The Co-Chaperone HspBP1 Is a Novel Component of Stress Granules that Regulates Their Formation
Abstract
1. Introduction
2. Materials and Methods
2.1. Cell Culture and Exposure to Stress
2.2. Antibodies and Pharmacological Reagents
2.3. Immunofluorescence and Microscopy
2.4. SG Quantification and Statistical Analyses
2.5. Protein Crosslinking and Indirect Immunoprecipitation
2.6. Mass Spectrometry
2.7. Oligo(dT) Binding Assay
2.8. In Vitro RNA Homopolymer Binding Assay
2.9. Western Blotting
2.10. In Situ Hybridization of PolyA-RNA
2.11. Transient Transfection
3. Results and Discussion
3.1. HspBP1 Is a Component of Oxidant-Induced SGs, but Does Not Concentrate in Cytoplasmic Processing Bodies
3.2. HspBP1 Binds the SG Marker Proteins G3BP1, HuR and TIA
3.3. HspBP1 Association with SGs during Granule Formation and Recovery
3.4. HspBP1 Binds PolyA-RNA In Vivo and Homopolymers In Vitro
3.5. HspBP1 Knockdown Impairs SG Formation
3.6. Full Length HspBP1 Overexpression Induces the Formation of Cytoplasmic SGs in the Absence of Stress
3.7. The hsp70-Interaction Domains of HspBP1 Stimulate SG Assembly in the Absence of Stress
3.8. Effects of Pharmacological Inhibition of hsp70 on SGs
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
- Bracher, A.; Verghese, J. GrpE, Hsp110/Grp170, HspBP1/Sil1 and BAG domain proteins: Nucleotide exchange factors for Hsp70 molecular chaperones. Subcell. Biochem. 2015, 78, 1–33. [Google Scholar] [PubMed]
- Kim, Y.E.; Hipp, M.S.; Bracher, A.; Hayer-Hartl, M.; Hartl, F.U. Molecular chaperone functions in protein folding and proteostasis. Annu. Rev. Biochem. 2013, 82, 323–355. [Google Scholar] [CrossRef] [PubMed]
- Brandvold, K.R.; Morimoto, R.I. The chemical biology of molecular chaperones - Implications for modulation of proteostasis. J. Mol. Biol. 2015, 427, 2931–2947. [Google Scholar] [CrossRef] [PubMed]
- Hipp, M.S.; Kasturi, P.; Hartl, F.U. The proteostasis network and its decline in ageing. Nat. Rev. Mol. Cell Biol. 2019, 20, 421–435. [Google Scholar] [CrossRef] [PubMed]
- Joshi, S.; Wang, T.; Araujo, T.L.S.; Sharma, S.; Brodsky, J.L.; Chiosis, G. Adapting to stress—Chaperome networks in cancer. Nature Reviews Cancer 2018, 18, 562–575. [Google Scholar] [CrossRef]
- Rosenzweig, R.; Nillegoda, N.B.; Mayer, M.P.; Bukau, B. The Hsp70 chaperone network. Nat. Rev. Mol. Cell Biol. 2019, 20, 665–680. [Google Scholar] [CrossRef]
