Sp1-binding Elements in the Promoter of RAGE Are Essential for Amphoterin-mediated Gene Expression in Cultured Neuroblastoma Cells

Abstract

Receptor for AGE (RAGE) and the polypeptide amphoterin are highly expressed and co-localized in neurons of the developing central nervous system of the rat. In vitro, the interaction of amphoterin with neuronal RAGE induces neurite outgrowth. We tested the hypothesis that interaction of amphoterin with neuronal cells enhances RAGE expression, thereby providing a mechanism by which amphoterin-mediated regulation of RAGE might contribute to promotion of neurite growth and spreading. Incubation of cultured neuroblastoma cells with amphoterin resulted in increased transcription and translation of RAGE, a process largely inhibited in the presence of anti-RAGE IgG but not by nonimmune IgG. To begin to delineate molecular mechanisms underlying these findings, we identified multiple putative binding elements within the 5′-flanking region of the RAGE gene for Sp1, a transcription factor that has been critically linked to the process of normal development. DNase I footprinting and electrophoretic mobility shift assays demonstrated multiple functional Sp1-binding sites within the region −245 to −40 of the RAGE promoter. Transient transfection of cultured SK-N-SH neuroblastoma cells with chimeric 5′-deletion constructs linked to luciferase reporter revealed that the region containing Sp1-binding elements did not contribute uniquely to basal expression of the RAGE gene. Simultaneous mutation of the multiple Sp1-binding elements in this region did not affect basal promoter function; however, promoter responsiveness to amphoterin was markedly attenuated. These results point to Sp1-dependent mechanisms underlying amphoterin-mediated increases in RAGE expression in neuroblastoma cells and further link amphoterin-RAGE interaction to development of the nervous system. Receptor for AGE (RAGE) and the polypeptide amphoterin are highly expressed and co-localized in neurons of the developing central nervous system of the rat. In vitro, the interaction of amphoterin with neuronal RAGE induces neurite outgrowth. We tested the hypothesis that interaction of amphoterin with neuronal cells enhances RAGE expression, thereby providing a mechanism by which amphoterin-mediated regulation of RAGE might contribute to promotion of neurite growth and spreading. Incubation of cultured neuroblastoma cells with amphoterin resulted in increased transcription and translation of RAGE, a process largely inhibited in the presence of anti-RAGE IgG but not by nonimmune IgG. To begin to delineate molecular mechanisms underlying these findings, we identified multiple putative binding elements within the 5′-flanking region of the RAGE gene for Sp1, a transcription factor that has been critically linked to the process of normal development. DNase I footprinting and electrophoretic mobility shift assays demonstrated multiple functional Sp1-binding sites within the region −245 to −40 of the RAGE promoter. Transient transfection of cultured SK-N-SH neuroblastoma cells with chimeric 5′-deletion constructs linked to luciferase reporter revealed that the region containing Sp1-binding elements did not contribute uniquely to basal expression of the RAGE gene. Simultaneous mutation of the multiple Sp1-binding elements in this region did not affect basal promoter function; however, promoter responsiveness to amphoterin was markedly attenuated. These results point to Sp1-dependent mechanisms underlying amphoterin-mediated increases in RAGE expression in neuroblastoma cells and further link amphoterin-RAGE interaction to development of the nervous system. receptor for AGE advanced glycation end products analysis of variance bovine serum albumin fetal bovine serum electrophoretic mobility shift assay phosphate-buffered saline polymerase chain reaction enzyme-linked immunosorbent assay 1,4-piperazinediethanesulfonic acid glyceraldehyde-3-phosphate dehydrogenase interleukin 6. The receptor for AGE (RAGE),1 a member of the immunoglobulin superfamily of cell-surface molecules, interacts with a range of ligands (1Schmidt A.M. Vianna M. Gerlach M. Brett J. Ryan J. Kao J. Esposito C. Hegarty H. Hurley W. Clauss M. Wang F. Pan Y.C. Tsang T.C. Stern D. J. Biol. Chem. 1992; 267: 