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Pituitary

, Volume 9, Issue 2, pp 109–120 | Cite as

Phosphoproteomic analysis of the human pituitary

  • Sarka Beranova-Giorgianni
  • Yingxin Zhao
  • Dominic M. Desiderio
  • Francesco GiorgianniEmail author
Article

Abstract

The pituitary is the central endocrine gland that regulates the functions of various target organs in the human body. Because of the pivotal regulatory role of the pituitary, it is essential to define on a global scale the components of the pituitary protein machinery, including a comprehensive characterization of the post-translational modifications of the pituitary proteins. Of particular interest is the examination of the phosphorylation status of the pituitary in health and disease. Towards the goal of global profiling of pituitary protein phosphorylation, we report here the application of the in-gel IEF-LC-MS/MS approach to the study of the pituitary phosphoproteome. The analytical strategy combined isoelectric focusing in immobilized pH gradient strips with immobilized metal ion affinity chromatography and mass spectrometry. With this method, a total of 50 phosphorylation sites were characterized in 26 proteins. Because the investigation involved primary tissue, the findings provide a direct glimpse into the phosphoprotein machinery operating within the human pituitary tissue microenvironment.

Keywords

Human pituitary tissue Isoelectric focusing Mass spectrometry Phosphoproteome 

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References

  1. 1.
    Asa SL, Ezzat S (2002) The pathogenesis of pituitary tumours. Nat Rev Cancer 2:836–849PubMedCrossRefGoogle Scholar
  2. 2.
    Heaney AP, Melmed S (2004) Molecular targets in pituitary tumours. Nat Rev Cancer 4:285–295PubMedCrossRefGoogle Scholar
  3. 3.
    Noble ME, Endicott JA, Johnson LN (2004) Protein kinase inhibitors: Insights into drug design from structure. Sci. 303:1800–1805PubMedCrossRefGoogle Scholar
  4. 4.
    Beranova-Giorgianni S, Giorgianni F, Desiderio DM (2002) Analysis of the proteome in the human pituitary. Proteomics 2:534–542PubMedCrossRefGoogle Scholar
  5. 5.
    Zhan X, Desiderio DM (2003) A reference map of a human pituitary adenoma proteome. Proteomics 3:699–713PubMedCrossRefGoogle Scholar
  6. 6.
    Zhan X, Evans CO, Oyesiku NM, Desiderio DM (2003) Proteomics, transcriptomics analyses of secretagogin down-regulation in human non-functional pituitary adenomas. Pituitary 6:189–202PubMedCrossRefGoogle Scholar
  7. 7.
    Giorgianni F, Beranova-Giorgianni S, Desiderio DM (2004) Identification and characterization of phosphorylated proteins in the human pituitary. Proteomics 4:587–598PubMedCrossRefGoogle Scholar
  8. 8.
    Giorgianni F, Desiderio DM, Beranova-Giorgianni S (2003) Proteome analysis using isoelectric focusing in immobilized pH gradient gels followed by mass spectrometry. Electrophoresis 24:253–259PubMedCrossRefGoogle Scholar
  9. 9.
    Zhao Y, Giorgianni F, Desiderio DM, Fang B, Beranova-Giorgianni S (2005) Toward a global analysis of the human pituitary proteome by multiple gel-based technology. Anal Chem 77:5324–5331PubMedCrossRefGoogle Scholar
  10. 10.
    Gatlin CL, Kleemann GR, Hays LG, Link AJ, Yates JR III (1998) Protein identification at the low femtomole level from silver-stained gels using a new fritless electrospray interface for liquid chromatography-microspray and nanospray mass spectrometry. Anal Biochem 263:93–101PubMedCrossRefGoogle Scholar
  11. 11.
    Giorgianni F, Cappiello A, Beranova-Giorgianni S, Palma P, Trufelli H, Desiderio DM (2004) LC-MS/MS analysis of peptides with methanol as organic modifier: Improved limits of detection. Anal Chem 76:7028–7038PubMedCrossRefGoogle Scholar
  12. 12.
    Obenauer JC, Cantley LC, Yaffe MB (2003) Scansite 2.0: Proteome-wide prediction of cell signaling interactions using short sequence motifs. Nucleic Acids Res 31:3635–3641PubMedCrossRefGoogle Scholar
  13. 13.
    Olsen JV, Ong SE, Mann M (2004) Trypsin cleaves exclusively C-terminal to arginine and lysine residues. Mol Cell Proteomics 3:608–614PubMedCrossRefGoogle Scholar
