G. wrote the manuscript.

Corresponding author

Correspondence to Kun-Liang Guan.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary Information (PDF 5907 kb)

Supplementary Table 1

Supplementary Information (XLSX 14 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Russell, R., Tian, Y., Yuan, H. et al. ULK1 induces autophagy by phosphorylating Beclin-1 and activating VPS34 lipid kinase. Nat Cell Biol 15, 741–750 (2013). https://doi.org/10.1038/ncb2757

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ncb2757

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing