A specific heat shock protein enhances the expression of mammalian olfactory receptor proteins.

Multiple trials failed to express significant amounts of olfactory receptors in heterologous cells as they are typically retained in the endoplasmic reticulum (ER). Evidence is accumulating that cell-type-specific accessory proteins regulate the folding of olfactory receptors, their exit from the ER, and the trafficking to the plasma membrane of the olfactory cilia where the receptors gain access to odorants. We found Hsc70t, a testis-enriched variant of the Hsp70 family of heat shock proteins which is specifically expressed in post-meiotic germ cells, in the olfactory epithelium of mouse and human. Cotransfected HEK293 cells with Hsc70t and different green fluorescent protein-tagged odorant receptors (ORs) from mouse and man showed a significantly enhanced OR expression. Hsc70t expression also changed the amount of cells functionally expressing olfactory receptors at the cell surface as the number of cells responding to odorants in Ca2+-imaging experiments significantly increased. Our results show that Hsc70t helps expression of ORs in heterologous cell systems and helped the characterization of an "orphan" human olfactory receptor.

[1]  W. Chirico.  Dissociation of complexes between 70 kDa stress proteins and presecretory proteins is facilitated by a cytosolic factor. , 1992, Biochemical and biophysical research communications.

[2]  Harumi Saito,et al.  RTP Family Members Induce Functional Expression of Mammalian Odorant Receptors , 2004, Cell.

[3]  H. Sakano,et al.  Functional identification and reconstitution of an odorant receptor in single olfactory neurons. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[4]  F. Echeverri,et al.  The human olfactory receptor repertoire , 2001, Genome Biology.

[5]  Ari Helenius,et al.  Quality control in the endoplasmic reticulum , 2003, Nature Reviews Molecular Cell Biology.

[6]  R. Morimoto,et al.  Small subclass of rat olfactory neurons with specific bulbar projections is reactive with monoclonal antibodies to the HSP70 heat shock protein , 1994, The Journal of comparative neurology.

[7]  S. Bröer,et al.  Xenopus laevis Oocytes. , 2003, Methods in molecular biology.

[8]  W. Welch,et al.  Involvement of 70-kD heat-shock proteins in peroxisomal import , 1994, The Journal of cell biology.

[9]  Melanie G. Lee,et al.  RAMPs regulate the transport and ligand specificity of the calcitonin-receptor-like receptor , 1998, Nature.

[10]  P. Vanderhaeghen,et al.  Olfactory receptors are displayed on dog mature sperm cells , 1993, The Journal of cell biology.

[11]  D. Sparks,et al.  Human Olfactory Receptor Neurons Express Heat Shock Protein 70: Age-Related Trends , 1995, The Annals of otology, rhinology, and laryngology.

[12]  R. Axel,et al.  Odorant Receptors on Axon Termini in the Brain , 2004, Science.

[13]  Cornelia I Bargmann,et al.  Odorant Receptor Localization to Olfactory Cilia Is Mediated by ODR-4, a Novel Membrane-Associated Protein , 1998, Cell.

[14]  M. Matsumoto,et al.  Haploid specific activations of protamine 1 and hsc70t genes in mouse spermatogenesis. , 1993, Biochimica et biophysica acta.

[15]  H. Breer,et al.  Olfactory Receptor Proteins in Axonal Processes of Chemosensory Neurons , 2004, The Journal of Neuroscience.

[16]  Patrice Mollard,et al.  A single olfactory receptor specifically binds a set of odorant molecules , 2002, The European journal of neuroscience.

[17]  Jason C. Young,et al.  Pathways of chaperone-mediated protein folding in the cytosol , 2004, Nature Reviews Molecular Cell Biology.

[18]  M. Lerner,et al.  Functional expression of olfactory-adrenergic receptor chimeras and intracellular retention of heterologously expressed olfactory receptors. , 1997, Brain research. Molecular brain research.

[19]  R. Schekman,et al.  The role of stress proteins in membrane biogenesis. , 1988, Trends in biochemical sciences.

[20]  G. Gisselmann,et al.  Odorant receptor heterodimerization in the olfactory system of Drosophila melanogaster , 2005, Nature Neuroscience.

[21]  U. Klein,et al.  A Novel Interaction between Adrenergic Receptors and the α-Subunit of Eukaryotic Initiation Factor 2B* , 1997, The Journal of Biological Chemistry.

[22]  Peter Mombaerts,et al.  Odorant Receptor Expression Defines Functional Units in the Mouse Olfactory System , 2002, The Journal of Neuroscience.

[23]  C. J. Reed,et al.  Heat shock protein 70 in the rat nasal cavity: localisation and response to hyperthermia , 2004, Archives of Toxicology.

[24]  M. Okabe,et al.  Functional characterization of a mouse testicular olfactory receptor and its role in chemosensing and in regulation of sperm motility , 2004, Journal of Cell Science.

[25]  N. Tsunekawa,et al.  The Hsp70 homolog gene, Hsc70t, is expressed under translational control during mouse spermiogenesis , 1999, Molecular reproduction and development.

[26]  R. Kaufman,et al.  The mammalian unfolded protein response. , 2003, Annual review of biochemistry.