  14. 14.
    Ballif BA, Villen J, Beausoleil SA, Schwartz D, Gygi SP (2004) Phosphoproteomic analysis of the developing mouse brain. Mol Cell Proteomics 3:1093–1101PubMedCrossRefGoogle Scholar
  15. 15.
    Schwartz JC, Senko MW, Syka JE (2002) A two-dimensional quadrupole ion trap mass spectrometer. J Am Soc Mass Spectrom 13:659–669PubMedCrossRefGoogle Scholar
  16. 16.
    Harper ME, Barrera-Saldana HA, Saunders GF (1982) Chromosomal localization of the human placental lactogen-growth hormone gene cluster to 17q22-24. Am J Hum Genet 34:227–234PubMedGoogle Scholar
  17. 17.
    Frohman LA, Kineman RD, Kamegai J, Park S, Teixeira LT, Coschigano KT, Kopchic JJ (2000) Secretagogues and the somatotrope: Signaling and proliferation. Recent Prog Horm Res 55:269–290PubMedGoogle Scholar
  18. 18.
    Kojima M, Hosoda H, Matsuo H, Kangawa K (2001) Ghrelin: Discovery of the natural endogenous ligand for the growth hormone secretagogue receptor. Trends Endocrinol Metab 12:118–122PubMedCrossRefGoogle Scholar
  19. 19.
    Kato Y, Murakami Y, Sohmiya M, Nishiki M (2002) Regulation of human growth hormone secretion and its disorders. Intern Med 41:7–13PubMedGoogle Scholar
  20. 20.
    Melmed S (2003) Mechanisms for pituitary tumorigenesis: The plastic pituitary. J Clin Invest 112:1603–1618PubMedCrossRefGoogle Scholar
  21. 21.
    Binder G (2002) Isolated growth hormone deficiency, the GH-1 gene: Update 2002. Horm Res 58 (Suppl 3):2–6PubMedCrossRefGoogle Scholar
  22. 22.
    Yoshizato H, Tanaka M, Nakai N, Nakao N, Nakashima K (2004) Growth hormone (GH)-stimulated insulin-like growth factor I gene expression is mediated by a tyrosine phosphorylation pathway depending on C-terminal region of human GH receptor in human GH receptor-expressing Ba/F3 cells. Endocrinology 145:214–220PubMedCrossRefGoogle Scholar
  23. 23.
    Fryburg DA, Louard RJ, Gerow KE, Gelfand RA, Barrett EJ (1992) Growth hormone stimulates skeletal muscle protein synthesis and antagonizes insulin’s antiproteolytic action in humans. Diabetes 41:424–429PubMedGoogle Scholar
  24. 24.
    Florini JR, Ewton DZ, Coolican SA (1996) Growth hormone and the insulin-like growth factor system in myogenesis. Endocr Rev 17:481–517PubMedCrossRefGoogle Scholar
  25. 25.
    Ivashkiv LB (1995) Cytokines, STATs: How can signals achieve specificity?. Immunity 3:1–4PubMedCrossRefGoogle Scholar
  26. 26.
    Silva CM, Lu H, Day RN (1996) Characterization and cloning of STAT5 from IM-9 cells and its activation by growth hormone. Mol Endocrinol 10:508–518PubMedCrossRefGoogle Scholar
  27. 27.
    Sotiropoulos A, Moutoussamy S, Renaudie F, Clauss M, Kayser C, Gouilleux F, Kelly PA, Finidori J (1996) Differential activation of Stat3 and Stat5 by distinct regions of the growth hormone receptor. Mol Endocrinol 10:998–1009PubMedCrossRefGoogle Scholar
  28. 28.
    de Vos AM, Ultsch M, Kossiakoff AA (1992) Human growth hormone and extracellular domain of its receptor: Crystal structure of the complex. Science 255:306–312PubMedGoogle Scholar
  29. 29.
    Lewis UJ, Sinha YN, Lewis GP (2000) Structure and properties of members of the hGH family: A review. Endocr J 47(Suppl):S1–S8PubMedGoogle Scholar
  30. 30.
    Baumann G (1999) Growth hormone heterogeneity in human pituitary, plasma. Horm Res 51(Suppl 1):2–6PubMedCrossRefGoogle Scholar
  31. 31.
    Liberti JP, Antoni BA, Chlebowski JF (1985) Naturally-occurring pituitary growth hormone is phosphorylated. Biochem Biophys Res Commun 128:713–720PubMedGoogle Scholar
  32. 32.
    Liberti JP, Joshi GS (1986) Synthesis and secretion of phosphorylated growth hormone by rat pituitary glands in vitro. Biochem Biophys Res Commun 137:806–812PubMedCrossRefGoogle Scholar
  33. 33.
    Aramburo C, Montiel JL, Proudman JA, Berghman LR, Scanes CG (1992) Phosphorylation of prolactin and growth hormone. J Mol Endocrinol 8:183–191PubMedCrossRefGoogle Scholar
  34. 34.
    Bennett HP, Browne CA, Solomon S (1981) Biosynthesis of phosphorylated forms of corticotropin-related peptides. Proc Natl Acad Sci USA 78:4713–4717PubMedCrossRefGoogle Scholar