- Shomura, Y.; Dragovic, Z.; Chang, H.C.; Tzvetkov, N.; Young, J.C.; Brodsky, J.L.; Guerriero, V.; Hartl, F.U.; Bracher, A. Regulation of Hsp70 function by HspBP1: Structural analysis reveals an alternate mechanism for Hsp70 nucleotide exchange. Mol. Cell 2005, 17, 367–379. [Google Scholar]
- Kabani, M.; McLellan, C.; Raynes, D.A.; Guerriero, V.; Brodsky, J.L. HspBP1, a homologue of the yeast Fes1 and Sls1 proteins, is an Hsc70 nucleotide exchange factor. FEBS Lett. 2002, 531, 339–342. [Google Scholar] [CrossRef]
- Zhao, T.; Hong, Y.; Yin, P.; Li, S.; Li, X.J. Differential HspBP1 expression accounts for the greater vulnerability of neurons than astrocytes to misfolded proteins. Proc. Natl. Acad. Sci. USA 2017, 114, E7803–e7811. [Google Scholar] [CrossRef]
- Tanimura, S.; Hirano, A.I.; Hashizume, J.; Yasunaga, M.; Kawabata, T.; Ozaki, K.; Kohno, M. Anticancer drugs up-regulate HspBP1 and thereby antagonize the prosurvival function of Hsp70 in tumor cells. J. Biol. Chem. 2007, 282, 35430–35439. [Google Scholar] [CrossRef]
- Yang, Z.; Zhuang, L.; Szatmary, P.; Wen, L.; Sun, H.; Lu, Y.; Xu, Q.; Chen, X. Upregulation of heat shock proteins (HSPA12A, HSP90B1, HSPA4, HSPA5 and HSPA6) in tumour tissues is associated with poor outcomes from HBV-related early-stage hepatocellular carcinoma. Int. J. Med. Sci. 2015, 12, 256–263. [Google Scholar] [CrossRef] [PubMed]
- Graner, M.W.; Raynes, D.A.; Bigner, D.D.; Guerriero, V. Heat shock protein 70-binding protein 1 is highly expressed in high-grade gliomas, interacts with multiple heat shock protein 70 family members, and specifically binds brain tumor cell surfaces. Cancer Sci. 2009, 100, 1870–1879. [Google Scholar] [CrossRef] [PubMed]
- Raynes, D.A.; Graner, M.W.; Bagatell, R.; McLellan, C.; Guerriero, V. Increased expression of the Hsp70 cochaperone HspBP1 in tumors. Tumour Biol. 2003, 24, 281–285. [Google Scholar] [CrossRef] [PubMed]
- Yashin, D.V.; Dukhanina, E.A.; Kabanova, O.D.; Romanova, E.A.; Lukyanova, T.I.; Tonevitskii, A.G.; Raynes, D.A.; Gnuchev, N.V.; Guerriero, V.; Georgiev, G.P.; et al. The heat shock-binding protein (HspBP1) protects cells against the cytotoxic action of the Tag7-Hsp70 complex. J. Biol. Chem. 2011, 286, 10258–10264. [Google Scholar] [CrossRef]
- Gottwald, E.; Herschbach, M.; Lahni, B.; Miesfeld, R.L.; Kunz, S.; Raynes, D.A.; Guerriero, V. Expression of the cochaperone HspBP1 is not coordinately regulated with Hsp70 expression. Cell Biol. Int. 2006, 30, 553–558. [Google Scholar] [CrossRef]
- Mahboubi, H.; Stochaj, U. Quantitative analysis of the interplay between hsc70 and its co-chaperone HspBP1 PeerJ. 2015, 3, e1530. 3.