14987-14997Abstract Full Text PDF PubMed Google Scholar, 2Neeper M. Schmidt A.M. Brett J. Yan S.D. Wang F. Pan Y.C. Elliston K. Stern D. Shaw A. J. Biol. Chem. 1992; 267: 14998-15004Abstract Full Text PDF PubMed Google Scholar). In pathophysiologic circumstances, advanced glycation end products (AGEs) result from nonenzymatic glycation and oxidation of proteins and lipids and accumulate in conditions such as diabetes, renal failure, and amyloidoses (3Brownlee M. Cerami A. Vlassara H. N. Engl. J. Med. 1988; 318: 1315-1320Crossref PubMed Scopus (2317) Google Scholar, 4Miyata T. Oda O. Inagi R. Iida Y. Araki N. Yamada Y. Horiuchi S. Taniguchi N. Maeda K. Kinoshita T. J. Clin. Invest. 1992; 92: 1243-1252Crossref Scopus (409) Google Scholar). AGEs interact with RAGE in a manner linked to perturbation of vascular and inflammatory cells (5Schmidt A.M. Yan S.D. Brett J. Mora R. Stern D. J. Clin. Invest. 1993; 92: 2155-2168Crossref Scopus (269) Google Scholar, 6Yan S.D. Schmidt A.M. Anderson G. Zhang J. Brett J. Zou Y.S. Pinsky D. Stern D. J. Biol. Chem. 1994; 269: 9889-9897Abstract Full Text PDF PubMed Google Scholar, 7Wautier J.-L. Wautier M.P. Schmidt A.M. Anderson G.M. Zoukourian C. Capron L. Chappey O. Yan S.D. Brett J. Guillausseau P.J. Stern D. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 7742-7746Crossref PubMed Scopus (323) Google Scholar, 8Schmidt A.-M. Hori O. Chen J. Li J.F. Crandall J. Zhang J. Cao R. Yan S.D. Brett J. Stern D. J. Clin. Invest. 1995; 96: 1395-1403Crossref PubMed Scopus (809) Google Scholar, 9Schmidt A.M. Weidman E. Lalla E. Yan S.D. Hori O. Cao R. Brett J. Lamster I. J. Periodontal Res. 1996; 31: 508-515Crossref PubMed Scopus (206) Google Scholar). In addition, RAGE interacts with amyloid β-peptide, consequences of which include enhanced neuronal and microglial oxidant stress and dysfunction (10Yan S.D. Chen X. Fu J. Chen M. Zhu H. Roher A. Slattery T. Nagashima M. Morser J. Migheli A. Nawroth P. Godman G. Stern D. Schmidt A.M. Nature. 1996; 382: 685-691Crossref PubMed Scopus (1783) Google Scholar, 11Yan S.-D. Zhu H. Fu J. Yan S.-F. Roher A. Tourtellotte W. Rajavashisth T. Chen X. Stern D. Schmidt A.-M. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 5296-5301Crossref PubMed Scopus (400) Google Scholar). These processes may be important in the pathogenesis of neurodegenerative disorders such as Alzheimer's disease. The expression of RAGE is strikingly enhanced in cells affected in settings associated with increased accumulation of AGEs and amyloid β-peptide (12Abel M. Ritthaler U. Zhang Y. Deng Y. Schmidt A.M. Greten J. Sernau T. Wahl P. Andrassy K. Ritz E. Stern D. Nawroth P.P. Nephrol. Dial. Transplant. 1995; 10: 1662-1667PubMed Google Scholar, 13Soulis T. Thallas V. Youssef S. Gilbert R.E. McWilliam B.G. Murray-McIntosh R.P. Cooper M.E. Diabetologia. 1997; 40: 619-628Crossref PubMed Scopus (198) Google Scholar, 14Schmidt A.M. Yan S.D. Stern D. Nat. Med. 1995; 1: 1002-1004Crossref PubMed Scopus (124) Google Scholar). We recently identified that an important means by which the expression of RAGE might be regulated in inflammatory-type milieu such as diabetic vasculature and wounds, and Alzheimer brain, is via two functional NF-kB-binding elements located with the 5′-flanking region of the RAGE gene (15Li J. Schmidt A.M. J. Biol. Chem. 1997; 272: 16498-16506Abstract Full Text Full Text PDF PubMed Scopus (442) Google Scholar). Transfection of 5′-deletion luciferase reporter gene constructs containing two mutated NF-kB-binding sites into either endothelial cells or vascular smooth muscle cells significantly diminished RAGE promoter responsiveness to prototypic inflammatory stimuli such as lipopolysaccharide (15Li J. Schmidt A.M. J. Biol. Chem. 1997; 272: 16498-16506Abstract Full Text Full Text PDF PubMed Scopus (442) Google Scholar). Our studies have also identified that the expression of RAGE is enhanced in the developing central nervous system, a setting in which excessive accumulation of AGEs and amyloid β-peptide is not likely (16Hori O. Brett J. Slattery T. Cao R. Zhang J. Chen J. Nagashima M. Lundh E.R. Vijay S. Nitecki D. Morser J. Stern D. Schmidt A.M. J. Biol. Chem. 1995; 270: 25752-25761Abstract Full Text Full Text PDF PubMed Scopus (1007) Google Scholar). Indeed, the increased expression of RAGE in neurons of the cerebrum, hippocampus, and cerebellum co-localized with that of the polypeptide amphoterin. In in vitro studies, amphoterin bound neuronal RAGE in a dose-dependent manner and mediated neurite outgrowth. In the presence of blockade of neuronal RAGE, employing either anti-RAGE F(ab′)2 or soluble RAGE, the extracellular two-thirds of RAGE (17Schmidt A.