Functional Diversification and Specialization of Cytosolic 70-kDa Heat Shock Proteins

A fundamental question in molecular evolution is how protein functional differentiation alters the ability of cells and organisms to cope with stress and survive. To answer this question we used two paralogous Hsp70s from mouse and explored whether these highly similar cytosolic molecular chaperones, which apart their temporal expression have been considered functionally interchangeable, are differentiated with respect to their lipid-binding function. We demonstrate that the two proteins bind to diverse lipids with different affinities and therefore are functionally specialized. The observed lipid-binding patterns may be related with the ability of both Hsp70s to induce cell death by binding to a particular plasma-membrane lipid, and the potential of only one of them to promote cell survival by binding to a specific lysosomal-membrane lipid. These observations reveal that two seemingly identical proteins differentially modulate cellular adaptation and survival by having acquired specialized functions via sequence divergence. Therefore, this study provides an evolutionary paradigm, where promiscuity, specificity, sub- and neo-functionalization orchestrate one of the most conserved systems in nature, the cellular stress-response.

[1]  D. Kültz,et al.  Molecular and evolutionary basis of the cellular stress response. , 2005, Annual review of physiology.

[2]  M. Febbraio,et al.  Mechanisms of stress-induced cellular HSP72 release: implications for exercise-induced increases in extracellular HSP72. , 2005, Exercise immunology review.

[3]  Liisa Holm,et al.  DaliLite workbench for protein structure comparison , 2000, Bioinform..

[4]  Radhey S. Gupta,et al.  Phylogenetic analysis of 70 kD heat shock protein sequences suggests a chimeric origin for the eukaryotic cell nucleus , 1994, Current Biology.

[5]  P. Cosson,et al.  Separation and Characterization of Late Endosomal Membrane Domains* , 2002, The Journal of Biological Chemistry.

[6]  L. Brocchieri,et al.  hsp70 genes in the human genome: Conservation and differentiation patterns predict a wide array of overlapping and specialized functions , 2008, BMC Evolutionary Biology.

[7]  I. Horváth,et al.  Membrane-associated stress proteins: more than simply chaperones. , 2008, Biochimica et biophysica acta.

[8]  M. Molls,et al.  Binding of heat shock protein 70 to extracellular phosphatidylserine promotes killing of normoxic and hypoxic tumor cells , 2009, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[9]  Mark Johnson,et al.  NCBI BLAST: a better web interface , 2008, Nucleic Acids Res..

[10]  C. Lingwood,et al.  The ATPase Domain of hsp70 Possesses a Unique Binding Specificity for 3′-Sulfogalactolipids* , 2001, The Journal of Biological Chemistry.

[11]  G. van Meer,et al.  Lipid map of the mammalian cell , 2011, Journal of Cell Science.

[12]  M. Nei,et al.  MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. , 2011, Molecular biology and evolution.

[13]  R. Morimoto,et al.  The human cytosolic molecular chaperones hsp90, hsp70 (hsc70) and hdj‐1 have distinct roles in recognition of a non‐native protein and protein refolding. , 1996, The EMBO journal.

[14]  Thierry Ferreira,et al.  Lipid‐Induced ER Stress: Synergistic Effects of Sterols and Saturated Fatty Acids , 2009, Traffic.

[15]  D. Moreira,et al.  The early evolution of lipid membranes and the three domains of life , 2012, Nature Reviews Microbiology.

[16]  S. Antonarakis,et al.  Gene duplication: a drive for phenotypic diversity and cause of human disease. , 2007, Annual review of genomics and human genetics.

[17]  Brian D. Slaughter,et al.  Non-uniform membrane diffusion enables steady-state cell polarization via vesicular trafficking , 2013, Nature Communications.

[18]  L. Hightower,et al.  Purification and initial characterization of the 71-kilodalton rat heat-shock protein and its cognate as fatty acid binding proteins. , 1986, Biochemistry.

[19]  S. Pizzo,et al.  Binding of Activated α2-Macroglobulin to Its Cell Surface Receptor GRP78 in 1-LN Prostate Cancer Cells Regulates PAK-2-dependent Activation of LIMK* , 2005, Journal of Biological Chemistry.

[20]  P. Srivastava,et al.  Heat‐Shock Proteins , 2003, Current protocols in immunology.

[21]  Clinton H Joiner,et al.  Phosphatidylserine externalization in sickle red blood cells: associations with cell age, density, and hemoglobin F. , 2003, Blood.

[22]  A. Maio Extracellular heat shock proteins, cellular export vesicles, and the Stress Observation System: A form of communication during injury, infection, and cell damage , 2010, Cell Stress and Chaperones.

[23]  D. Sheff,et al.  Intracellular phosphatidylserine is essential for retrograde membrane traffic through endosomes , 2011, Proceedings of the National Academy of Sciences.