- Chaudhary, P.; Khan, S.Z.; Rawat, P.; Augustine, T.; Raynes, D.A.; Guerriero, V.; Mitra, D. HSP70 binding protein 1 (HspBP1) suppresses HIV-1 replication by inhibiting NF-kappaB mediated activation of viral gene expression. Nucleic Acids Res. 2016, 44, 1613–1629. [Google Scholar] [CrossRef]
- Khong, A.; Matheny, T.; Jain, S.; Mitchell, S.F.; Wheeler, J.R.; Parker, R. The Stress Granule Transcriptome Reveals Principles of mRNA Accumulation in Stress Granules. Mol. Cell 2017, 68, 808–820. [Google Scholar] [CrossRef]
- Mahboubi, H.; Stochaj, U. Nucleoli and Stress Granules: Connecting Distant Relatives. Traffic 2014, 15, 1179–1193. [Google Scholar] [CrossRef]
- Kedersha, N.; Ivanov, P.; Anderson, P. Stress granules and cell signaling: More than just a passing phase? Trends Biochem. Sci. 2013, 38, 494–506. [Google Scholar] [CrossRef]
- Protter, D.S.W.; Parker, R. Principles and Properties of Stress Granules. Trends Cell Biol. 2016, 26, 668–679. [Google Scholar] [CrossRef]
- Wang, Z.; Zhang, H. Phase Separation, Transition, and Autophagic Degradation of Proteins in Development and Pathogenesis. Trends Cell Biol. 2019, 29, 417–427. [Google Scholar] [CrossRef] [PubMed]
- Mahboubi, H.; Stochaj, U. Cytoplasmic stress granules: Dynamic modulators of cell signaling and disease. Biochim. Biophys. Acta 2017, 1863, 884–895. [Google Scholar] [CrossRef] [PubMed]
- Arimoto, K.; Fukuda, H.; Imajoh-Ohmi, S.; Saito, H.; Takekawa, M. Formation of stress granules inhibits apoptosis by suppressing stress-responsive MAPK pathways. Nat. Cell Biol. 2008, 10, 1324–1332. [Google Scholar] [CrossRef] [PubMed]
- Mahboubi, H.; Koromilas, A.; Stochaj, U. AMP-kinase Activation Alters Oxidant-induced Stress Granule Assembly by Modulating Cell Signaling and Microtubule Organization. Mol. Pharmacol. 2016, 90, 460–468. [Google Scholar] [CrossRef] [PubMed]
- Anderson, P.; Kedersha, N.; Ivanov, P. Stress granules, P-bodies and cancer. Biochim. Biophys. Acta 2015, 1859, 861–870. [Google Scholar] [CrossRef] [PubMed]
- Shukla, S.; Parker, R. Hypo- and Hyper-Assembly Diseases of RNA-Protein Complexes. Trends Mol. Med. 2016, 22, 615–628. [Google Scholar] [CrossRef]
- Poblete-Duran, N.; Prades-Perez, Y.; Vera-Otarola, J.; Soto-Rifo, R.; Valiente-Echeverria, F. Who Regulates Whom? An Overview of RNA Granules and Viral Infections. Viruses 2016, 8, 180. [Google Scholar] [CrossRef]
- Baradaran-Heravi, Y.; Van Broeckhoven, C.; van der Zee, J. Stress granule mediated protein aggregation and underlying gene defects in the FTD-ALS spectrum. Neurobiol. Dis. 2020, 134, 104639. [Google Scholar] [CrossRef]
- Moujaber, O.; Stochaj, U. Cytoplasmic RNA granules in somatic maintenance. Gerontology 2018, 64, 485–494. [Google Scholar] [CrossRef]
- Gilks, N.; Kedersha, N.; Ayodele, M.; Shen, L.; Stoecklin, G.; Dember, L.M.; Anderson, P. Stress Granule Assembly Is Mediated by Prion-like Aggregation of TIA-1. Mol. Biol. Cell 2004, 15, 5383–5398. [Google Scholar] [CrossRef]
- Tourriere, H.; Chebli, K.; Zekri, L.; Courselaud, B.; Blanchard, J.M.; Bertrand, E.; Tazi, J. The RasGAP-associated endoribonuclease G3BP assembles stress granules. J. Cell Biol. 2003, 160, 823–831. [Google Scholar] [CrossRef]
