Abstract
Heat shock protein 70 (Hsp70) is an evolutionarily highly conserved molecular chaperone that promotes the survival of stressed cells by inhibiting lysosomal membrane permeabilization1,2,3,4,5, a hallmark of stress-induced cell death6,7,8,9,10. Clues to its molecular mechanism of action may lay in the recently reported stress- and cancer-associated translocation of a small portion of Hsp70 to the lysosomal compartment1,11. Here we show that Hsp70 stabilizes lysosomes by binding to an endolysosomal anionic phospholipid bis(monoacylglycero)phosphate (BMP), an essential co-factor for lysosomal sphingomyelin metabolism12,13,14. In acidic environments Hsp70 binds with high affinity and specificity to BMP, thereby facilitating the BMP binding and activity of acid sphingomyelinase (ASM). The inhibition of the Hsp70–BMP interaction by BMP antibodies or a point mutation in Hsp70 (Trp90Phe), as well as the pharmacological and genetic inhibition of ASM, effectively revert the Hsp70-mediated stabilization of lysosomes. Notably, the reduced ASM activity in cells from patients with Niemann–Pick disease (NPD) A and B—severe lysosomal storage disorders caused by mutations in the sphingomyelin phosphodiesterase 1 gene (SMPD1) encoding for ASM15—is also associated with a marked decrease in lysosomal stability, and this phenotype can be effectively corrected by treatment with recombinant Hsp70. Taken together, these data open exciting possibilities for the development of new treatments for lysosomal storage disorders and cancer with compounds that enter the lysosomal lumen by the endocytic delivery pathway.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 51 print issues and online access
$199.00 per year
only $3.90 per issue
Rent or buy this article
Prices vary by article type
from$1.95
to$39.95
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Nylandsted, J. et al. Heat shock protein 70 promotes cell survival by inhibiting lysosomal membrane permeabilization. J. Exp. Med. 200, 425–435 (2004)
Hwang, J. H. et al. Spontaneous activation of pancreas trypsinogen in heat shock protein 70.1 knock-out mice. Pancreas 31, 332–336 (2005)
Gyrd-Hansen, M. et al. Apoptosome-independent activation of lysosomal cell death pathway by caspase-9. Mol. Cell. Biol. 26, 7880–7891 (2006)
Bivik, C., Rosdahl, I. & Ollinger, K. Hsp70 protects against UVB induced apoptosis by preventing release of cathepsins and cytochrome c in human melanocytes. Carcinogenesis 28, 537–544 (2007)
Doulias, P. T. et al. Involvement of heat shock protein-70 in the mechanism of hydrogen peroxide-induced DNA damage: the role of lysosomes and iron. Free Radic. Biol. Med. 42, 567–577 (2007)
Brunk, U. T., Neuzil, J. & Eaton, J. W. Lysosomal involvement in apoptosis. Redox Rep. 6, 91–97 (2001)
Leist, M. & Jäättelä, M. Four deaths and a funeral: from caspases to alternative mechanisms. Nature Rev. Mol. Cell Biol. 2, 589–598 (2001)
Guicciardi, M. E., Leist, M. & Gores, G. J. Lysosomes in cell death. Oncogene 23, 2881–2890 (2004)
Stoka, V., Turk, V. & Turk, B. Lysosomal cysteine cathepsins: signaling pathways in apoptosis. Biol. Chem. 388, 555–560 (2007)
Kirkegaard, T. & Jäättelä, M. Lysosomal involvement in cell death and cancer. Biochim. Biophys. Acta 1793, 746–754 (2009)
Mambula, S. S. & Calderwood, S. K. Heat shock protein 70 is secreted from tumor cells by a nonclassical pathway involving lysosomal endosomes. J. Immunol. 177, 7849–7857 (2006)
Quintern, L. E. et al. Acid sphingomyelinase from human urine: purification and characterization. Biochim. Biophys. Acta 922, 323–336 (1987)
