SemOpenAlex |

SemOpenAlex

Matches in SemOpenAlex for { <https://semopenalex.org/work/W2042871707> ?p ?o ?g. }

Showing items 1 to 100 of ±128 with 100 items per page.

W2042871707 endingPage "20212" @default.
W2042871707 startingPage "20206" @default.
W2042871707 abstract "Human lipocalin-1 (Lcn-1, also called tear lipocalin), a member of the lipocalin structural superfamily, is produced by a number of glands and tissues and is known to bind an unusually large array of hydrophobic ligands. Apart from its specific function in stabilizing the lipid film of human tear fluid, it is suggested to act as a physiological scavenger of potentially harmful lipophilic compounds, in general. To characterize proteins involved in the reception, detoxification, or degradation of these ligands, a cDNA phage-display library from human pituitary gland was constructed and screened for proteins interacting with Lcn-1. Using this method an Lcn-1 interacting phage was isolated that expressed a novel human protein. Molecular cloning and analysis of the entire cDNA indicated that it encodes a 55-kDa protein, lipocalin-1 interacting membrane receptor (LIMR), with nine putative transmembrane domains. The cell membrane location of this protein was confirmed by immunocytochemistry and Western blot analysis of membrane fractions of human NT2 cells. Independent biochemical investigations using a recombinant N-terminal fragment of LIMR also demonstrated a specific interaction with Lcn-1 in vitro. Based on these data, we suggest LIMR to be a receptor of Lcn-1 ligands. These findings constitute the first report of cloning of a lipocalin interacting, plasma membrane-located receptor, in general. In addition, a sequence comparison supports the biological relevance of this novel membrane protein, because genes with significant nucleotide sequence similarity are present in Takifugu rubripes, Drosophila melanogaster, Caenorhabditis elegans, Mus musculus, Bos taurus, and Sus scrofa. According to data derived from the human genome sequencing project, the LIMR-encoding gene has to be mapped on human chromosome 12, and its intron/exon organization could be established. The entire LIMR-encoding gene consists of about 13.7 kilobases in length and contains 16 introns with a length between 91 and 3438 base pairs. Human lipocalin-1 (Lcn-1, also called tear lipocalin), a member of the lipocalin structural superfamily, is produced by a number of glands and tissues and is known to bind an unusually large array of hydrophobic ligands. Apart from its specific function in stabilizing the lipid film of human tear fluid, it is suggested to act as a physiological scavenger of potentially harmful lipophilic compounds, in general. To characterize proteins involved in the reception, detoxification, or degradation of these ligands, a cDNA phage-display library from human pituitary gland was constructed and screened for proteins interacting with Lcn-1. Using this method an Lcn-1 interacting phage was isolated that expressed a novel human protein. Molecular cloning and analysis of the entire cDNA indicated that it encodes a 55-kDa protein, lipocalin-1 interacting membrane receptor (LIMR), with nine putative transmembrane domains. The cell membrane location of this protein was confirmed by immunocytochemistry and Western blot analysis of membrane fractions of human NT2 cells. Independent biochemical investigations using a recombinant N-terminal fragment of LIMR also demonstrated a specific interaction with Lcn-1 in vitro. Based on these data, we suggest LIMR to be a receptor of Lcn-1 ligands. These findings constitute the first report of cloning of a lipocalin interacting, plasma membrane-located receptor, in general. In addition, a sequence comparison supports the biological relevance of this novel membrane protein, because genes with significant nucleotide sequence similarity are present in Takifugu rubripes, Drosophila melanogaster, Caenorhabditis elegans, Mus musculus, Bos taurus, and Sus scrofa. According to data derived from the human genome sequencing project, the LIMR-encoding gene has to be mapped on human chromosome 12, and its intron/exon organization could be established. The entire LIMR-encoding gene consists of about 13.7 kilobases in length and contains 16 introns with a length between 91 and 3438 base pairs. retinol-binding protein lipocalin-1 interacting membrane protein lipocalin-1 (also called tear lipocalin or von Ebner's gland protein) nickel-nitrilotriacetic acid N-methyl-d-asparate phosphate-buffered saline polymerase chain reaction base pair(s) Tris-buffered saline bovine serum albumin expressed sequence tag reverse transcription group of overlapping clones The protein superfamily of lipocalins consists of small, mainly secretory proteins defined on the basis of conserved amino acid sequence motifs and their common structure. Functionally they share several properties including the ability to bind/transport a remarkable array of small hydrophobic molecules, the formation of macromolecular complexes, and the binding to specific cell surface receptors (1Flower D.R. Biochem. J. 1996; 318: 1-14Crossref PubMed Scopus (1372) Google Scholar, 2Akerstrom B. Flower D.R. Salier J-P. Biochim. Biophys. Acta. 2000; 1482: 1-8Crossref PubMed Scopus (233) Google Scholar, 3Flower D.R. North A.C.T. Sansom C.E. Biochim. Biophys. Acta. 2000; 1482: 9-24Crossref PubMed Scopus (689) Google Scholar). Whereas a large number of various lipophilic ligands able to bind to lipocalins are known, only limited data are available concerning the identity of lipocalin receptors. There is clear evidence of a specific receptor for plasma retinol-binding protein (RBP)1 (4Bavik C.O. Busch C. Eriksson U. J. Biol. Chem. 1992; 267: 23035-23042Abstract Full Text PDF PubMed Google Scholar, 5Smeland S. Bjerknes T. Malaba L. Eskild W. Norum K.R. Blomhoff R. Biochem. J. 1995; 305: 419-424Crossref PubMed Scopus (58) Google Scholar, 6Sivaprasadarao A. Boudjelal M. Findlay J.B.C. Biochem. J. 1994; 302: 245-251Crossref PubMed Scopus (41) Google Scholar, 7Sundaram M. Sivaprasadarao A. DeSousa M.M. Findlay J.B.C. J. Biol. Chem. 1998; 273: 3336-3342Abstract Full Text Full Text PDF PubMed Scopus (100) Google Scholar) and more indirect evidence for receptors for α1-microglobulin (8Wester L. Michaelsson E. Holmdahl R. Olofsson T. Akerstrom B. Scand. J. Immunol. 