- Bley, N.; Lederer, M.; Pfalz, B.; Reinke, C.; Fuchs, T.; Glaß, M.; Möller, B.; Hüttelmaier, S. Stress granules are dispensable for mRNA stabilization during cellular stress. Nucleic Acids Res. 2015, 43, e26. [Google Scholar] [CrossRef]
- Alberti, S.; Mateju, D.; Mediani, L.; Carra, S. Granulostasis: Protein Quality Control of RNP Granules. Front. Mol. Neurosci. 2017, 10, 84. [Google Scholar] [CrossRef] [PubMed]
- Ganassi, M.; Mateju, D.; Bigi, I.; Mediani, L.; Poser, I.; Lee, H.O.; Seguin, S.J.; Morelli, F.F.; Vinet, J.; Leo, G.; et al. A Surveillance Function of the HSPB8-BAG3-HSP70 Chaperone Complex Ensures Stress Granule Integrity and Dynamism. Mol. Cell 2016, 63, 796–810. [Google Scholar] [CrossRef] [PubMed]
- Cuesta, R.; Laroia, G.; Schneider, R.J. Chaperone hsp27 inhibits translation during heat shock by binding eIF4G and facilitating dissociation of cap-initiation complexes. Genes Dev. 2000, 14, 1460–1470. [Google Scholar] [PubMed]
- Kedersha, N.L.; Gupta, M.; Li, W.; Miller, I.; Anderson, P. RNA-Binding Proteins TIA-1 and TIAR Link the Phosphorylation of eIF-2α to the Assembly of Mammalian Stress Granules. J. Cell Biol. 1999, 147, 1431–1442. [Google Scholar] [CrossRef] [PubMed]
- Jain, S.; Wheeler, J.R.; Walters, R.W.; Agrawal, A.; Barsic, A.; Parker, R. ATPase modulated stress granules contain a diverse proteome and substructure. Cell 2016, 164, 487–498. [Google Scholar] [CrossRef] [PubMed]
- Vertii, A.; Hakim, C.; Kotlyarov, A.; Gaestel, M. Analysis of properties of small heat shock protein Hsp25 in MAPK-activated protein kinase 2 (MK2)-deficient cells: MK2-dependent insolubilization of Hsp25 oligomers correlates with susceptibility to stress. J. Biol. Chem. 2006, 281, 26966–26975. [Google Scholar] [CrossRef]
- Bengoechea, R.; Pittman, S.K.; Tuck, E.P.; True, H.L.; Weihl, C.C. Myofibrillar disruption and RNA-binding protein aggregation in a mouse model of limb-girdle muscular dystrophy 1D. Hum. Mol. Genet. 2015, 24, 6588–6602. [Google Scholar] [CrossRef]
- Rajgor, D.; Shanahan, C.M. RNA granules and cytoskeletal links. Biochem. Soc. Trans. 2014, 42, 1206–1210. [Google Scholar] [CrossRef]
- Buchan, J.R. mRNP granules. Assembly, function, and connections with disease. RNA Biol. 2014, 11, 1019–1030. [Google Scholar] [CrossRef] [PubMed]
- Moser, J.J.; Fritzler, M.J. Relationship of other cytoplasmic ribonucleoprotein bodies (cRNPB) to GW/P bodies. Adv. Exp. Med. Biol. 2013, 768, 213–242. [Google Scholar] [PubMed]
- Gallouzi, I.E.; Brennan, C.M.; Stenberg, M.G.; Swanson, M.S.; Eversole, A.; Maizels, N.; Steitz, J.A. HuR binding to cytoplasmic mRNA is perturbed by heat shock. Proc. Natl. Acad. Sci. USA 2000, 97, 3073–3078. [Google Scholar] [CrossRef] [PubMed]
- Aizer, A.; Brody, Y.; Ler, L.W.; Sonenberg, N.; Singer, R.H.; Shav-Tal, Y. The Dynamics of Mammalian P Body Transport, Assembly, and Disassembly In Vivo. Mol. Biol. Cell 2008, 19, 4154–4166. [Google Scholar] [CrossRef] [PubMed]
- Gyurko, D.M.; Soti, C.; Stetak, A.; Csermely, P. System level mechanisms of adaptation, learning, memory formation and evolvability: The role of chaperone and other networks. Curr. Protein Pept. Sci. 2014, 15, 171–188. [Google Scholar] [CrossRef] [PubMed]