Linke, T. et al. Stimulation of acid sphingomyelinase activity by lysosomal lipids and sphingolipid activator proteins. Biol. Chem. 382, 283–290 (2001)
Kolter, T. & Sandhoff, K. Principles of lysosomal membrane digestion: stimulation of sphingolipid degradation by sphingolipid activator proteins and anionic lysosomal lipids. Annu. Rev. Cell Dev. Biol. 21, 81–103 (2005)
Ferlinz, K., Hurwitz, R. & Sandhoff, K. Molecular basis of acid sphingomyelinase deficiency in a patient with Niemann-Pick disease type A. Biochem. Biophys. Res. Commun. 179, 1187–1191 (1991)
Daugaard, M., Rohde, M. & Jäättelä, M. The heat shock protein 70 family: highly homologous proteins with overlapping and distinct functions. FEBS Lett. 581, 3702–3710 (2007)
Kobayashi, T. et al. A lipid associated with the antiphospholipid syndrome regulates endosome structure and function. Nature 392, 193–197 (1998)
Kölzer, M., Werth, N. & Sandhoff, K. Interactions of acid sphingomyelinase and lipid bilayers in the presence of the tricyclic antidepressant desipramine. FEBS Lett. 559, 96–98 (2004)
Holopainen, J. M., Subramanian, M. & Kinnunen, P. K. Sphingomyelinase induces lipid microdomain formation in a fluid phosphatidylcholine/sphingomyelin membrane. Biochemistry 37, 17562–17570 (1998)
Goñi, F. M. & Alonso, A. Sphingomyelinases: enzymology and membrane activity. FEBS Lett. 531, 38–46 (2002)
Utermöhlen, O., Herz, J., Schramm, M. & Kronke, M. Fusogenicity of membranes: the impact of acid sphingomyelinase on innate immune responses. Immunobiology 213, 307–314 (2008)
Smith, E. L. & Schuchman, E. H. The unexpected role of acid sphingomyelinase in cell death and the pathophysiology of common diseases. FASEB J. 22, 3419–3431 (2008)
Tidwell, J. L., Houenou, L. J. & Tytell, M. Administration of Hsp70 in vivo inhibits motor and sensory neuron degeneration. Cell Stress Chaperones 9, 88–98 (2004)
Yu, Q., Kent, C. R. & Tytell, M. Retinal uptake of intravitreally injected Hsc/Hsp70 and its effect on susceptibility to light damage. Mol. Vis. 7, 48–56 (2001)
Flerov, M. A. et al. The use of HSP70 for prevention of consequences of unavoidable stress in rats. Bull. Exp. Biol. Med. 136, 120–122 (2003)
Brunk, U. T., Dalen, H., Roberg, K. & Hellquist, H. B. Photo-oxidative disruption of lysosomal membranes causes apoptosis of cultured human fibroblasts. Free Radic. Biol. Med. 23, 616–626 (1997)
Holopainen, J. M., Saily, M., Caldentey, J. & Kinnunen, P. K. The assembly factor P17 from bacteriophage PRD1 interacts with positively charged lipid membranes. Eur. J. Biochem. 267, 6231–6238 (2000)
Zhao, H. & Kinnunen, P. K. Binding of the antimicrobial peptide temporin L to liposomes assessed by Trp fluorescence. J. Biol. Chem. 277, 25170–25177 (2002)
Rohde, M. et al. Members of the heat-shock protein 70 family promote cancer cell growth by distinct mechanisms. Genes Dev. 19, 570–582 (2005)
Bligh, E. G. & Dyer, W. J. A rapid method of total lipid extraction and purification. Can. J. Biochem. Physiol. 37, 911–917 (1959)
Guex, N. & Peitsch, M. C. SWISS-MODEL and the Swiss-PdbViewer: an environment for comparative protein modeling. Electrophoresis 18, 2714–2723 (1997)
Sriram, M., Osipiuk, J., Freeman, B., Morimoto, R. & Joachimiak, A. Human Hsp70 molecular chaperone binds two calcium ions within the ATPase domain. Structure 5, 403–414 (1997)
Morshauser, R. C. et al. High-resolution solution structure of the 18 kDa substrate-binding domain of the mammalian chaperone protein Hsc70. J. Mol. Biol. 289, 1387–1403 (1999)