1998; 48: 1-7Crossref PubMed Scopus (33) Google Scholar), major urinary protein (9Böcskei Z. Groom C.R. Flower D.R. Wright C.R. Phillips S.E. Cavaggioni A. Findlay J.B. North A.C. Nature. 1992; 360: 186-188Crossref PubMed Scopus (346) Google Scholar), β-lactoglobulin (10Papiz M.Z. Sawyer L. Eliopoulos E.E. North A.C. Findlay J.B. Sivaprasadarao R. Jones T.A. Newcomer M.E. Kraulis P.J. Nature. 1986; 324: 383-385Crossref PubMed Scopus (847) Google Scholar), olfactory-binding protein (11Boudjelal M. Sivaprasadarao A. Findlay B.B. Biochem. J. 1996; 317: 23-27Crossref PubMed Scopus (36) Google Scholar), α1-acid glycoprotein (12Andersen U.O. Bog-Hansen S. Kirkeby S. Acta Histochem. 1999; 101: 113-119Crossref PubMed Scopus (7) Google Scholar), and glycodelin (13Miller R.E. Fayen J.D. Chakraborty S. Weber M.C. Tykocinski M.L. FEBS Lett. 1998; 436: 455-460Crossref PubMed Scopus (34) Google Scholar), but with the exception of megalin, which seems to be an endocytic receptor for a variety of soluble macromolecules including several lipocalins, no specific lipocalin receptor has been fully characterized so far (14Flower D.R. Biochim. Biophys. Acta. 2000; 1482: 327-336Crossref PubMed Scopus (119) Google Scholar). This lack of knowledge is a major disadvantage in understanding the biological function of many lipocalin members.We have identified Lcn-1 (identical with tear lipocalin or von Ebner's gland protein) as a human member of the lipocalin superfamily (15Redl B. Holzfeind P. Lottspeich F. J. Biol. Chem. 1992; 267: 20282-20287Abstract Full Text PDF PubMed Google Scholar). It is produced by a number of secretory glands and tissues, including lacrimal and lingual salivary glands, prostate, and mucosal glands of the tracheobronchial tree, nasal mucosa, and sweat glands and by some neuroendocrine tissues (15Redl B. Holzfeind P. Lottspeich F. J. Biol. Chem. 1992; 267: 20282-20287Abstract Full Text PDF PubMed Google Scholar, 16Lassagne H. Gachon A.M. Exp. Eye Res. 1993; 55: 605-609Crossref Scopus (28) Google Scholar, 17Bläker M. Kock K. Ahlers C. Buck F. Schmale H. Biochim. Biophys. Acta. 1993; 1172: 131-137Crossref PubMed Scopus (77) Google Scholar, 18Holzfeind P. Merschak P. Rogatsch H. Culig Z. Feichtinger H. Klocker H. Redl B. FEBS Lett. 1996; 395: 95-98Crossref PubMed Scopus (39) Google Scholar, 19Redl B. Wojnar P. Ellemunter H. Feichtinger H. Lab. Invest. 1998; 78: 1121-1129PubMed Google Scholar, 20Scalfari F. Castagna M. Fattori B. Andreini I. Maremman C. Pelosi P. Comp. Biochem. Physiol. B Biochem. Mol. Biol. 1997; 118: 819-824Crossref Scopus (23) Google Scholar, 21Lacazette E. Gachon A.M. Pitiot G. Hum. Mol. Genet. 2000; 9: 289-301Crossref PubMed Scopus (59) Google Scholar). Lcn-1 is an unusual member of the lipocalin superfamily, because it is able to bind a broad array of various lipophilic ligands in vitro and in vivo(15Redl B. Holzfeind P. Lottspeich F. J. Biol. Chem. 1992; 267: 20282-20287Abstract Full Text PDF PubMed Google Scholar, 22Glasgow B.J. Abduragimov A.R. Farahakhsh Z.T. Faull K.F. Hubbell W.L. Curr. Eye Res. 1995; 14: 363-372Crossref PubMed Scopus (163) Google Scholar) and was demonstrated to exhibit cysteine proteinase inhibition and nonspecific endonuclease activity in vitro (23van't Hof W. Blankenvoorde M.F.J. Veerman E.C.I. Nieuw Amerongen A.V. J. Biol. Chem. 1997; 272: 1837-1841Abstract Full Text Full Text PDF PubMed Scopus (118) Google Scholar, 24Yusifov T.N. Abduragimov A.R. Gasymov O.K. Glasgow B.J. Biochem. J. 2000; 347: 815-819Crossref PubMed Scopus (49) Google Scholar). Although the biological relevance of its multiple activities has still to be established, its main function seems to be scavenging of lipophilic, potentially harmful molecules, thus acting as a protection factor for cells and tissues (25Redl B. Biochim. Biophys. Acta. 2000; 1482: 241-248Crossref PubMed Scopus (104) Google Scholar). However, the mechanism of clearance or detoxification of the putative harmful ligands is unknown.In the present study we have used a phage display-based technique for interaction screening of a complex cDNA expression library with Lcn-1 as bait to isolate proteins that may be involved in the reception or degradation of Lcn-1-specific ligands. Here we describe the identification, molecular cloning, expression, and subcellular localization of a novel Lcn-1 interacting membrane protein. Our findings set the stage for exploring the molecular mechanism of the lipocalin-receptor interaction in more detail. Moreover, this is the first successful attempt of identifying a lipocalin receptor using phage-display techniques.DISCUSSIONLcn-1 is of considerable interest, because it was supposed to be a member of the immunocalin subfamily of lipocalins, which are involved in modulation of immune and inflammatory responses, and is one of the lipocalins used as a biochemical marker of disease in man (33Lögdberg L. Wester L. Biochim. Biophys. Acta. 2000; 1482: 284-297Crossref PubMed Scopus (167) Google Scholar, 34Xu S. Venge P. Biochim. Biophys. Acta. 2000; 1482: 289-307Crossref Scopus (241) Google Scholar). Functionally, it seems to act as a physiological scavenger of lipophilic, potentially harmful compounds. In search for identification of Lcn-1 interacting proteins involved in the reception or detoxification of Lcn-1 ligands we have applied a phage-display technology using a complex cDNA library and expression screening against Lcn-1 as a bait. Using this method we have identified and characterized a novel human cell membrane protein, LIMR, which is likely to be a receptor for lipophilic ligands physiologically bound to Lcn-1. The predicted topology of LIMR, consisting of 487 amino acids, is complex. It contains nine putative transmembrane domains, with the N terminus orientated outside of the cell. The outside orientation of the N terminus is supported by the result from phage display and the fact that an antiserum raised against this part of the protein reacted with NT2 cells. From phage display it is also evident that the N terminus of LIMR represents the Lcn-1 