  35. 35.
    Eipper BA, Mains RE (1982) Phosphorylation of pro-adrenocorticotropin/endorphin-derived peptides. J Biol Chem 257:4907–4915PubMedGoogle Scholar
  36. 36.
    Huttner WB, Gerdes HH, Rosa P (1991) The granin (chromogranin/secretogranin) family. Trends Biochem Sci 16:27–30PubMedCrossRefGoogle Scholar
  37. 37.
    Aunis D, Metz-Boutigue MH (2000) Chromogranins: Current concepts. Structural and functional aspects. Adv Exp Med Biol 482:21–38PubMedCrossRefGoogle Scholar
  38. 38.
    Gadroy P, Stridsberg M, Capon C, Michalski JC, Strub JM, van Dorsselaer A, Aunis D, Metz-Boutigue MH (1998) Phosphorylation and O-glycosylation sites of human chromogranin A (CGA79-439) from urine of patients with carcinoid tumors. J Biol Chem 273:34087–34097PubMedCrossRefGoogle Scholar
  39. 39.
    Aunis D, Metz-Boutigue MH (2000) Chromogranins: Current concepts. Structural and functional aspects. Adv Exp Med Biol 482:21–38PubMedGoogle Scholar
  40. 40.
    Dahma H, Gourlet P, Vandermeers A, Vandermeers-Piret MC, Robberecht P (2001) Evidence that the chromogranin B fragment 368-417 extracted from a pheochromocytoma is phosphorylated. Peptides 22:1491–1499PubMedCrossRefGoogle Scholar
  41. 41.
    Gasnier C, Lugardon K, Ruh O, Strub JM, Aunis D, Metz-Boutigue MH (2004) Characterization and location of post-translational modifications on chromogranin B from bovine adrenal medullary chromaffin granules. Proteomics 4:1789–1801PubMedCrossRefGoogle Scholar
  42. 42.
    Blom N, Gammeltoft S, Brunak S (1999) Sequence and structure-based prediction of eukaryotic protein phosphorylation sites. J Mol Biol 294:1351–1362PubMedCrossRefGoogle Scholar
  43. 43.
    Gerdes HH, Glombik MM (2000) Signal-mediated sorting of chromogranins to secretory granules. Adv Exp Med Biol 482:41–54PubMedGoogle Scholar
  44. 44.
    Kromer A, Glombik MM, Huttner WB, Gerdes HH (1998) Essential role of the disulfide-bonded loop of chromogranin B for sorting to secretory granules is revealed by expression of a deletion mutant in the absence of endogenous granin synthesis. J Cell Biol 140:1331–1346PubMedCrossRefGoogle Scholar
  45. 45.
    Gorr SU, Shioi J, Cohn DV (1989) Interaction of calcium with porcine adrenal chromogranin A (secretory protein-I) and chromogranin B (secretogranin I). Am J Physiol 257:E247–E254PubMedGoogle Scholar
  46. 46.
    Wynick D, Small CJ, Bacon A, Holmes FE, Norman M, Ormandy CJ, Kilic E, Kerr NC, Ghatei M, Talamantes F, Bloom SR, Pachnis V (1998) Galanin regulates prolactin release and lactotroph proliferation. Proc Natl Acad Sci USA 95:12671–12676PubMedCrossRefGoogle Scholar
  47. 47.
    Bacon A, Holmes FE, Small CJ, Ghatei M, Mahoney S, Bloom S, Wynick D (2002) Transgenic over-expression of galanin in injured primary sensory neurons. Neuroreport 13:2129–2132PubMedCrossRefGoogle Scholar
  48. 48.
    Wynick D, Small CJ, Bloom SR, Pachnis V (1998) Targeted disruption of the murine galanin gene. Ann NY Acad Sci 863:22–47PubMedCrossRefGoogle Scholar
  49. 49.
    Mazarati AM (2004) Galanin and galanin receptors in epilepsy. Neuropeptides 38:331–343PubMedCrossRefGoogle Scholar
  50. 50.
    She YM, Huang YW, Zhang L, Trimble WS (2004) Septin 2 phosphorylation: Theoretical and mass spectrometric evidence for the existence of a single phosphorylation site in vivo. Rapid Commun Mass Spectrom 18:1123–1130PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science + Business Media, LLC 2006

Authors and Affiliations

  • Sarka Beranova-Giorgianni
    • 1
    • 2
    • 3
  • Yingxin Zhao
    • 2
  • Dominic M. Desiderio
    • 1
    • 3
  • Francesco Giorgianni
    • 1
    • 3
    Email author
  1. 1.Charles B. Stout Neuroscience Mass Spectrometry LaboratoryThe University of Tennessee Health Science CenterMemphisUSA
  2. 2.Department of Pharmaceutical SciencesThe University of Tennessee Health Science CenterMemphisUSA
  3. 3.Department of NeurologyThe University of Tennessee Health Science Center MemphisUSA

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