Acknowledgements
We thank E. Gulbins and A. Riehle for the NPD A (no. 2) cells, the Charité Berlin and the Johannes Gutenberg-Universität Mainz for the NPD B fibroblasts, C. Ejsing and H. Schulze for advice, and J. Grüenberg, B. Margulis and the Developmental Studies Hybridoma Bank for antibodies. This work was supported by grants from the Danish Cancer Society, the Danish National Research Foundation, the Danish Medical Research Council, the European Commission FP7 (APO-SYS), the Meyer Foundation, the Novo Nordisk Foundation, and the Association for International Cancer Research for M.J., the Finnish Academy and the Sigrid Juselius Foundation for P.K.J.K., Volkswagenstiftung for A.G.R. and C.A., Deutsche Forschungsgemeinschaft (SFB 645, SPP 1267) and EU (LipidomicNet) for K.S., and the Polish Ministry of Science and High Education for A.Z.
Author Contributions T.K. and M.J. designed the study, analysed the data and wrote the paper. T.K. performed experiments for Figs 1a–e, 2b, e, f, 3b, d, f, 4a–c, Supplementary Figs 1, 4 and Supplementary Movies. A.G.R. and C.A. performed experiments for Figs 2d, 3a and Supplementary Fig. 6, and provided NPD cell lines. K.S. contributed to the BIAcore data and provided rASM. N.H.T.P. performed experiments for Fig. 3e and Supplementary Fig. 3. P.K.J.K. designed the biophysical studies on Hsp–lipid interactions. A.K.M. and I.M. performed experiments for Fig. 2a, e. O.D.O. and J.K. performed experiments for Figs 3c, 4d and Supplementary Figs 2 and 5. A.Z. validated the functionality of rHsp70 and its mutants, and provided proteins for control. J.N. provided input to study design. All authors discussed the results and commented on the manuscript.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
The authors declare no competing financial interests.
Supplementary information
Supplementary Information
This file contains Supplementary Figures S1-S6 with Legends and Legends for Supplementary Movies 1 - 2. (PDF 2954 kb)
Supplementary Movie 1
Photo-oxidation-induced lysosomal membrane permeabilization in U-2-OS cells labeled with acridine orange (see Supplementary Information file for full Legend). (MOV 5577 kb)
Supplementary Movie 2
Massive photo-oxidation-induced lysosomal membrane permeabilization in NPDA fibroblasts labeled with acridine orange (see Supplementary Information file for full Legend). (AVI 3804 kb)
Rights and permissions
About this article
Cite this article
Kirkegaard, T., Roth, A., Petersen, N. et al. Hsp70 stabilizes lysosomes and reverts Niemann–Pick disease-associated lysosomal pathology. Nature 463, 549–553 (2010). https://doi.org/10.1038/nature08710
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/nature08710
This article is cited by
-
MLKL polymerization-induced lysosomal membrane permeabilization promotes necroptosis
Cell Death & Differentiation (2024)
-
Progranulin and GPNMB: interactions in endo-lysosome function and inflammation in neurodegenerative disease
Journal of Neuroinflammation (2023)
-
Lysosomal phospholipase A2 contributes to the biosynthesis of the atypical late endosome lipid bis(monoacylglycero)phosphate
Communications Biology (2023)
-
RIPK1 inhibition contributes to lysosomal membrane stabilization in ischemic astrocytes via a lysosomal Hsp70.1B-dependent mechanism
Acta Pharmacologica Sinica (2023)
-
Heat shock protein A2 is a novel extracellular vesicle-associated protein
Scientific Reports (2023)
Comments
By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.