- Mahboubi, H.; Seganathy, E.; Kong, D.; Stochaj, U. Identification of Novel Stress Granule Components That Are Involved in Nuclear Transport. PLoS ONE 2013, 8, e68356. [Google Scholar] [CrossRef]
- Mahboubi, H.; Kodiha, M.; Stochaj, U. Automated detection and quantification of granular cell compartments. Microsc. Microanal. 2013, 19, 617–628. [Google Scholar] [CrossRef]
- Crampton, N.; Kodiha, M.; Shrivastava, S.; Umar, R.; Stochaj, U. Oxidative stress inhibits nuclear protein export by multiple mechanisms that target FG nucleoporins and Crm1. Mol. Biol. Cell 2009, 20, 5106–5116. [Google Scholar] [CrossRef]
- Shevchenko, A.; Wilm, M.; Vorm, O.; Mann, M. Mass spectrometric sequencing of proteins silver-stained polyacrylamide gels. Anal. Chem. 1996, 68, 850–858. [Google Scholar] [CrossRef]
- Kodiha, M.; Tran, D.; Qian, C.; Morogan, A.; Presley, J.F.; Brown, C.M.; Stochaj, U. Oxidative stress mislocalizes and retains transport factor importin-alpha and nucleoporins Nup153 and Nup88 in nuclei where they generate high molecular mass complexes. Biochim. Biophys. Acta 2008, 1783, 405–418. [Google Scholar] [CrossRef]
- Kodiha, M.; Banski, P.; Stochaj, U. Interplay between MEK and PI3 kinase signaling regulates the subcellular localization of protein kinases ERK1/2 and Akt upon oxidative stress. FEBS Lett. 2009, 583, 1987–1993. [Google Scholar] [CrossRef] [PubMed]
- Mahboubi, H.; Barisé, R.; Stochaj, U. 5′-AMP-activated protein kinase alpha regulates stress granule biogenesis. Biochim. Biophys. Acta 2015, 1853, 1725–1737. [Google Scholar] [CrossRef] [PubMed]
- Anderson, P.; Kedersha, N. Stress granules: The Tao of RNA triage. Trends Biochem. Sci 2008, 33, 141–150. [Google Scholar] [CrossRef] [PubMed]
- Kedersha, N.; Stoecklin, G.; Ayodele, M.; Yacono, P.; Lykke-Andersen, J.; Fritzler, M.J.; Scheuner, D.; Kaufman, R.J.; Golan, D.E.; Anderson, P. Stress granules and processing bodies are dynamically linked sites of mRNP remodeling. J. Cell Biol. 2005, 169, 871–884. [Google Scholar] [CrossRef]
- Yamamoto, Y.; Izawa, S. Adaptive response in stress granule formation and bulk translational repression upon a combined stress of mild heat shock and mild ethanol stress in yeast. Genes Cells 2013, 18, 974–984. [Google Scholar] [CrossRef]
- Mateju, D.; Franzmann, T.M.; Patel, A.; Kopach, A.; Boczek, E.E.; Maharana, S.; Lee, H.O.; Carra, S.; Hyman, A.A.; Alberti, S. An aberrant phase transition of stress granules triggered by misfolded protein and prevented by chaperone function. The EMBO Journal 2017, 36, 1669–1687. [Google Scholar] [CrossRef]
- Ciuzan, O.; Hancock, J.; Pamfil, D.; Wilson, I.; Ladomery, M. The evolutionarily conserved multifunctional glycine-rich RNA-binding proteins play key roles in development and stress adaptation. Physiol. Plant. 2015, 153, 1–11. [Google Scholar] [CrossRef]
- Pinol-Roma, S.; Choi, Y.D.; Matunis, M.J.; Dreyfuss, G. Immunopurification of heterogeneous nuclear ribonucleoprotein particles reveals an assortment of RNA-binding proteins. Genes Dev. 1988, 2, 215–227. [Google Scholar] [CrossRef]
