Skip to main content

Advertisement

SpringerLink
Log in

A Selective Human Bombesin Receptor Subtype-3 Peptide Agonist Mediates CREB Phosphorylation and Transactivation

  • Published:
Journal of Molecular Neuroscience Aims and scope Submit manuscript

Abstract

The native ligand for the G protein-coupled bombesin receptor subtype-3 (BRS-3) has currently not been identified. Studies in mice showed robust BRS-3 expression in the hypothalamic satiety centers, and genetic receptor inactivation resulted in obesity, diabetes, and hypertension. BRS-3 was also detected in normal human pancreatic islet cells suggesting a critical role of BRS-3 in regulating energy metabolism and satiety via central and peripheral mechanisms of action. The cyclic AMP response element binding protein (CREB) is a main regulator of pancreatic β-cell gene expression required for glucose homeostasis and islet cell survival, and hypothalamic regulation of satiety. Therefore, in this study we examined whether agonist-dependent hBRS-3 stimulation mediates CREB activation. A selective hBRS-3 peptide agonist and two non-selective hBRS-3 peptide agonists were used to activate ectopically expressed hBRS-3. Stimulation with hBRS-3 peptide agonists resulted in transient calcium mobilization, whereby the selective peptide agonist acted exclusively via hBRS-3 but not through the gastrin-releasing peptide receptor (GRP-R). A selective high-affinity GRP-R antagonist did not inhibit hBRS-3-mediated calcium signals. We also found time-dependent CREB phosphorylation in response to the selective hBRS-3 activation, which was abrogated by pretreatment with protein kinase A and protein kinase C inhibitors. Human BRS-3 agonists also stimulated CREB transactivation and resulted in modest increases of CRE-dependent gene transcription. These changes were significantly reduced after pretreatment with inhibitors of PKA, PKC, and MEK-1. Thus, our results suggest that hBRS-3 agonist-dependent signaling mediates CREB phosphorylation and transactivation through partially PKA, PKC, and MEK-1 pathways.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Abbreviations

Bn:

Bombesin

Bn-r:

Bombesin receptors

BRS-3:

Bombesin receptor subtype-3

GRP-R:

Gastrin-releasing peptide receptor

CREB:

Cyclic AMP response element binding protein

DMEM:

Dulbecco’s minimum essential medium

MAPK:

Mitogen-activated protein kinase

MEK-1:

Mitogen-activated protein/extracellular regulated kinase kinase

NMB-R:

Neuromedin B receptor

PKA:

Protein kinase A

PKC:

Protein kinase C

References

  • Arias J, Alberts AS, Brindle P et al (1994) Activation of cAMP and mitogen responsive genes relies on a common nuclear factor. Nature 370(6486):226–229

    Article  PubMed  CAS  Google Scholar 

  • Branson R, Potoczna N, Kral JG, Lentes KU, Hoehe MR, Horber FF (2003) Binge eating as a major phenotype of melanocortin 4 receptor gene mutations. N Engl J Med 348(12):1096–1103

    Article  PubMed  CAS  Google Scholar 

  • Brindle P, Linke S, Montminy M (1993) Protein-kinase-A-dependent activator in transcription factor CREB reveals new role for CREM repressors. Nature 364(6440):821–824

    Article  PubMed  CAS  Google Scholar 

  • Coll AP (2010) Treating obesity? It’s in the bag! Cell Metab 11(2):95–96

    Article  PubMed  CAS  Google Scholar 

  • Dalle S, Longuet C, Costes S et al (2004) Glucagon promotes cAMP-response element-binding protein phosphorylation via activation of ERK1/2 in MIN6 cell line and isolated islets of Langerhans. J Biol Chem 279(19):20345–20355

    Article  PubMed  CAS  Google Scholar 

  • Eckert B, Schwaninger M, Knepel W (1996) Calcium-mobilizing insulin secretagogues stimulate transcription that is directed by the cyclic adenosine 3′,5′-monophosphate/calcium response element in a pancreatic islet beta-cell line. Endocrinology 137(1):225–233

    Article  PubMed  CAS  Google Scholar 

  • Farooqi IS, Keogh JM, Yeo GS, Lank EJ, Cheetham T, O’Rahilly S (2003) Clinical spectrum of obesity and mutations in the melanocortin 4 receptor gene. N Engl J Med 348(12):1085–1095