[27]  R. Morimoto,et al.  Odorants as cell‐type specific activators of a heat shock response in the rat olfactory mucosa , 2001, The Journal of comparative neurology.

[28]  M. Mayer,et al.  Hsp70 chaperones: Cellular functions and molecular mechanism , 2005, Cellular and Molecular Life Sciences.

[29]  M. Spehr,et al.  Particulate Adenylate Cyclase Plays a Key Role in Human Sperm Olfactory Receptor-mediated Chemotaxis* , 2004, Journal of Biological Chemistry.

[30]  Stuart Firestein,et al.  A pharmacological profile of the aldehyde receptor repertoire in rat olfactory epithelium , 2004, The Journal of physiology.

[31]  R. Morimoto,et al.  Development and further characterization of a small subclass of rat olfactory receptor neurons that shows immunoreactivity for the HSP70 heat shock protein , 1999, The Journal of comparative neurology.

[32]  Nicholas J. Hoogenraad,et al.  Molecular Chaperones Hsp90 and Hsp70 Deliver Preproteins to the Mitochondrial Import Receptor Tom70 , 2003, Cell.

[33]  A. Gimelbrant,et al.  Truncation Releases Olfactory Receptors from the Endoplasmic Reticulum of Heterologous Cells , 1999, Journal of neurochemistry.

[34]  L. Walter,et al.  Comparative analysis of the three major histocompatibility complex-linked heat shock protein 70 (Hsp70) genes of the rat , 2005, Immunogenetics.

[35]  Y. Saijoh,et al.  Genomic structure of the spermatid-specific hsp70 homolog gene located in the class III region of the major histocompatibility complex of mouse and man. , 1998, Journal of biochemistry.

[36]  Elizabeth A. Craig,et al.  A subfamily of stress proteins facilitates translocation of secretory and mitochondrial precursor polypeptides , 1988, Nature.

[37]  Wulf Paschen,et al.  Shutdown of Translation: Lethal or Protective? Unfolded Protein Response versus Apoptosis , 2003, Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism.

[38]  Timothy S McClintock,et al.  Trafficking prerogatives of olfactory receptors. , 2003, Neuroreport.

[39]  S. Firestein,et al.  The olfactory receptor gene superfamily of the mouse , 2002, Nature Neuroscience.

[40]  L. Pott,et al.  Functional expression of odorant receptors of the zebrafish Danio rerio and of the nematode C. elegans in HEK293 cells. , 1997, Chemical senses.

[41]  Cori Bargmann,et al.  Polarized Dendritic Transport and the AP-1 μ1 Clathrin Adaptor UNC-101 Localize Odorant Receptors to Olfactory Cilia , 2001, Neuron.

[42]  G Vassart,et al.  Specific repertoire of olfactory receptor genes in the male germ cells of several mammalian species. , 1997, Genomics.

[43]  F. Urano,et al.  Transcriptional and translational control in the Mammalian unfolded protein response. , 2002, Annual review of cell and developmental biology.

[44]  K. Mikoshiba,et al.  Functional expression of a mammalian odorant receptor. , 1998, Science.

[45]  W. Pak,et al.  Cyclophilin-related protein RanBP2 acts as chaperone for red/green opsin , 1996, Nature.

[46]  F. Echeverri,et al.  Endoplasmic Reticulum Retention, Degradation, and Aggregation of Olfactory G‐Protein Coupled Receptors , 2003, Traffic.

[47]  Hanns Hatt,et al.  Specificity and Sensitivity of a Human Olfactory Receptor Functionally Expressed in Human Embryonic Kidney 293 Cells andXenopus Laevis Oocytes , 1999, The Journal of Neuroscience.

[48]  Jeffrey A. Riffell,et al.  Identification of a Testicular Odorant Receptor Mediating Human Sperm Chemotaxis , 2003, Science.

[49]  C. J. Reed,et al.  Induction of heat shock protein 70 in rat olfactory epithelium by toxic chemicals: in vitro and in vivo studies , 2005, Archives of Toxicology.

[50]  Christopher M Dobson,et al.  Principles of protein folding, misfolding and aggregation. , 2004, Seminars in cell & developmental biology.

[51]  C. Milner,et al.  Structure and expression of the three MHC-linked HSP70 genes , 2004, Immunogenetics.

[52]  Dietmar Krautwurst,et al.  Identification of Ligands for Olfactory Receptors by Functional Expression of a Receptor Library , 1998, Cell.

[53]  A. Gimelbrant,et al.  A nuclear matrix attachment region is highly homologous to a conserved domain of olfactory receptors , 1997, Journal of Molecular Neuroscience.

[54]  M. Matsumoto,et al.  Cloning of a hsp70-related gene expressed in mouse spermatids. , 1990, Biochemical and biophysical research communications.

[55]  Ivan Rodriguez,et al.  Axon Guidance of Mouse Olfactory Sensory Neurons by Odorant Receptors and the β2 Adrenergic Receptor , 2004, Cell.

[56]  Hiroshi Kataoka,et al.  Molecular Bases of Odor Discrimination: Reconstitution of Olfactory Receptors that Recognize Overlapping Sets of Odorants , 2001, The Journal of Neuroscience.