- Nazarov, I.B.; Bakhmet, E.I.; Tomilin, A.N. KH-Domain Poly(C)-Binding Proteins as Versatile Regulators of Multiple Biological Processes. Biochemistry (Mosc.) 2019, 84, 205–219. [Google Scholar] [CrossRef]
- Niforou, K.; Cheimonidou, C.; Trougakos, I.P. Molecular chaperones and proteostasis regulation during redox imbalance. Redox Biol 2014, 2, 323–332. [Google Scholar] [CrossRef]
- McLellan, C.A.; Raynes, D.A.; Guerriero, V. HspBP1, an Hsp70 cochaperone, has two structural domains and is capable of altering the conformation of the Hsp70 ATPase domain. J. Biol. Chem. 2003, 278, 19017–19022. [Google Scholar] [CrossRef]
- Walters, R.W.; Muhlrad, D.; Garcia, J.; Parker, R. Differential effects of Ydj1 and Sis1 on Hsp70-mediated clearance of stress granules in Saccharomyces cerevisiae. RNA 2015, 21, 1669–1671. [Google Scholar] [CrossRef]
- Mazroui, R.; Di Marco, S.; Kaufman, R.J.; Gallouzi, I.E. Inhibition of the ubiquitin-proteasome system induces stress granule formation. Mol. Biol. Cell 2007, 18, 2603–2618. [Google Scholar] [CrossRef]
- Shrestha, L.; Young, J.C. Function and Chemotypes of Human Hsp70 Chaperones. Curr. Top. Med. Chem. 2016, 16, 2812–2828. [Google Scholar] [CrossRef]
- Elbaum-Garfinkle, S.; Kim, Y.; Szczepaniak, K.; Chen, C.C.; Eckmann, C.R.; Myong, S.; Brangwynne, C.P. The disordered P granule protein LAF-1 drives phase separation into droplets with tunable viscosity and dynamics. Proc. Natl. Acad. Sci. USA 2015, 112, 7189–7194. [Google Scholar] [CrossRef]
- Sheu-Gruttadauria, J.; MacRae, I.J. Phase Transitions in the Assembly and Function of Human miRISC. Cell 2018, 173, 946–957. [Google Scholar] [CrossRef]
- Celetti, G.; Paci, G.; Caria, J.; VanDelinder, V.; Bachand, G.; Lemke, E.A. The liquid state of FG-nucleoporins mimics permeability barrier properties of nuclear pore complexes. J. Cell Biol. 2020, 219, pii: e201907157. [Google Scholar] [CrossRef]
- Bounedjah, O.; Desforges, B.; Wu, T.D.; Pioche-Durieu, C.; Marco, S.; Hamon, L.; Curmi, P.A.; Guerquin-Kern, J.L.; Pietrement, O.; Pastre, D. Free mRNA in excess upon polysome dissociation is a scaffold for protein multimerization to form stress granules. Nucleic Acids Res. 2014, 42, 8678–8691. [Google Scholar] [CrossRef]
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Mahboubi, H.; Moujaber, O.; Kodiha, M.; Stochaj, U. The Co-Chaperone HspBP1 Is a Novel Component of Stress Granules that Regulates Their Formation. Cells 2020, 9, 825. https://doi.org/10.3390/cells9040825
Mahboubi H, Moujaber O, Kodiha M, Stochaj U. The Co-Chaperone HspBP1 Is a Novel Component of Stress Granules that Regulates Their Formation. Cells. 2020; 9(4):825. https://doi.org/10.3390/cells9040825
Chicago/Turabian StyleMahboubi, Hicham, Ossama Moujaber, Mohamed Kodiha, and Ursula Stochaj. 2020. "The Co-Chaperone HspBP1 Is a Novel Component of Stress Granules that Regulates Their Formation" Cells 9, no. 4: 825. https://doi.org/10.3390/cells9040825
APA StyleMahboubi, H., Moujaber, O., Kodiha, M., & Stochaj, U. (2020). The Co-Chaperone HspBP1 Is a Novel Component of Stress Granules that Regulates Their Formation. Cells, 9(4), 825. https://doi.org/10.3390/cells9040825