-M. Hasu M. Popov D. Zhang J.-H. Yan S.-D. Brett J. Cao R. Kuwabara K. Costache G. Simionescu N. Simionescu M. Stern D. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 8807-8811Crossref PubMed Scopus (279) Google Scholar, 18Wautier J.-L. Zoukourian C. Chappey O. Wautier M.P. Guillausseau P.J. Cao R. Hori O. Stern D. Schmidt A.M. J. Clin. Invest. 1996; 97: 238-243Crossref PubMed Scopus (492) Google Scholar), amphoterin-mediated neurite outgrowth was inhibited (16Hori O. Brett J. Slattery T. Cao R. Zhang J. Chen J. Nagashima M. Lundh E.R. Vijay S. Nitecki D. Morser J. Stern D. Schmidt A.M. J. Biol. Chem. 1995; 270: 25752-25761Abstract Full Text Full Text PDF PubMed Scopus (1007) Google Scholar). We hypothesized that ligation of neuronal RAGE by amphoterin in the developing nervous system might be, at least in part, one means by which the expression of RAGE is enhanced in that setting. Such findings would suggest contributory mechanisms by which amphoterin-mediated regulation of RAGE promotes outgrowth of neurites. Since examination of the promoter of RAGE revealed multiple putative binding elements for Sp1, a transcription factor essential for early embryonic development (19Marin M. Karis A. Visser P. Grosveld F. Philipsen S. Cell. 1997; 89: 619-628Abstract Full Text Full Text PDF PubMed Scopus (440) Google Scholar, 20Saffer J.D. Jackson S.P. Annarella M.B. Mol. Cell. Biol. 1991; 11: 2189-2199Crossref PubMed Scopus (483) Google Scholar), we tested the hypothesis that amphoterin-mediated activation of Sp1 might be important in regulation of RAGE expression in developing neurons. In this study, we demonstrate that interaction of amphoterin with RAGE on cultured neuroblastoma cells up-regulates transcription and translation of RAGE. The presence of functional Sp1-binding elements within the promoter of RAGE appears necessary for this outcome. These findings further suggest a role for amphoterin-RAGE interaction in the development of the nervous system and implicate RAGE as a gene whose regulation in the process of neuronal development is, at least in part, mediated by Sp1. SK-N-SH cells were obtained from American Type Tissue Corporation (Rockville, MD). SK-N-SH cells are derived from human neuroblastoma, and their neuronal origin has been extensively characterized (21Biedler J.L. Helson L. Spengler B.A. Cancer Res. 1973; 33: 2643-2652PubMed Google Scholar). Cells were grown in Medium 199 containing fetal bovine serum (FBS) (10%) (Life Technologies, Inc.) in the presence of penicillin (100 units/ml) and streptomycin (100 μg/ml) in humidified incubators containing CO2 (5%). Monospecific, polyclonal anti-human RAGE IgG was used to prepare an ELISA for the detection of human RAGE. SK-N-SH cells were cultured in Medium 199 containing FBS (1%) for 16 h and then completely starved of serum for another 24 h. At the end of that time, cells were exposed to the indicated amount of amphoterin and characterized as (16Hori O. Brett J. Slattery T. Cao R. Zhang J. Chen J. Nagashima M. Lundh E.R. Vijay S. Nitecki D. Morser J. Stern D. Schmidt A.M. J. Biol. Chem. 1995; 270: 25752-25761Abstract Full Text Full Text PDF PubMed Scopus (1007) Google or bovine serum albumin for 24 h. Cells were with and in containing and at for h and then at for The was and the The was then with of in to of of a 16 h at sites on the were in containing were then with anti-RAGE IgG μg/ml) and characterized as O. Brett J. Slattery T. Cao R. Zhang J. Chen J. Nagashima M. Lundh E.R. Vijay S. Nitecki D. Morser J. Stern D. Schmidt A.M. J. Biol. Chem. 1995; 270: 25752-25761Abstract Full Text Full Text PDF PubMed Scopus (1007) Google and sites of binding IgG with was human RAGE and from a expression system. The of detection in the assay is SK-N-SH cells were cultured in Medium 199 containing FBS (1%) for 16 h to of serum for another 24 h. At the end of that time, cells were exposed to either amphoterin μg/ml) or μg/ml) for 24 h. with cells were in containing