    Article  PubMed  CAS  Google Scholar 

  • Fathi Z, Corjay MH, Shapira H et al (1993) BRS-3: a novel bombesin receptor subtype selectively expressed in testis and lung carcinoma cells. J Biol Chem 268(8):5979–5984

    PubMed  CAS  Google Scholar 

  • Fathi Z, Way JW, Corjay MH, Viallet J, Sausville EA, Battey JF (1996) Bombesin receptor structure and expression in human lung carcinoma cell lines. J Cell Biochem Suppl 24:237–246

    Article  PubMed  CAS  Google Scholar 

  • Fleischmann A, Laderach U, Friess H, Buechler MW, Reubi JC (2000) Bombesin receptors in distinct tissue compartments of human pancreatic diseases. Lab Invest 80(12):1807–1817

    Article  PubMed  CAS  Google Scholar 

  • Gorbulev V, Akhundova A, Buchner H, Fahrenholz F (1992) Molecular cloning of a new bombesin receptor subtype expressed in uterus during pregnancy. Eur J Biochem 208(2):405–410

    Article  PubMed  CAS  Google Scholar 

  • Gorbulev V, Akhundova A, Grzeschik KH, Fahrenholz F (1994) Organization and chromosomal localization of the gene for the human bombesin receptor subtype expressed in pregnant uterus. FEBS Lett 340(3):260–264

    Article  PubMed  CAS  Google Scholar 

  • Guan XM, Chen H, Dobbelaar PH et al (2010) Regulation of energy homeostasis by bombesin receptor subtype-3: selective receptor agonists for the treatment of obesity. Cell Metab 11(2):101–112

    Article  PubMed  CAS  Google Scholar 

  • Guan XM, Metzger JM, Yang L et al (2011) Antiobesity effect of MK-5046, a novel bombesin receptor subtype-3 agonist. J Pharmacol Exp Ther 336(2):356–364

    Article  PubMed  CAS  Google Scholar 

  • Guo C, Guzzo PR, Hadden M et al (2010) Synthesis of 7-benzyl-5-(piperidin-1-yl)-6,7,8,9-tetrahydro-3H-pyrazolo[3,4-c][2,7]naph thyridin-1-ylamine and its analogs as bombesin receptor subtype-3 agonists. Bioorg Med Chem Lett 20(9):2785–2789

    Article  PubMed  CAS  Google Scholar 

  • Hadden M, Goodman A, Guo C et al (2010) Synthesis and SAR of heterocyclic carboxylic acid isosteres based on 2-biarylethylimidazole as bombesin receptor subtype-3 (BRS-3) agonists for the treatment of obesity. Bioorg Med Chem Lett 20(9):2912–2915

    Article  PubMed  CAS  Google Scholar 

  • He S, Dobbelaar PH, Liu J et al (2010) Discovery of substituted biphenyl imidazoles as potent, bioavailable bombesin receptor subtype-3 agonists. Bioorg Med Chem Lett 20(6):1913–1917

    Article  PubMed  CAS  Google Scholar 

  • Hou X, Wei L, Harada A, Tatamoto K (2006) Activation of bombesin receptor subtype-3 stimulates adhesion of lung cancer cells. Lung Cancer 54(2):143–148

    Article  PubMed  Google Scholar 

  • Huszar D, Lynch CA, Fairchild-Huntress V et al (1997) Targeted disruption of the melanocortin-4 receptor results in obesity in mice. Cell 88(1):131–141

    Article  PubMed  CAS  Google Scholar 

  • Jennings CA, Harrison DC, Maycox PR, Crook B, Smart D, Hervieu GJ (2003) The distribution of the orphan bombesin receptor subtype-3 in the rat CNS. Neuroscience 120(2):309–324

    Article  PubMed  CAS  Google Scholar 

  • Jensen RT, Battey JF, Spindel ER, Benya RV (2008) International Union of Pharmacology. LXVIII. Mammalian bombesin receptors: nomenclature, distribution, pharmacology, signaling, and functions in normal and disease states. Pharmacol Rev 60(1):1–42

    Article  PubMed  CAS  Google Scholar 

  • Jhala US, Canettieri G, Screaton RA et al (2003) cAMP promotes pancreatic beta-cell survival via CREB-mediated induction of IRS2. Genes Dev 17(13):1575–1580

    Article  PubMed  CAS  Google Scholar 

  • Kuiper P., Verspaget H.W., Biemond I., et al. (2010) Expression and ligand binding of bombesin receptors in pulmonary and intestinal carcinoids. The role of bombesin in carcinoids. J Endocrinol Invest [Epub ahead of print].

  • Kwok RP, Lundblad JR, Chrivia JC et al (1994) Nuclear protein CBP is a coactivator for the transcription factor CREB. Nature 370(6486):223–226

    Article  PubMed  CAS  Google Scholar 

  • Ladenheim EE, Hamilton NL, Behles RR et al (2008) Factors contributing to obesity in bombesin receptor subtype-3-deficient mice. Endocrinology 149(3):971–978

    Article  PubMed  CAS  Google Scholar 

  • Lee JY, Lee JH, Kim DG, Jahng JW (2003) Dexamethasone blocks the refeeding-induced phosphorylation of cAMP response element-binding protein in the rat hypothalamus. Neurosci Lett 344(2):107–111

    Article  PubMed  CAS  Google Scholar 

  • List JF, Habener JF (2003) Defective melanocortin 4 receptors in hyperphagia and morbid obesity. N Engl J Med 348(12):1160–1163

    Article  PubMed  Google Scholar 

  • Liu J, Lao ZJ, Zhang J et al (2002a) Molecular basis of the pharmacological difference between rat and human bombesin receptor subtype-3 (BRS-3). Biochemistry 41(28):8954–8960

    Article  PubMed  CAS  Google Scholar 

  • Liu W, Chin-Chance C, Lee EJ, Lowe WL Jr (2002b) Activation of phosphatidylinositol 3-kinase contributes to insulin-like growth factor I-mediated inhibition of pancreatic beta-cell death. Endocrinology 143(10):3802–3812

    Article  PubMed  CAS  Google Scholar 

  • Liu J, He S, Jian T et al (2010) Synthesis and SAR of derivatives based on 2-biarylethylimidazole as bombesin receptor subtype-3 (BRS-3) agonists for the treatment of obesity. Bioorg Med Chem Lett 20(7):2074–2077

    Article  PubMed  CAS  Google Scholar 

  • Lo MM, Chobanian HR, Palyha O et al (2011) Pyridinesulfonylureas and pyridinesulfonamides as selective bombesin receptor subtype-3 (BRS-3) agonists. Bioorg Med Chem Lett 21(7):2040–2043

    Article  PubMed  CAS  Google Scholar 

  • Maekawa F, Quah HM, Tanaka K, Ohki-Hamazaki H (2004) Leptin resistance and enhancement of feeding facilitation by melanin-concentrating hormone in mice lacking bombesin receptor subtype-3. Diabetes 53(3):570–576

    Article  PubMed  CAS  Google Scholar 

  • Mantey SA, Weber HC, Sainz E et al (1997) Discovery of a high affinity radioligand for the human orphan receptor, bombesin receptor subtype 3, which demonstrates that it has a unique pharmacology compared with other mammalian bombesin receptors. J Biol Chem 272(41):26062–26071

    Article  PubMed  CAS  Google Scholar 

  • Mantey SA, Coy DH, Pradhan TK et al (2001) Rational design of a peptide agonist that interacts selectively with the orphan receptor, bombesin receptor subtype 3. J Biol Chem 276(12):9219–9229

    Article  PubMed  CAS  Google Scholar 

  • Mantey SA, Entsuah L, Coy DH et al (2002) Identification of a selective agonist for the orphan receptor, BRS-3. Gastroenterology 122(4, Supplement):A-135

    Google Scholar 

  • Mantey SA, Coy DH, Entsuah LK, Jensen RT (2004) Development of bombesin analogs with conformationally restricted amino acid substitutions with enhanced selectivity for the orphan receptor human bombesin receptor subtype 3. J Pharmacol Exp Ther 310(3):1161–1170

    Article  PubMed  CAS  Google Scholar 

  • Matsumoto K, Yamada K, Wada E, Hasegawa T, Usui Y, Wada K (2003) Bombesin receptor subtype-3 modulates plasma insulin concentration. Peptides 24(1):83–90

    Article  PubMed  CAS  Google Scholar 

  • Mayr B, Montminy M (2001) Transcriptional regulation by the phosphorylation-dependent factor CREB. Nat Rev Mol Cell Biol 2(8):599–609