[24]  K. Katoh,et al.  MAFFT Multiple Sequence Alignment Software Version 7: Improvements in Performance and Usability , 2013, Molecular biology and evolution.

[25]  Kenneth H. Wolfe,et al.  Turning a hobby into a job: How duplicated genes find new functions , 2008, Nature Reviews Genetics.

[26]  S. Dowler,et al.  Protein Lipid Overlay Assay , 2002, Science's STKE.

[27]  Sunhwan Jo,et al.  PBEQ-Solver for online visualization of electrostatic potential of biomolecules , 2008, Nucleic Acids Res..

[28]  N. Arispe,et al.  Lipid interaction differentiates the constitutive and stress-induced heat shock proteins Hsc70 and Hsp70 , 2002, Cell stress & chaperones.

[29]  M. Lemmon,et al.  Pleckstrin homology (PH) domains and phosphoinositides. , 2007, Biochemical Society symposium.

[30]  Douglas M. Cry,et al.  Cooperation of the molecular chaperone Ydj1 with specific Hsp70 homologs to suppress protein aggregation , 1995 .

[31]  D. Pain,et al.  Mt-Hsp70 Homolog, Ssc2p, Required for Maturation of Yeast Frataxin and Mitochondrial Iron Homeostasis* , 1998, The Journal of Biological Chemistry.

[32]  John Eric Wilson Isozymes of mammalian hexokinase: structure, subcellular localization and metabolic function , 2003, Journal of Experimental Biology.

[33]  Huiping Zhou,et al.  ER Stress and Lipid Metabolism in Adipocytes , 2012, Biochemistry research international.

[34]  T. Haines A new look at Cardiolipin. , 2009, Biochimica et biophysica acta.

[35]  A. Finazzi-Agro’,et al.  Molecular identification of albumin and Hsp70 as cytosolic anandamide-binding proteins. , 2009, Chemistry & biology.

[36]  D. Kültz,et al.  Evolution of the cellular stress proteome: from monophyletic origin to ubiquitous function , 2003, Journal of Experimental Biology.

[37]  G. Balogh,et al.  Lipidomics reveals membrane lipid remodelling and release of potential lipid mediators during early stress responses in a murine melanoma cell line. , 2010, Biochimica et biophysica acta.

[38]  Eoin Fahy,et al.  Subcellular organelle lipidomics in TLR-4-activated macrophages1[S] , 2010, Journal of Lipid Research.

[39]  E. Craig,et al.  Major heat shock gene of Drosophila and the Escherichia coli heat-inducible dnaK gene are homologous. , 1984, Proceedings of the National Academy of Sciences of the United States of America.

[40]  E. Craig,et al.  Functional Specificity Among Hsp70 Molecular Chaperones , 1997, Science.

[41]  M. Jäättelä,et al.  Connecting Hsp70, sphingolipid metabolism and lysosomal stability , 2010, Cell cycle.

[42]  D. Murray,et al.  Binding of phosphoinositide-specific phospholipase C-zeta (PLC-zeta) to phospholipid membranes: potential role of an unstructured cluster of basic residues. , 2007, The Journal of biological chemistry.

[43]  M. Kabani,et al.  Multiple Hsp70 Isoforms in the Eukaryotic Cytosol: Mere Redundancy or Functional Specificity? , 2008, Current genomics.

[44]  L. Santambrogio,et al.  Microautophagy of cytosolic proteins by late endosomes. , 2011, Developmental cell.

[45]  T. Berendonk,et al.  Convergent evolution of heat-inducibility during subfunctionalization of the Hsp70 gene family , 2013, BMC Evolutionary Biology.

[46]  E. Dainese,et al.  Intracellular trafficking of anandamide: new concepts for signaling. , 2010, Trends in biochemical sciences.

[47]  C. Steinem,et al.  Hsp70 Translocates into the Plasma Membrane after Stress and Is Released into the Extracellular Environment in a Membrane-Associated Form that Activates Macrophages1 , 2008, The Journal of Immunology.

[48]  P. Gray,et al.  Extracellular Heat Shock Proteins in Cell Signaling and Immunity , 2007, Annals of the New York Academy of Sciences.

[49]  G. Schmitz,et al.  Tumor-Specific Hsp70 Plasma Membrane Localization Is Enabled by the Glycosphingolipid Gb3 , 2008, PloS one.

[50]  J. Hopwood,et al.  Lipid composition of microdomains is altered in a cell model of Gaucher disease** This work was supported by a National Health and Medical Research Council project grant in Australia. Published, JLR Papers in Press, April 21, 2008. , 2008, Journal of Lipid Research.

[51]  Robert H. Michell,et al.  Inositol derivatives: evolution and functions , 2008, Nature Reviews Molecular Cell Biology.

[52]  C. Bucana,et al.  Differentiation-dependent expression of phosphatidylserine in mammalian plasma membranes: quantitative assessment of outer-leaflet lipid by prothrombinase complex formation. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[53]  M. Lemmon,et al.  Determining selectivity of phosphoinositide-binding domains. , 2006, Methods.