interacting domain. A notable feature of LIMR is a large central cytoplasmatic loop consisting of 82 amino acids, whereas all of the other loops are rather small (between 18 and 27 amino acids). Both immunocytochemical analysis and cell fractionation experiments gave clear evidence of LIMR being located within the cell plasma membrane. Therefore, it has to be characterized as the first human cell surface membrane-located lipocalin receptor.Although there is clear experimental evidence that many lipocalins bind to specific cell surface receptors, isolation of these proteins or cloning of the encoding genes has failed so far. Because of its important role in retinol transport and retinol supply of cells, a large number of investigations have dealt with the receptor for the classical lipocalin RBP. Thereby, the gene encoding a 63-kDa protein, suspected to be a receptor of bovine plasma retinol-binding protein, was cloned (35Bavik C.O. Levy F. Hellmann U. Wernstedt C. Eriksson U. J. Biol. Chem. 1993; 268: 20540-20546Abstract Full Text PDF PubMed Google Scholar). However, several consecutive reports demonstrated that it is not an integral membrane receptor but rather a membrane-associated protein involved in enzymatic processing of retinol (36Nicoletti A. Wong D.J. Kawase K. Gibson L.H. Yang-feng T.L. Richards J.E. Thompson D.A. Hum. Mol. Genet. 1995; 4: 641-649Crossref PubMed Scopus (105) Google Scholar, 37Redmond T.M., Yu, S. Lee E. Bok D. Hamasaki D. Chen N. Goletz P. Ma J.X. Crouch R.K. Pfeifer K. Nat. Genet. 1998; 20: 344-351Crossref PubMed Scopus (776) Google Scholar). Because no other specific RBP receptor has been identified so far, there is still no consensus whether retinol transfer into the cell is receptor-driven or proceeds via passive diffusion (7Sundaram M. Sivaprasadarao A. DeSousa M.M. Findlay J.B.C. J. Biol. Chem. 1998; 273: 3336-3342Abstract Full Text Full Text PDF PubMed Scopus (100) Google Scholar, 38Noy N. Xu Z.J. Biochemistry. 1990; 29: 3613-3619Google Scholar). There is also convincing evidence that in some cells RBP undergoes internalization by a receptor-mediated endocytotic process (39Senoo H. Smeland S. Malaba L. Bjerknes T. Stang E. Roos N. Berg T. Norum K.R. Blomhoff R. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 3616-3620Crossref PubMed Scopus (57) Google Scholar).Another well characterized protein that is supposed to interact with lipocalins is megalin. This protein, also called gp330, is an epithelial endocytic receptor with a single transmembrane region and shares some characteristic features with the low density lipoprotein receptor family (40Saito A. Pietromonaco S. Loo A.K. Farquhar M.G. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 9725-9729Crossref PubMed Scopus (492) Google Scholar, 41Korenberg J.R. Argraves K.M. Chen X.N. Tran H. Strickland D.K. Argraves W.S. Genomics. 1994; 22: 88-93Crossref PubMed Scopus (42) Google Scholar). From studies on a megalin knockout mouse it was concluded to interact with different lipocalins, including RBP, major urinary protein, olfactory-binding protein, and α1-microglobulin (42Leheste J.R. Rolinski B. Vorum H. Hilpert J. Nykjaer A. Jacobsen C. Aucouturier P. Moskaug J.O. Otto A. Christensen E.I. Willnow T.E. Am. J. Pathol. 1999; 155: 1361-1370Abstract Full Text Full Text PDF PubMed Scopus (342) Google Scholar). However, because it also binds a variety of other macromolecular ligands, such as thyroglobulin, apolipoprotein A, clusterin, albumin, and insulin, its specific role in lipocalin binding remains unclear.With exception of these two defined molecules the interaction of other lipocalins with putative receptors is only marginally characterized, and a number of interactions described are likely to be of no physiological relevance. However, those receptors that have been characterized in some detail show no consensus in biochemical or biophysical properties, suggesting a great diversity of receptor structure (14Flower D.R. Biochim. Biophys. Acta. 2000; 1482: 327-336Crossref PubMed Scopus (119) Google Scholar).There is also no consensus in the mechanism of interaction between lipocalins and their receptors. In principle, lipocalin receptors have been found in both groups, carbohydrate-binding receptors and receptors binding via protein-protein interaction. An example for the first group seems to be the receptor for α1-acid glycoprotein, a highly sialylated serum lipocalin. Inhibition of α1-acid glycoprotein binding to several cells could be inhibited by simple sugars, like mannose and GlcNAc, suggesting that this interaction is mediated by carbohydrates and that the receptor is of a lectin-type (43Andersen U.O. Kirkeby S. Bog-Hansen T.C. J. Mol. Recognit. 1996; 9: 364-367Crossref PubMed Google Scholar). Our investigations clearly show that LIMR is an example for a receptor-type binding via direct protein interaction, because it was isolated by means of prokaryotic expression, and a recombinant LIMR produced inE. coli interacts in vitro with Lcn-1.Apart from its interaction with Lcn-1 the precise physiological function of LIMR is unknown to date. Supposing that it is involved in detoxification of ligands bound to Lcn-1, LIMR could be directly involved in the transfer of ligands to the cell, where they undergo detoxification. Alternatively, it could act as a detoxification proteinper se, probably by displaying an enzymatic activity. Indeed, membrane-associated enzymes involved in modification of lipid molecules have been described, e.g. human enzymes involved in sterol or cholesterol modification (44Robinson G.W. Tsay Y.H. Kienzle B.K. Smith-Monroy C.A. Bishop R.W. Mol. Cell. Biol. 1993; 13: 2706-2717Crossref PubMed Scopus (146) Google Scholar, 45Silve S. Dupuy P.H. Labit-Lebouteiller C. Kaghad M. Chalon P. Rahier A. Taton M. Lupker J. Shire D. Loison G. J. Biol. Chem. 1996; 271: 22434-22440Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar). Interestingly, a recently identified human Δ7-sterol reductase, involved in reduction of 7-dehydrocholesterol to cholesterol, shows a similar overall topography to LIMR, including nine transmembrane regions and a molecular mass of 55 kDa (46Moebius F.F. Fitzky B.U. Lee J.N. Paik Y.K. Glossmann H. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 1899-1902Crossref PubMed Scopus (186) Google Scholar).Although many aspects concerning the precise function of LIMR are speculative at the moment, our investigations might have important consequences for the isolation and characterization of lipocalin receptors, in general. First, we could demonstrate that the phage-display strategy applied is a reliable method for isolating such receptors. Second, our findings constitute the first report of identifying a lipocalin receptor that is, in contrast to the RBP-associated p63, clearly a plasma membrane-located protein. Furthermore, the availability of a cloned gene is a vital first step for elucidating the structural requirements necessary for lipocalin-receptor interaction. Finally, the identification of an Lcn-1-specific plasma membrane receptor will add new hints to the precise physiological function of Lcn-1 and may help to dissect putative detoxification pathways for the ligands bound. The protein superfamily of lipocalins consists of small, mainly secretory proteins defined on the basis of conserved amino acid sequence motifs and their common structure. Functionally they share several properties including the ability to bind/transport a remarkable array of small hydrophobic molecules, the formation of macromolecular complexes, and the binding to specific cell surface receptors (1Flower D.R. Biochem. J. 1996; 318: 1-14Crossref PubMed Scopus (1372) Google Scholar, 2Akerstrom B. Flower D.R. Salier J-P. Biochim. Biophys. Acta. 2000; 1482: 1-8Crossref PubMed Scopus (233) Google Scholar, 3Flower D.R. North A.C.T. Sansom C.E. Biochim. Biophys. Acta. 2000; 1482: 9-24Crossref PubMed Scopus (689) Google Scholar). Whereas a large number of various lipophilic ligands able to bind to lipocalins are known, only limited data are available concerning the identity of lipocalin receptors. There is clear evidence of a specific receptor for plasma retinol-binding protein (RBP)1 (4Bavik C.O. Busch C. Eriksson U. J. Biol. Chem. 1992; 267: 23035-23042Abstract Full Text PDF PubMed Google Scholar, 5Smeland S. Bjerknes T. Malaba L. Eskild W. Norum K.R. Blomhoff R. Biochem. J. 1995; 305: 419-424Crossref PubMed Scopus (58) Google Scholar, 6Sivaprasadarao A. Boudjelal M. Findlay J.B.C. Biochem. J. 1994; 302: 245-251Crossref PubMed Scopus (41) Google Scholar, 7Sundaram M. Sivaprasadarao A. DeSousa M.M. Findlay J.B.C. J. Biol. Chem. 1998; 273: 3336-3342Abstract Full Text Full Text PDF PubMed Scopus (100) Google Scholar) and more indirect evidence for receptors for α1-microglobulin (8Wester L. Michaelsson E. Holmdahl R. Olofsson T. Akerstrom B. Scand. J. Immunol. 1998; 48: 1-7Crossref PubMed Scopus (33) Google Scholar), major urinary protein (9Böcskei Z. Groom C.R. Flower D.R. Wright C.R. Phillips S.E. Cavaggioni A. Findlay J.B. North A.C. Nature. 1992; 360: 186-188Crossref PubMed Scopus (346) Google Scholar), β-lactoglobulin (10Papiz M.Z. Sawyer L. Eliopoulos E.E. North A.C. Findlay J.B. Sivaprasadarao R. Jones T.A. Newcomer M.E. Kraulis P.J. Nature. 1986; 324: 383-385Crossref PubMed Scopus (847) Google Scholar), olfactory-binding protein (11Boudjelal M. Sivaprasadarao A. Findlay B.B. Biochem. J. 1996; 317: 23-27Crossref PubMed Scopus (36) Google Scholar), α1-acid glycoprotein (12Andersen U.O. Bog-Hansen S. Kirkeby S. Acta Histochem. 1999; 101: 113-119Crossref PubMed Scopus (7) Google Scholar), and glycodelin (13Miller R.E. Fayen J.D. Chakraborty S. Weber M.C. Tykocinski M.L. FEBS Lett. 1998; 436: 455-460Crossref PubMed Scopus (34) Google Scholar), but with the exception of megalin, which seems to be an endocytic receptor for a variety of soluble macromolecules including several lipocalins, no specific lipocalin receptor has been fully characterized so far (14Flower D.R. Biochim. Biophys. Acta. 2000; 1482: 327-336Crossref PubMed Scopus (119) Google Scholar). This lack of knowledge is a major disadvantage in understanding the biological function of many lipocalin members. We have identified Lcn-1 (identical with tear lipocalin or von Ebner's gland protein) as a human member of the lipocalin superfamily (15Redl B. Holzfeind P. Lottspeich F. J. Biol. Chem. 1992; 267: 20282-20287Abstract Full Text PDF PubMed Google Scholar). It is produced by a number of secretory glands and tissues, including lacrimal and lingual salivary glands, prostate, and mucosal glands of the tracheobronchial tree, nasal mucosa, and sweat glands and by some neuroendocrine tissues (15Redl B. Holzfeind P. Lottspeich F. J. Biol. Chem. 1992; 267: 20282-20287Abstract Full Text PDF PubMed Google Scholar, 16Lassagne H. Gachon A.M. Exp. Eye Res. 1993; 55: 605-609Crossref Scopus (28) Google Scholar, 17Bläker M. Kock K. Ahlers C. Buck F. Schmale H. Biochim. Biophys. Acta. 1993; 1172: 131-137Crossref PubMed Scopus (77) Google Scholar, 18Holzfeind P. Merschak P. Rogatsch H. Culig Z. Feichtinger H. Klocker H. Redl B. FEBS Lett. 1996; 395: 95-98Crossref PubMed Scopus (39) Google Scholar, 19Redl B. Wojnar P. Ellemunter H. Feichtinger H. Lab. Invest. 1998; 78: 1121-1129PubMed Google Scholar, 20Scalfari F. Castagna M. Fattori B. Andreini I. Maremman C. Pelosi P. Comp. Biochem. Physiol. B Biochem. Mol. Biol. 1997; 118: 819-824Crossref Scopus (23) Google Scholar, 21Lacazette E. Gachon A.M. Pitiot G. Hum. Mol. Genet. 