    Article  PubMed  CAS  Google Scholar 

  • Metzger JM, Gagen K, Raustad KA et al (2010) Body temperature as a mouse pharmacodynamic response to bombesin receptor subtype-3 agonists and other potential obesity treatments. Am J Physiol Endocrinol Metab 299(5):E816–E824

    Article  PubMed  CAS  Google Scholar 

  • Must A, Spadano J, Coakley EH, Field AE, Colditz G, Dietz WH (1999) The disease burden associated with overweight and obesity. JAMA 282(16):1523–1529

    Article  PubMed  CAS  Google Scholar 

  • Nakamichi Y, Wada E, Aoki K et al (2004) Functions of pancreatic beta cells and adipocytes in bombesin receptor subtype-3-deficient mice. Biochem Biophys Res Commun 318(3):698–703

    Article  PubMed  CAS  Google Scholar 

  • Oetjen E, Diedrich T, Eggers A, Eckert B, Knepel W (1994) Distinct properties of the cAMP-responsive element of the rat insulin I gene. J Biol Chem 269(43):27036–27044

    PubMed  CAS  Google Scholar 

  • Ohki-Hamazaki H, Watase K, Yamamoto K et al (1997) Mice lacking bombesin receptor subtype-3 develop metabolic defects and obesity. Nature 390(6656):165–169

    Article  PubMed  CAS  Google Scholar 

  • Porcher C, Juhem A, Peinnequin A, Bonaz B (2005) Bombesin receptor subtype-3 is expressed by the enteric nervous system and by interstitial cells of Cajal in the rat gastrointestinal tract. Cell Tissue Res 320(1):21–31

    Article  PubMed  CAS  Google Scholar 

  • Pradhan TK, Katsuno T, Taylor JE et al (1998) Identification of a unique ligand which has high affinity for all four bombesin receptor subtypes. Eur J Pharmacol 343(2–3):275–287

    Article  PubMed  CAS  Google Scholar 

  • Qu X, Xiao D, Weber HC (2002) Human gastrin-releasing peptide receptor mediates sustained CREB phosphorylation and transactivation in HuTu 80 duodenal cancer cells. FEBS Lett 527(1–3):109–113

    Article  PubMed  CAS  Google Scholar 

  • Reubi JC, Wenger S, Schmuckli-Maurer J, Schaer JC, Gugger M (2002) Bombesin receptor subtypes in human cancers: detection with the universal radioligand (125)I-[D-TYR(6), beta-ALA(11), PHE(13), NLE(14)] bombesin(6–14). Clin Cancer Res 8(4):1139–1146

    PubMed  CAS  Google Scholar 

  • Ryan RR, Weber HC, Hou W et al (1998a) Ability of various bombesin receptor agonists and antagonists to alter intracellular signaling of the human orphan receptor BRS-3. J Biol Chem 273(22):13613–13624

    Article  PubMed  CAS  Google Scholar 

  • Ryan RR, Weber HC, Mantey SA et al (1998b) Pharmacology and intracellular signaling mechanisms of the native human orphan receptor BRS-3 in lung cancer cells. J Pharmacol Exp Ther 287(1):366–380

    PubMed  CAS  Google Scholar 

  • Sancho V, Moody TW, Mantey SA et al (2010) Pharmacology of putative selective hBRS-3 receptor agonists for human bombesin receptors (BnR): affinities, potencies and selectivity in multiple native and BnR transfected cells. Peptides 31(8):1569–1578

    Article  PubMed  CAS  Google Scholar 

  • Schulz S, Rocken C (2006) Immunohistochemical detection of bombesin receptor subtypes GRP-R and BRS-3 in human tumors using novel antipeptide antibodies. Virchows Arch 449(4):421–427

    Article  PubMed  CAS  Google Scholar 

  • Sheriff S, Chance WT, Fischer JE, Balasubramaniam A (1997) Neuropeptide Y treatment and food deprivation increase cyclic AMP response element-binding in rat hypothalamus. Mol Pharmacol 51(4):597–604

    PubMed  CAS  Google Scholar 

  • Shimizu-Albergine M, Ippolito DL, Beavo JA (2001) Downregulation of fasting-induced cAMP response element-mediated gene induction by leptin in neuropeptide Y neurons of the arcuate nucleus. J Neurosci 21(4):1238–1246

