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. 2013 Jun 6;4(7):520–528. doi: 10.1007/s13238-013-3005-1

Angiotensin IV upregulates the activity of protein phosphatase 1α in Neura-2A cells

Dan Wang 13005,23005, Peng Xue 13005, Xiu Lan Chen 13005, Zhen Sheng Xie 13005, Fu Quan Yang 13005, Li Zheng 13005,✉, Tao Xu 13005,✉
PMCID: PMC4875510  PMID: 23744339

Abstract

The peptide angiotensin IV (Ang IV) is a derivative of angiotensin II. While insulin regulated amino peptidase (IRAP) has been proposed as a potential receptor for Ang IV, the signalling pathways of Ang IV through IRAP remain elusive. We applied high-resolution mass spectrometry to perform a systemic quantitative phosphoproteome of Neura-2A (N2A) cells treated with and without Ang IV using sta ble-isotope labeling by amino acids in cell culture (SILAC), and identified a reduction in the phosphorylation of a major Ser/Thr protein phosphorylase 1 (PP1) upon Ang IV treatment. In addition, spinophilin (spn), a PP1 regulatory protein that plays important functions in the neural system, was expressed at higher levels. Immunoblotting revealed decreased phosphorylation of p70S6 kinase (p70S6K) and the major cell cycle modulator retinoblastoma protein (pRB). These changes are consistent with an observed decrease in cell proliferation. Taken together, our study suggests that Ang IV functions via regulating the activity of PP1.

Electronic Supplementary Material

Supplementary material is available for this article at 10.1007/s13238-013-3005-1 and is accessible for authorized users.

Keywords: angiotensin IV, PP1α, cell proliferation, p70S6 kinase, spinophilin

Electronic supplementary material

13238_2013_3005_MOESM1_ESM.pdf (207.1KB, pdf)

Supplementary material, approximately 203 KB.

Footnotes

Electronic Supplementary Material

Supplementary material is available for this article at 10.1007/s13238-013-3005-1 and is accessible for authorized users.

Contributor Information

Li Zheng, Email: zhengli@moon.ibp.ac.cn.

Tao Xu, Email: xutao@ibp.ac.cn.