2000; 9: 289-301Crossref PubMed Scopus (59) Google Scholar). Lcn-1 is an unusual member of the lipocalin superfamily, because it is able to bind a broad array of various lipophilic ligands in vitro and in vivo(15Redl B. Holzfeind P. Lottspeich F. J. Biol. Chem. 1992; 267: 20282-20287Abstract Full Text PDF PubMed Google Scholar, 22Glasgow B.J. Abduragimov A.R. Farahakhsh Z.T. Faull K.F. Hubbell W.L. Curr. Eye Res. 1995; 14: 363-372Crossref PubMed Scopus (163) Google Scholar) and was demonstrated to exhibit cysteine proteinase inhibition and nonspecific endonuclease activity in vitro (23van't Hof W. Blankenvoorde M.F.J. Veerman E.C.I. Nieuw Amerongen A.V. J. Biol. Chem. 1997; 272: 1837-1841Abstract Full Text Full Text PDF PubMed Scopus (118) Google Scholar, 24Yusifov T.N. Abduragimov A.R. Gasymov O.K. Glasgow B.J. Biochem. J. 2000; 347: 815-819Crossref PubMed Scopus (49) Google Scholar). Although the biological relevance of its multiple activities has still to be established, its main function seems to be scavenging of lipophilic, potentially harmful molecules, thus acting as a protection factor for cells and tissues (25Redl B. Biochim. Biophys. Acta. 2000; 1482: 241-248Crossref PubMed Scopus (104) Google Scholar). However, the mechanism of clearance or detoxification of the putative harmful ligands is unknown. In the present study we have used a phage display-based technique for interaction screening of a complex cDNA expression library with Lcn-1 as bait to isolate proteins that may be involved in the reception or degradation of Lcn-1-specific ligands. Here we describe the identification, molecular cloning, expression, and subcellular localization of a novel Lcn-1 interacting membrane protein. Our findings set the stage for exploring the molecular mechanism of the lipocalin-receptor interaction in more detail. Moreover, this is the first successful attempt of identifying a lipocalin receptor using phage-display techniques. DISCUSSIONLcn-1 is of considerable interest, because it was supposed to be a member of the immunocalin subfamily of lipocalins, which are involved in modulation of immune and inflammatory responses, and is one of the lipocalins used as a biochemical marker of disease in man (33Lögdberg L. Wester L. Biochim. Biophys. Acta. 2000; 1482: 284-297Crossref PubMed Scopus (167) Google Scholar, 34Xu S. Venge P. Biochim. Biophys. Acta. 2000; 1482: 289-307Crossref Scopus (241) Google Scholar). Functionally, it seems to act as a physiological scavenger of lipophilic, potentially harmful compounds. In search for identification of Lcn-1 interacting proteins involved in the reception or detoxification of Lcn-1 ligands we have applied a phage-display technology using a complex cDNA library and expression screening against Lcn-1 as a bait. Using this method we have identified and characterized a novel human cell membrane protein, LIMR, which is likely to be a receptor for lipophilic ligands physiologically bound to Lcn-1. The predicted topology of LIMR, consisting of 487 amino acids, is complex. It contains nine putative transmembrane domains, with the N terminus orientated outside of the cell. The outside orientation of the N terminus is supported by the result from phage display and the fact that an antiserum raised against this part of the protein reacted with NT2 cells. From phage display it is also evident that the N terminus of LIMR represents the Lcn-1 interacting domain. A notable feature of LIMR is a large central cytoplasmatic loop consisting of 82 amino acids, whereas all of the other loops are rather small (between 18 and 27 amino acids). Both immunocytochemical analysis and cell fractionation experiments gave clear evidence of LIMR being located within the cell plasma membrane. Therefore, it has to be characterized as the first human cell surface membrane-located lipocalin receptor.Although there is clear experimental evidence that many lipocalins bind to specific cell surface receptors, isolation of these proteins or cloning of the encoding genes has failed so far. Because of its important role in retinol transport and retinol supply of cells, a large number of investigations have dealt with the receptor for the classical lipocalin RBP. Thereby, the gene encoding a 63-kDa protein, suspected to be a receptor of bovine plasma retinol-binding protein, was cloned (35Bavik C.O. Levy F. Hellmann U. Wernstedt C. Eriksson U. J. Biol. Chem. 1993; 268: 20540-20546Abstract Full Text PDF PubMed Google Scholar). However, several consecutive reports demonstrated that it is not an integral membrane receptor but rather a membrane-associated protein involved in enzymatic processing of retinol (36Nicoletti A. Wong D.J. Kawase K. Gibson L.H. Yang-feng T.L. Richards J.E. Thompson D.A. Hum. Mol. Genet. 1995; 4: 641-649Crossref PubMed Scopus (105) Google Scholar, 37Redmond T.M., Yu, S. Lee E. Bok D. Hamasaki D. Chen N. Goletz P. Ma J.X. Crouch R.K. Pfeifer K. Nat. Genet. 1998; 20: 344-351Crossref PubMed Scopus (776) Google Scholar). Because no other specific RBP receptor has been identified so far, there is still no consensus whether retinol transfer into the cell is receptor-driven or proceeds via passive diffusion (7Sundaram M. Sivaprasadarao A. DeSousa M.M. Findlay J.B.C. J. Biol. Chem. 1998; 273: 3336-3342Abstract Full Text Full Text PDF PubMed Scopus (100) Google Scholar, 38Noy N. Xu Z.J. Biochemistry. 1990; 29: 3613-3619Google Scholar). There is also convincing evidence that in some cells RBP undergoes internalization by a receptor-mediated endocytotic process (39Senoo H. Smeland S. Malaba L. Bjerknes T. Stang E. Roos N. Berg T. Norum K.R. Blomhoff R. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 3616-3620Crossref PubMed Scopus (57) Google Scholar).Another well characterized protein that is supposed to interact with lipocalins is megalin. This protein, also called gp330, is an epithelial endocytic receptor with a single transmembrane region and shares some characteristic features with the low density lipoprotein receptor family (40Saito A. Pietromonaco S. Loo A.K. Farquhar M.G. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 9725-9729Crossref PubMed Scopus (492) Google Scholar, 41Korenberg J.R. Argraves K.M. Chen X.N. Tran H. Strickland D.K. Argraves W.S. Genomics. 1994; 22: 88-93Crossref PubMed Scopus (42) Google Scholar). From studies on a megalin knockout mouse it was concluded to interact with different lipocalins, including RBP, major urinary protein, olfactory-binding protein, and α1-microglobulin (42Leheste J.R. Rolinski B. Vorum H. Hilpert J. Nykjaer A. Jacobsen C. Aucouturier P. Moskaug J.O. Otto A. Christensen E.I. Willnow T.E. Am. J. Pathol. 