further to cells were with polyclonal anti-human RAGE IgG for h at The was then in and cells were in IgG for h at with cells were in and to were nonimmune IgG for detection of cell-surface RAGE on SK-N-SH cells was as (16Hori O. Brett J. Slattery T. Cao R. Zhang J. Chen J. Nagashima M. Lundh E.R. Vijay S. Nitecki D. Morser J. Stern D. Schmidt A.M. J. Biol. Chem. 1995; 270: 25752-25761Abstract Full Text Full Text PDF PubMed Scopus (1007) Google Scholar). SK-N-SH cells were grown in Medium 199 for 16 h and then completely of serum for 24 h. that time, cells were then with the indicated of amphoterin or was from SK-N-SH cells The of was by at a was to electrophoretic on a was a and linked by a The was then with the human RAGE or glyceraldehyde-3-phosphate dehydrogenase with the system at with containing and then with containing at for the was to at for was used in studies to in the RAGE by transcription assay was as A.-M. Hori O. Chen J. Li J.F. Crandall J. Zhang J. Cao R. Yan S.D. Brett J. Stern D. J. Clin. Invest. 1995; 96: 1395-1403Crossref PubMed Scopus (809) Google Scholar, P. R. J. Biol. Chem. Full Text PDF PubMed Google Scholar). of SK-N-SH cells with amphoterin μg/ml) or μg/ml) for 24 h at cells were with and by were then in containing and for on and at for at The were with as and at for at The were and were in of reaction containing and of and at for h. The was as and of were in containing and to human RAGE and was at for h at for h in as containing for h and exposed to binding to the the human RAGE and were by with for at and was the and with a DNase I footprinting assay was to the binding of the putative Sp1-binding sites located within the 5′-flanking region of the human RAGE gene as revealed by analysis J. P. O. Res. PubMed Scopus Google Scholar). DNase I footprinting assay was with with Sp1 the region from to of the human RAGE gene was at the of the with by the DNase I of the was in binding containing μg/ml) in the presence or of the indicated amount of the human Sp1 and on for h. To this was then for at by DNase in for at The reaction was by the of to The reaction was then with one The was to and was for on for The were then to at for was then with and The was in to for and then on for at least The were then were then on at with an SK-N-SH cells were with and cultured in Medium 199 containing FBS (1%) for 16 h. of serum for another 24 cells were exposed to the indicated of amphoterin or for h at The was with and into were as E. P. M. W. Res. PubMed Scopus Google Scholar). the was to for at at The was then in and and on for to which was then The was for and the were to at for at which the were The was with and in and at with The was then to at for at and the was The of was then at an on the results of DNase I footprinting were J. P. O. Res. PubMed Scopus Google Scholar). of was with for were with of the indicated for at to with the for Sp1 were obtained from and were (Life for the and was with human Sp1, or for h to the of the indicated of the RAGE promoter and constructs of the RAGE promoter were by into the of from the 5′-flanking region of the human RAGE gene and to the of Sp1 a mutation these two binding sites was constructs for transfection were by system and luciferase of from the 5′-flanking region of the RAGE gene expressed in cultured SK-N-SH neuroblastoma was as indicated in the for of the Sp1-binding sites in the 5′-flanking region of the human RAGE gene by of the in of the The for the Sp1-binding sites are in luciferase from cultured SK-N-SH cells with the constructs either basal or conditions are for the of or of basal conditions are as basal conditions of the cells with the presence of or multiple on luciferase with that of in is as in the setting of with the presence of and or and not Sp1 on luciferase with was Sp1 was mutated either or in and luciferase significantly with that of in were Sp1 was on luciferase with Sp1 were and in luciferase was with that of Sp1-binding sites were mutated a in luciferase with that of was SK-N-SH cells were grown and as Transfection of the cells was (Life Technologies, Inc.) to the cells were the of to The for transfection was of and which as to of Cells were exposed to the of and for h. of the was and the for h. to studies of of the RAGE cells were with and in as that serum was for 24 h to