    PubMed  CAS  Google Scholar 

  • Spiegelman BM, Flier JS (2001) Obesity and the regulation of energy balance. Cell 104(4):531–543

    Article  PubMed  CAS  Google Scholar 

  • Tan YR, Qi MM, Qin XQ et al (2006) Wound repair and proliferation of bronchial epithelial cells enhanced by bombesin receptor subtype 3 activation. Peptides 27(7):1852–1858

    Article  PubMed  CAS  Google Scholar 

  • Tan YR, Qin XQ, Xiang Y et al (2007) PPARalpha and AP-2alpha regulate bombesin receptor subtype 3 expression in ozone-stressed bronchial epithelial cells. Biochem J 405(1):131–137

    PubMed  CAS  Google Scholar 

  • Wang Y, Zhang M, Tan Y et al (2007) BRS-3 activation transforms the effect of human bronchial epithelial cells from PGE2 mediated inhibition to TGF-beta1 dependent promotion on proliferation and collagen synthesis of lung fibroblasts. Cell Biol Int 31(12):1495–1500

    Article  PubMed  CAS  Google Scholar 

  • Weber HC, Hampton LL, Jensen RT, Battey JF (1998) Structure and chromosomal localization of the mouse bombesin receptor subtype 3 gene. Gene 211(1):125–131

    Article  PubMed  CAS  Google Scholar 

  • Weber HC, Walters J, Leyton J et al (2001) A bombesin receptor subtype-3 peptide increases nuclear oncogene expression in a MEK-1 dependent manner in human lung cancer cells. Eur J Pharmacol 412(1):13–20

    Article  PubMed  CAS  Google Scholar 

  • Wu JM, Nitecki DE, Biancalana S, Feldman RI (1996) Discovery of high affinity bombesin receptor subtype 3 agonists. Mol Pharmacol 50(5):1355–1363

    PubMed  CAS  Google Scholar 

  • Xiao D, Wang J, Hampton LL, Weber HC (2001) The human gastrin-releasing peptide receptor gene structure, its tissue expression and promoter. Gene 264(1):95–103

    Article  PubMed  CAS  Google Scholar 

  • Yamada K, Wada E, Imaki J, Ohki-Hamazaki H, Wada K (1999) Hyperresponsiveness to palatable and aversive taste stimuli in genetically obese (bombesin receptor subtype-3-deficient) mice. Physiol Behav 66(5):863–867

    Article  PubMed  CAS  Google Scholar 

  • Yamada K, Ohki-Hamazaki H, Wada K (2000) Differential effects of social isolation upon body weight, food consumption, and responsiveness to novel and social environment in bombesin receptor subtype-3 (BRS-3) deficient mice. Physiol Behav 68(4):555–561

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

We thank Dr. G. Yaney and Jamie Fairbanks (Boston University) for their invaluable assistance with the intracellular calcium measurements. Grant Support: This work was supported in part by a National Cancer Center Fellowship (to X.Q.) and NIH grant DK46200.

Author information

Authors and Affiliations

  1. Department of Physiology, Xiangya Medicine School, Central South University, Changsha, Hunan, 410078, People’s Republic of China

    Xiaoqun Qin & Xiangping Qu

  2. Section of Gastroenterology, Boston University School of Medicine, 650 Albany Street, EBRC, Room 515, Boston, MA, 02118-2518, USA

    Xiangping Qu

  3. Peptide Research Laboratories, Tulane University Health Sciences Center, New Orleans, LA, USA

    David Coy

  4. Section of Gastroenterology and Department of Pathology and Laboratory Medicine, Boston University School of Medicine, 650 Albany Street, EBRC, Room 515, Boston, MA, 02118-2518, USA

    H. Christian Weber

Authors
  1. Xiaoqun Qin
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xiangping Qu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. David Coy
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. H. Christian Weber
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to H. Christian Weber.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Qin, X., Qu, X., Coy, D. et al. A Selective Human Bombesin Receptor Subtype-3 Peptide Agonist Mediates CREB Phosphorylation and Transactivation. J Mol Neurosci 46, 88–99 (2012). https://doi.org/10.1007/s12031-011-9675-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12031-011-9675-3

Keywords

Search

Navigation