1999; 155: 1361-1370Abstract Full Text Full Text PDF PubMed Scopus (342) Google Scholar). However, because it also binds a variety of other macromolecular ligands, such as thyroglobulin, apolipoprotein A, clusterin, albumin, and insulin, its specific role in lipocalin binding remains unclear.With exception of these two defined molecules the interaction of other lipocalins with putative receptors is only marginally characterized, and a number of interactions described are likely to be of no physiological relevance. However, those receptors that have been characterized in some detail show no consensus in biochemical or biophysical properties, suggesting a great diversity of receptor structure (14Flower D.R. Biochim. Biophys. Acta. 2000; 1482: 327-336Crossref PubMed Scopus (119) Google Scholar).There is also no consensus in the mechanism of interaction between lipocalins and their receptors. In principle, lipocalin receptors have been found in both groups, carbohydrate-binding receptors and receptors binding via protein-protein interaction. An example for the first group seems to be the receptor for α1-acid glycoprotein, a highly sialylated serum lipocalin. Inhibition of α1-acid glycoprotein binding to several cells could be inhibited by simple sugars, like mannose and GlcNAc, suggesting that this interaction is mediated by carbohydrates and that the receptor is of a lectin-type (43Andersen U.O. Kirkeby S. Bog-Hansen T.C. J. Mol. Recognit. 1996; 9: 364-367Crossref PubMed Google Scholar). Our investigations clearly show that LIMR is an example for a receptor-type binding via direct protein interaction, because it was isolated by means of prokaryotic expression, and a recombinant LIMR produced inE. coli interacts in vitro with Lcn-1.Apart from its interaction with Lcn-1 the precise physiological function of LIMR is unknown to date. Supposing that it is involved in detoxification of ligands bound to Lcn-1, LIMR could be directly involved in the transfer of ligands to the cell, where they undergo detoxification. Alternatively, it could act as a detoxification proteinper se, probably by displaying an enzymatic activity. Indeed, membrane-associated enzymes involved in modification of lipid molecules have been described, e.g. human enzymes involved in sterol or cholesterol modification (44Robinson G.W. Tsay Y.H. Kienzle B.K. Smith-Monroy C.A. Bishop R.W. Mol. Cell. Biol. 1993; 13: 2706-2717Crossref PubMed Scopus (146) Google Scholar, 45Silve S. Dupuy P.H. Labit-Lebouteiller C. Kaghad M. Chalon P. Rahier A. Taton M. Lupker J. Shire D. Loison G. J. Biol. Chem. 1996; 271: 22434-22440Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar). Interestingly, a recently identified human Δ7-sterol reductase, involved in reduction of 7-dehydrocholesterol to cholesterol, shows a similar overall topography to LIMR, including nine transmembrane regions and a molecular mass of 55 kDa (46Moebius F.F. Fitzky B.U. Lee J.N. Paik Y.K. Glossmann H. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 1899-1902Crossref PubMed Scopus (186) Google Scholar).Although many aspects concerning the precise function of LIMR are speculative at the moment, our investigations might have important consequences for the isolation and characterization of lipocalin receptors, in general. First, we could demonstrate that the phage-display strategy applied is a reliable method for isolating such receptors. Second, our findings constitute the first report of identifying a lipocalin receptor that is, in contrast to the RBP-associated p63, clearly a plasma membrane-located protein. Furthermore, the availability of a cloned gene is a vital first step for elucidating the structural requirements necessary for lipocalin-receptor interaction. Finally, the identification of an Lcn-1-specific plasma membrane receptor will add new hints to the precise physiological function of Lcn-1 and may help to dissect putative detoxification pathways for the ligands bound. Lcn-1 is of considerable interest, because it was supposed to be a member of the immunocalin subfamily of lipocalins, which are involved in modulation of immune and inflammatory responses, and is one of the lipocalins used as a biochemical marker of disease in man (33Lögdberg L. Wester L. Biochim. Biophys. Acta. 2000; 1482: 284-297Crossref PubMed Scopus (167) Google Scholar, 34Xu S. Venge P. Biochim. Biophys. Acta. 2000; 1482: 289-307Crossref Scopus (241) Google Scholar). Functionally, it seems to act as a physiological scavenger of lipophilic, potentially harmful compounds. In search for identification of Lcn-1 interacting proteins involved in the reception or detoxification of Lcn-1 ligands we have applied a phage-display technology using a complex cDNA library and expression screening against Lcn-1 as a bait. Using this method we have identified and characterized a novel human cell membrane protein, LIMR, which is likely to be a receptor for lipophilic ligands physiologically bound to Lcn-1. The predicted topology of LIMR, consisting of 487 amino acids, is complex. It contains nine putative transmembrane domains, with the N terminus orientated outside of the cell. The outside orientation of the N terminus is supported by the result from phage display and the fact that an antiserum raised against this part of the protein reacted with NT2 cells. From phage display it is also evident that the N terminus of LIMR represents the Lcn-1 interacting domain. A notable feature of LIMR is a large central cytoplasmatic loop consisting of 82 amino acids, whereas all of the other loops are rather small (between 18 and 27 amino acids). Both immunocytochemical analysis and cell fractionation experiments gave clear evidence of LIMR being located within the cell plasma membrane. Therefore, it has to be characterized as the first human cell surface membrane-located lipocalin receptor. Although there is clear experimental evidence that many lipocalins bind to specific cell surface receptors, isolation of these proteins or cloning of the encoding genes has failed so far. Because of its important role in retinol transport and retinol supply of cells, a large number of investigations have dealt with the receptor for the classical lipocalin RBP. Thereby, the gene encoding a 63-kDa protein, suspected to be a receptor of bovine plasma retinol-binding protein, was cloned (35Bavik C.O. Levy F. Hellmann U. Wernstedt C. Eriksson U. J. Biol. Chem. 1993; 268: 20540-20546Abstract Full Text PDF PubMed Google Scholar). However, several consecutive reports demonstrated that it is not an integral membrane receptor but rather a membrane-associated protein involved in enzymatic processing of retinol (36Nicoletti A. Wong D.J. Kawase K. Gibson L.H. Yang-feng T.L. Richards J.E. Thompson D.A. Hum. Mol. Genet. 