further or was to the 24 h to was and in a as and results were for analysis was of expression of RAGE in the developing neurons of the central nervous system with of expression of amphoterin. In to interaction of amphoterin with RAGE, at least in part, mediated this we tested the of amphoterin to the expression of RAGE. Incubation of cultured neuroblastoma cells with amphoterin for 24 h revealed an in for RAGE with with The of amphoterin were mediated by interaction with cell-surface RAGE as demonstrated by of RAGE expression in the presence of anti-RAGE IgG In of neuroblastoma cells with nonimmune IgG to to amphoterin Incubation of SK-N-SH cells with anti-RAGE IgG on expression of RAGE The enhanced for RAGE by amphoterin was of for RAGE were in the presence of transcription assay revealed that amphoterin-mediated in for RAGE was to enhanced transcription with enhanced for RAGE in cultured neuroblastoma increased RAGE was by ELISA in a dose-dependent manner with cells with an in RAGE was in the presence of amphoterin. these were mediated by RAGE was demonstrated by of amphoterin-mediated expression of RAGE in the presence of anti-RAGE but not nonimmune IgG Incubation of SK-N-SH cells with anti-RAGE IgG was cell-surface RAGE in the presence of amphoterin was further by and not these indicated that amphoterin expression of RAGE in cultured neuroblastoma cells in a In studies, we identified two putative binding sites for the transcription factor Sp1 Sp1 and Sp1 in the promoter of RAGE analysis by for Sp1 with in the RAGE promoter. To the putative Sp1-binding sites within the promoter of RAGE were DNase I footprinting analysis was human Sp1 and RAGE promoter from to of Sp1 sites and was demonstrated were Sp1 and Sp1 of the these with the Sp1 by P.J. Mol. Biol. Scopus Google revealed or one In to amphoterin activation of Sp1 binding in neuroblastoma SK-N-SH cells were with or amphoterin in the presence of anti-RAGE IgG or nonimmune IgG. were and employing either Sp1 or Sp1-binding as indicated in A. with of neuroblastoma cells with with amphoterin resulted in increased Sp1 binding increases in Sp1 were significantly in the presence of anti-RAGE IgG but not in the presence of nonimmune IgG Incubation of cells with anti-RAGE IgG was for Sp1 was in in which a of the was in the presence of Sp1 and the of amphoterin in binding of Sp1 were activation of Sp1 binding was in the presence of In the presence of of further in activation of Sp1 binding was In to transcription were in amphoterin-mediated increases in RAGE expression, and were putative binding sites for these transcription were identified in the RAGE promoter by In was of activation of or and In Sp1 was in cells with these assays employing IgG indicated a shift of the Sp1 in cells and shift was in the presence of IgG or nonimmune IgG not Incubation of Sp1 with Sp1 resulted in of the In with or was these that Sp1 was an important transcription factor in amphoterin-mediated increases in RAGE expression in SK-N-SH In to delineate elements within the promoter of RAGE to a of 5′-deletion gene of the RAGE promoter was in a as indicated in 6. The constructs were then into neuroblastoma In the of amphoterin of cells to the constructs containing one or Sp1-binding sites to basal promoter and but not in a manner significantly which putative Sp1-binding these is not to Sp1-binding elements were central in basal promoter as is that elements within these contribute to basal or luciferase was transfection with containing of that in In the presence of luciferase was increased in cells with or with basal conditions in luciferase was increased in cells with or with basal conditions in in cells with in luciferase was of the cells with amphoterin In cells with or of the cells with anti-RAGE IgG to with amphoterin significantly the amphoterin-mediated in luciferase in with amphoterin such that luciferase was not significantly that basal conditions In of the cells with nonimmune IgG was not with anti-RAGE IgG did not affect basal luciferase not In to the of putative Sp1-binding elements in basal promoter and responsiveness to was as indicated in basal conditions with or multiple of Sp1-binding sites on luciferase with putative Sp1-binding in These the that Sp1-binding sites are not central for of basal RAGE promoter In the setting