[54]  M. Stevenson,et al.  Mechanisms for Hsp70 secretion: crossing membranes without a leader. , 2007, Methods.

[55]  H. Kampinga,et al.  The HSP70 chaperone machinery: J proteins as drivers of functional specificity , 2010, Nature Reviews Molecular Cell Biology.

[56]  A. Zylicz,et al.  Hsp70 stabilizes lysosomes and reverts Niemann–Pick disease-associated lysosomal pathology , 2010, Nature.

[57]  L. Hein,et al.  Selective reduction of bis(monoacylglycero)phosphate ameliorates the storage burden in a THP-1 macrophage model of Gaucher disease[S] , 2013, Journal of Lipid Research.

[58]  N. Nikolaidis,et al.  Identification of Several Cytoplasmic HSP70 Genes from the Mediterranean Mussel (Mytilus galloprovincialis) and Their Long-Term Evolution in Mollusca and Metazoa , 2006, Journal of Molecular Evolution.

[59]  David E. Misek,et al.  Global Profiling of the Cell Surface Proteome of Cancer Cells Uncovers an Abundance of Proteins with Chaperone Function* , 2003, The Journal of Biological Chemistry.

[60]  W. Arap,et al.  Cell surface expression of the stress response chaperone GRP78 enables tumor targeting by circulating ligands. , 2004, Cancer cell.

[61]  F. Boas,et al.  Phosphatidylserine exposure and red cell viability in red cell aging and in hemolytic anemia. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[62]  R. Stahelin Lipid binding domains: more than simple lipid effectors This research was supported by grants from the American Heart Association (0735350N), the American Cancer Society (IRG-84-002-22), and the Indiana University School of Medicine. Published, JLR Papers in Press, November 13, 2008. , 2009, Journal of Lipid Research.

[63]  A. Herr,et al.  Shiga Toxin Binding to Glycolipids and Glycans , 2012, PloS one.

[64]  C. Gross,et al.  Heat shock protein 70 surface-positive tumor exosomes stimulate migratory and cytolytic activity of natural killer cells. , 2005, Cancer research.

[65]  Irina Zaitseva,et al.  Binding of Phosphoinositide-specific Phospholipase C-ζ (PLC-ζ) to Phospholipid Membranes , 2007, Journal of Biological Chemistry.

[66]  D. Irwin,et al.  Evolution of glucose utilization: glucokinase and glucokinase regulator protein. , 2014, Molecular phylogenetics and evolution.

[67]  J. Raulston,et al.  Hsp70s contain a specific sulfogalactolipid binding site. Differential aglycone influence on sulfogalactosyl ceramide binding by recombinant prokaryotic and eukaryotic hsp70 family members. , 2001, Biochemistry.

[68]  G. Meer,et al.  Membrane lipids: where they are and how they behave , 2008, Nature Reviews Molecular Cell Biology.

[69]  Jianzhi Zhang,et al.  Rapid Subfunctionalization Accompanied by Prolonged and Substantial Neofunctionalization in Duplicate Gene Evolution , 2005, Genetics.

[70]  K. Katoh,et al.  Improvements in Performance and Usability , 2013 .

[71]  L. Hightower,et al.  The 73 kilodalton heat shock cognate protein purified from rat brain contains nonesterified palmitic and stearic acids , 1986, Journal of cellular physiology.

[72]  D. Spitz,et al.  Aging augments mitochondrial susceptibility to heat stress. , 2009, American journal of physiology. Regulatory, integrative and comparative physiology.

[73]  A. De Maio Extracellular Hsp70: export and function. , 2014, Current protein & peptide science.

[74]  G. Multhoff Heat shock proteins in immunity. , 2006, Handbook of experimental pharmacology.

[75]  A. Force,et al.  The probability of duplicate gene preservation by subfunctionalization. , 2000, Genetics.

[76]  A. Maio,et al.  Hsc70 and Hsp70 interact with phosphatidylserine on the surface of PC12 cells resulting in a decrease of viability , 2004, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[77]  Masatoshi Nei,et al.  Concerted and nonconcerted evolution of the Hsp70 gene superfamily in two sibling species of nematodes. , 2004, Molecular biology and evolution.

[78]  E. Craig,et al.  Divergent functional properties of the ribosome-associated molecular chaperone Ssb compared with other Hsp70s. , 2001, Molecular biology of the cell.

[79]  D. Cyr Cooperation of the molecular chaperone Ydj1 with specific Hsp70 homologs to suppress protein aggregation. , 1995, FEBS letters.

[80]  C. Newgard,et al.  Differential Effects of Overexpressed Glucokinase and Hexokinase I in Isolated Islets , 1996, The Journal of Biological Chemistry.

[81]  P. Greimel,et al.  Lipid compartmentalization in the endosome system. , 2014, Seminars in cell & developmental biology.