1995; 4: 641-649Crossref PubMed Scopus (105) Google Scholar, 37Redmond T.M., Yu, S. Lee E. Bok D. Hamasaki D. Chen N. Goletz P. Ma J.X. Crouch R.K. Pfeifer K. Nat. Genet. 1998; 20: 344-351Crossref PubMed Scopus (776) Google Scholar). Because no other specific RBP receptor has been identified so far, there is still no consensus whether retinol transfer into the cell is receptor-driven or proceeds via passive diffusion (7Sundaram M. Sivaprasadarao A. DeSousa M.M. Findlay J.B.C. J. Biol. Chem. 1998; 273: 3336-3342Abstract Full Text Full Text PDF PubMed Scopus (100) Google Scholar, 38Noy N. Xu Z.J. Biochemistry. 1990; 29: 3613-3619Google Scholar). There is also convincing evidence that in some cells RBP undergoes internalization by a receptor-mediated endocytotic process (39Senoo H. Smeland S. Malaba L. Bjerknes T. Stang E. Roos N. Berg T. Norum K.R. Blomhoff R. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 3616-3620Crossref PubMed Scopus (57) Google Scholar). Another well characterized protein that is supposed to interact with lipocalins is megalin. This protein, also called gp330, is an epithelial endocytic receptor with a single transmembrane region and shares some characteristic features with the low density lipoprotein receptor family (40Saito A. Pietromonaco S. Loo A.K. Farquhar M.G. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 9725-9729Crossref PubMed Scopus (492) Google Scholar, 41Korenberg J.R. Argraves K.M. Chen X.N. Tran H. Strickland D.K. Argraves W.S. Genomics. 1994; 22: 88-93Crossref PubMed Scopus (42) Google Scholar). From studies on a megalin knockout mouse it was concluded to interact with different lipocalins, including RBP, major urinary protein, olfactory-binding protein, and α1-microglobulin (42Leheste J.R. Rolinski B. Vorum H. Hilpert J. Nykjaer A. Jacobsen C. Aucouturier P. Moskaug J.O. Otto A. Christensen E.I. Willnow T.E. Am. J. Pathol. 1999; 155: 1361-1370Abstract Full Text Full Text PDF PubMed Scopus (342) Google Scholar). However, because it also binds a variety of other macromolecular ligands, such as thyroglobulin, apolipoprotein A, clusterin, albumin, and insulin, its specific role in lipocalin binding remains unclear. With exception of these two defined molecules the interaction of other lipocalins with putative receptors is only marginally characterized, and a number of interactions described are likely to be of no physiological relevance. However, those receptors that have been characterized in some detail show no consensus in biochemical or biophysical properties, suggesting a great diversity of receptor structure (14Flower D.R. Biochim. Biophys. Acta. 2000; 1482: 327-336Crossref PubMed Scopus (119) Google Scholar). There is also no consensus in the mechanism of interaction between lipocalins and their receptors. In principle, lipocalin receptors have been found in both groups, carbohydrate-binding receptors and receptors binding via protein-protein interaction. An example for the first group seems to be the receptor for α1-acid glycoprotein, a highly sialylated serum lipocalin. Inhibition of α1-acid glycoprotein binding to several cells could be inhibited by simple sugars, like mannose and GlcNAc, suggesting that this interaction is mediated by carbohydrates and that the receptor is of a lectin-type (43Andersen U.O. Kirkeby S. Bog-Hansen T.C. J. Mol. Recognit. 1996; 9: 364-367Crossref PubMed Google Scholar). Our investigations clearly show that LIMR is an example for a receptor-type binding via direct protein interaction, because it was isolated by means of prokaryotic expression, and a recombinant LIMR produced inE. coli interacts in vitro with Lcn-1. Apart from its interaction with Lcn-1 the precise physiological function of LIMR is unknown to date. Supposing that it is involved in detoxification of ligands bound to Lcn-1, LIMR could be directly involved in the transfer of ligands to the cell, where they undergo detoxification. Alternatively, it could act as a detoxification proteinper se, probably by displaying an enzymatic activity. Indeed, membrane-associated enzymes involved in modification of lipid molecules have been described, e.g. human enzymes involved in sterol or cholesterol modification (44Robinson G.W. Tsay Y.H. Kienzle B.K. Smith-Monroy C.A. Bishop R.W. Mol. Cell. Biol. 1993; 13: 2706-2717Crossref PubMed Scopus (146) Google Scholar, 45Silve S. Dupuy P.H. Labit-Lebouteiller C. Kaghad M. Chalon P. Rahier A. Taton M. Lupker J. Shire D. Loison G. J. Biol. Chem. 1996; 271: 22434-22440Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar). Interestingly, a recently identified human Δ7-sterol reductase, involved in reduction of 7-dehydrocholesterol to cholesterol, shows a similar overall topography to LIMR, including nine transmembrane regions and a molecular mass of 55 kDa (46Moebius F.F. Fitzky B.U. Lee J.N. Paik Y.K. Glossmann H. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 1899-1902Crossref PubMed Scopus (186) Google Scholar). Although many aspects concerning the precise function of LIMR are speculative at the moment, our investigations might have important consequences for the isolation and characterization of lipocalin receptors, in general. First, we could demonstrate that the phage-display strategy applied is a reliable method for isolating such receptors. Second, our findings constitute the first report of identifying a lipocalin receptor that is, in contrast to the RBP-associated p63, clearly a plasma membrane-located protein. Furthermore, the availability of a cloned gene is a vital first step for elucidating the structural requirements necessary for lipocalin-receptor interaction. Finally, the identification of an Lcn-1-specific plasma membrane receptor will add new hints to the precise physiological function of Lcn-1 and may help to dissect putative detoxification pathways for the ligands bound. We are grateful to R. Crameri for providing the vector pJuFo and to C. Enzinger for the antiserum against NMDA. We thank M. Schöser, F. Marx, and B. Hörtnagl for helpful comments in preparing the manuscript." @default.