of with on luciferase from and or and of Sp1-binding sites was Sp1-binding was mutated in either or and luciferase in the presence of amphoterin in In the setting of and an was in the of that of Sp1 and Sp1 was luciferase was mutation of Sp1-binding sites identified within this of promoter responsiveness to amphoterin resulted These that Sp1 is important in promoter responsiveness to however, of Sp1-binding sites appears essential for promoter We have that amphoterin may interact with neuronal RAGE to increased transcription and translation of RAGE in an Sp1-dependent has been to be highly and expressed in embryonic by a expression is markedly attenuated. These findings, with the that amphoterin mediated neurite outgrowth in vitro, a role for amphoterin in the development of the nervous system J. H. J. Biol. Chem. 1991; Full Text PDF PubMed Google Scholar, H. R. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. R. J. J. H. J. Biol. Chem. 1991; Full Text PDF PubMed Google Scholar). Our identified RAGE as a neuronal for amphoterin as of to to RAGE, such as anti-RAGE F(ab′)2 or soluble RAGE, the extracellular two-thirds of RAGE, inhibited neurite outgrowth on with a role for the interaction of these two in neuronal in the developing nervous system, increased expression of RAGE was by and in in and neurons that co-localized with amphoterin (16Hori O. Brett J. Slattery T. Cao R. Zhang J. Chen J. Nagashima M. Lundh E.R. Vijay S. Nitecki D. Morser J. Stern D. Schmidt A.M. J. Biol. Chem. 1995; 270: 25752-25761Abstract Full Text Full Text PDF PubMed Scopus (1007) Google Scholar). to the in expression of amphoterin in the expression of RAGE in neurons in not completely is significantly In settings of enhanced neuronal however, such as in amyloid and oxidant stimuli increases in RAGE expression (10Yan S.D. Chen X. Fu J. Chen M. Zhu H. Roher A. Slattery T. Nagashima M. Morser J. Migheli A. Nawroth P. Godman G. Stern D. Schmidt A.M. Nature. 1996; 382: 685-691Crossref PubMed Scopus (1783) Google Scholar, 11Yan S.-D. Zhu H. Fu J. Yan S.-F. Roher A. Tourtellotte W. Rajavashisth T. Chen X. Stern D. Schmidt A.-M. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 5296-5301Crossref PubMed Scopus (400) Google Scholar). Our findings an important role for two functional binding elements for within the promoter of RAGE in this process (15Li J. Schmidt A.M. J. Biol. Chem. 1997; 272: 16498-16506Abstract Full Text Full Text PDF PubMed Scopus (442) Google Scholar). The studies implicate an important role for Sp1 in amphoterin-mediated increases in RAGE expression in cultured neuroblastoma Sp1, a transcription has been in of and responsiveness to perturbation A. T. J. Biol. Chem. 1995; 270: Full Text Full Text PDF PubMed Scopus Google Scholar, J.L. Chen X. Wang J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar, C. S. M. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar). Sp1 has been linked to the process of normal development. and (19Marin M. Karis A. Visser P. Grosveld F. Philipsen S. Cell. 1997; 89: 619-628Abstract Full Text Full Text PDF PubMed Scopus (440) Google recently that for of the gene for Sp1 demonstrate early in a range of was demonstrated in these the role of Sp1 in development of the nervous system. and J.D. Jackson S.P. Annarella M.B. Mol. Cell. Biol. 1991; 11: 2189-2199Crossref PubMed Scopus (483) Google that in early the for Sp1 was in Our findings not mechanisms in enhanced RAGE expression in developing neurons. as from the may with amphoterin to transcription and translation of RAGE. In this interaction sites for such as M. H. K. M. Res. 1992; PubMed Scopus Google and J. J. H. Res. 1992; PubMed Scopus Google Scholar), have been is that ligation of such sites may the of important in RAGE in the of multiple and to in expression of important in neuronal and In that is likely that a of and stimuli may contribute to increased expression of RAGE in developing neurons. studies suggest that regulation of RAGE in neurons is at least in part, by ligation by a polypeptide expressed in development. activation of Sp1 and binding to elements within the promoter of RAGE necessary for of RAGE the of Sp1 in embryonic these findings further suggest a role for amphoterin-RAGE interaction in the development of the nervous system. We the of Hori in the of these