W2042871707 created "2016-06-24" @default.
W2042871707 creator A5010030468 @default.
W2042871707 creator A5055809121 @default.
W2042871707 creator A5067305600 @default.
W2042871707 creator A5071423267 @default.
W2042871707 date "2001-01-01" @default.
W2042871707 modified "2023-10-13" @default.
W2042871707 title "Molecular Cloning of a Novel Lipocalin-1 Interacting Human Cell Membrane Receptor Using Phage Display" @default.
W2042871707 cites W10606439 @default.
W2042871707 cites W1501007768 @default.
W2042871707 cites W1501887512 @default.
W2042871707 cites W1507155737 @default.
W2042871707 cites W1587435244 @default.
W2042871707 cites W1835254399 @default.
W2042871707 cites W1859885259 @default.
W2042871707 cites W1964579574 @default.
W2042871707 cites W1973132656 @default.
W2042871707 cites W1976172969 @default.
W2042871707 cites W1976506214 @default.
W2042871707 cites W1985753275 @default.
W2042871707 cites W1989642228 @default.
W2042871707 cites W1991944268 @default.
W2042871707 cites W2003113302 @default.
W2042871707 cites W2021882216 @default.
W2042871707 cites W2024142971 @default.
W2042871707 cites W2026183815 @default.
W2042871707 cites W2026191433 @default.
W2042871707 cites W2038438679 @default.
W2042871707 cites W2039760937 @default.
W2042871707 cites W2040141270 @default.
W2042871707 cites W2040228918 @default.
W2042871707 cites W2043586365 @default.
W2042871707 cites W2055182510 @default.
W2042871707 cites W2061940389 @default.
W2042871707 cites W2063007989 @default.
W2042871707 cites W2071711022 @default.
W2042871707 cites W2072893670 @default.
W2042871707 cites W2074947829 @default.
W2042871707 cites W2083487562 @default.
W2042871707 cites W2094436192 @default.
W2042871707 cites W2112727307 @default.
W2042871707 cites W2115170303 @default.
W2042871707 cites W2120684043 @default.
W2042871707 cites W2128556903 @default.
W2042871707 cites W2147456532 @default.
W2042871707 cites W2148147338 @default.
W2042871707 cites W2166523594 @default.
W2042871707 cites W2280545067 @default.
W2042871707 cites W4294216491 @default.
W2042871707 doi "https://doi.org/10.1074/jbc.m101762200" @default.
W2042871707 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/11287427" @default.
W2042871707 hasPublicationYear "2001" @default.
W2042871707 type Work @default.
W2042871707 sameAs 2042871707 @default.
W2042871707 citedByCount "69" @default.
W2042871707 countsByYear W20428717072012 @default.
W2042871707 countsByYear W20428717072013 @default.
W2042871707 countsByYear W20428717072014 @default.
W2042871707 countsByYear W20428717072015 @default.
W2042871707 countsByYear W20428717072016 @default.
W2042871707 countsByYear W20428717072017 @default.
W2042871707 countsByYear W20428717072018 @default.
W2042871707 countsByYear W20428717072019 @default.
W2042871707 countsByYear W20428717072020 @default.
W2042871707 countsByYear W20428717072021 @default.
W2042871707 countsByYear W20428717072022 @default.
W2042871707 crossrefType "journal-article" @default.
W2042871707 hasAuthorship W2042871707A5010030468 @default.
W2042871707 hasAuthorship W2042871707A5055809121 @default.
W2042871707 hasAuthorship W2042871707A5067305600 @default.
W2042871707 hasAuthorship W2042871707A5071423267 @default.
W2042871707 hasBestOaLocation W20428717071 @default.
W2042871707 hasConcept C104317684 @default.
W2042871707 hasConcept C121050878 @default.
W2042871707 hasConcept C153911025 @default.
W2042871707 hasConcept C159654299 @default.
W2042871707 hasConcept C167625842 @default.
W2042871707 hasConcept C170493617 @default.
W2042871707 hasConcept C185592680 @default.
W2042871707 hasConcept C186268636 @default.
W2042871707 hasConcept C19924922 @default.
W2042871707 hasConcept C199360897 @default.
W2042871707 hasConcept C20299238 @default.
W2042871707 hasConcept C41008148 @default.
W2042871707 hasConcept C54355233 @default.
W2042871707 hasConcept C55493867 @default.
W2042871707 hasConcept C70721500 @default.
W2042871707 hasConcept C86803240 @default.
W2042871707 hasConcept C95444343 @default.
W2042871707 hasConceptScore W2042871707C104317684 @default.
W2042871707 hasConceptScore W2042871707C121050878 @default.
W2042871707 hasConceptScore W2042871707C153911025 @default.
W2042871707 hasConceptScore W2042871707C159654299 @default.
W2042871707 hasConceptScore W2042871707C167625842 @default.
W2042871707 hasConceptScore W2042871707C170493617 @default.
W2042871707 hasConceptScore W2042871707C185592680 @default.
W2042871707 hasConceptScore W2042871707C186268636 @default.