References

  1. Akiyama T, Ohuchi T, Sumida S, Matsumoto K, Toyoshima K. Phosphorylation of the retinoblastoma protein by cdk2. Proc Natl Acad Sci U S A. 1992;89:7900–7904. doi: 10.1073/pnas.89.17.7900. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Albiston AL, McDowall SG, Matsacos D, Sim P, Clune E, Mustafa T, Lee J, Mendelsohn FA, Simpson RJ, Connolly LM, et al. Evidence that the angiotensin IV (AT(4)) receptor is the enzyme insulin-regulated aminopeptidase. J Biol Chem. 2001;276:48623–48626. doi: 10.1074/jbc.C100512200. [DOI] [PubMed] [Google Scholar]
  3. Allen PB, Ouimet CC, Greengard P. Spinophilin, a novel protein phosphatase 1 binding protein localized to dendritic spines. Proc Natl Acad Sci U S A. 1997;94:9956–9961. doi: 10.1073/pnas.94.18.9956. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Berndt N. Protein dephosphorylation and the intracellular control of the cell number. Front Biosci. 1999;4:D22–42. doi: 10.2741/Berndt. [DOI] [PubMed] [Google Scholar]
  5. Berndt N, Dohadwala M, Liu CWY. Constitutively active protein phosphatase 1α causes Rb-dependent G1 arrest in human cancer cells. Curr Biol. 1997;7:375–386. doi: 10.1016/S0960-9822(06)00185-0. [DOI] [PubMed] [Google Scholar]
  6. Bettoun DJ, Buck DW, II, Lu J, Khalifa B, Chin WW, Nagpal S. A vitamin D receptor-Ser/Thr phosphatase-p70 S6 kinase complex and modulation of its enzymatic activities by the ligand. J Biol Chem. 2002;277:24847–24850. doi: 10.1074/jbc.C200187200. [DOI] [PubMed] [Google Scholar]
  7. Bialojan C, Takai A. Inhibitory effect of a marine-sponge toxin, okadaic acid, on protein phosphatases. Specificity and kinetics. Biochem J. 1988;256:283–290. doi: 10.1042/bj2560283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Bollen M. Combinatorial control of protein phosphatase-1. Trends Biochem Sci. 2001;26:426–431. doi: 10.1016/S0968-0004(01)01836-9. [DOI] [PubMed] [Google Scholar]
  9. Braszko JJ, Kupryszewski G, Witczuk B, Wisniewski K. Angiotensin II-(3–8)-hexapeptide affects motor activity, performance of passive avoidance and a conditioned avoidance response in rats. Neuroscience. 1988;27:777–783. doi: 10.1016/0306-4522(88)90182-0. [DOI] [PubMed] [Google Scholar]
  10. Buchsbaum RJ, Connolly BA, Feig LA. Regulation of p70 S6 kinase by complex formation between the Rac guanine nucleotide exchange factor (Rac-GEF) Tiam1 and the scaffold spinophilin. J Biol Chem. 2003;278:18833–18841. doi: 10.1074/jbc.M207876200. [DOI] [PubMed] [Google Scholar]
  11. Burnett PE, Blackshaw S, Lai MM, Qureshi IA, Burnett AF, Sabatini DM, Snyder SH. Neurabin is a synaptic protein linking p70 S6 kinase and the neuronal cytoskeleton. Proc Natl Acad Sci U S A. 1998;95:8351–8356. doi: 10.1073/pnas.95.14.8351. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Ceulemans H, Bollen M. Functional diversity of protein phosphatase-1, a cellular economizer and reset button. Physiol Rev. 2004;84:1–39. doi: 10.1152/physrev.00013.2003. [DOI] [PubMed] [Google Scholar]
  13. Chansel D, Czekalski S, Vandermeersch S, Ruffet E, Fournie-Zaluski MC, Ardaillou R. Characterization of angiotensin IV-degrading enzymes and receptors on rat mesangial cells. Am J Physiol. 1998;275:F535–542. doi: 10.1152/ajprenal.1998.275.4.F535. [DOI] [PubMed] [Google Scholar]
  14. Chansel D, Vandermeersch S, Oko A, Curat C, Ardaillou R. Effects of angiotensin IV and angiotensin-(1–7) on basal and angiotensin II-stimulated cytosolic Ca2+ in mesangial cells. Euro J Pharm. 2001;414:165–175. doi: 10.1016/S0014-2999(01)00791-9. [DOI] [PubMed] [Google Scholar]
  15. Cheng A, Dean N M, Honkanen RE. Serine/threonine protein phosphatase type 1γ1 is required for the completion of cytokinesis in human A549 lung carcinoma cells. J Biol Chem. 2000;275:1846–1854. doi: 10.1074/jbc.275.3.1846. [DOI] [PubMed] [Google Scholar]
  16. Chou MM, Blenis J. The 70 kDa S6 kinase: regulation of a kinase with multiple roles in mitogenic signalling. Curr Opin Cell Biol. 1995;7:806–814. doi: 10.1016/0955-0674(95)80064-6. [DOI] [PubMed] [Google Scholar]
  17. Chung J, Grammar T C, Lemon KP, Kazlauskas A, Blenis J. PDGF-and insulin-dependent pp70S6K activation mediated by phosphatidylinositol-3-OH kinase. Nature. 1994;370:71–75. doi: 10.1038/370071a0. [DOI] [PubMed] [Google Scholar]
  18. Cohen PTW. Protein phosphatase 1-targeted in many directions. J Cell Sci. 2002;115:241–256. doi: 10.1242/jcs.115.2.241. [DOI] [PubMed] [Google Scholar]
  19. Cui Z, Chen X, Lu B, Park SK, Xu T, Xie Z, Xue P, Hou J, Hang H, Yates JR. Preliminary quantitative profile of differential protein expression between rat L6 myoblasts and myotubes by stable isotope labeling with amino acids in cell culture. Proteomics. 2009;9:1274–1292. doi: 10.1002/pmic.200800354. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Dostal DE, Murahashi T, Peach MJ. Regulation of cytosolic calcium by angiotensins in vascular smooth muscle. Hypertension. 1990;15:815–822. doi: 10.1161/01.HYP.15.6.815. [DOI] [PubMed] [Google Scholar]
  21. Dou QP, An B, Will PL. Induction of a retinoblastoma phosphatase activity by anticancer drugs accompanies p53-independent G1 arrest and apoptosis. Proc Natl Acad Sci U S A. 1995;92:9019–9023. doi: 10.1073/pnas.92.20.9019. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Dulin N, Madhun ZT, Chang CH, Berti-Mattera L, Dickens D, Douglas JG. Angiotensin IV receptors and signaling in opossum kidney cells. Am J Physiol. 1995;269:F644–652. doi: 10.1152/ajprenal.1995.269.5.F644. [DOI] [PubMed] [Google Scholar]
  23. Dyson N. The regul ation of E2F by pRB-family proteins. Genes Dev. 1998;12:2245–2262. doi: 10.1101/gad.12.15.2245. [DOI] [PubMed] [Google Scholar]
  24. Ewen ME, Sluss HK, Sherr CJ, Matsushime H, Kato J-y, Livingston DM. Functional interactions of the retinoblastoma protein with mammalian D-type cyclins. Cell. 1993;73:487–497. doi: 10.1016/0092-8674(93)90136-E. [DOI] [PubMed] [Google Scholar]
  25. Fernandez A, Brautigan D L, Lamb N. Protein phosphatase type 1 in mammalian cell mitosis: chromosomal localization and involvement in mitotic exit. J Cell Biol. 1992;116:1421–1430. doi: 10.1083/jcb.116.6.1421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Fernando RN, Albiston AL, Chai SY. The insulinregulated aminopeptidase IRAP is colocalised with GLUT4 in the mouse hippocampus-potential role in modulation of glucose uptake in neurones? Euro J Neurosci. 2008;28:588–598. doi: 10.1111/j.1460-9568.2008.06347.x. [DOI] [PubMed] [Google Scholar]
  27. Gräff J, Koshibu K, Jouvenceau A, Dutar P, Mansuy IM. Protein phosphatase 1-dependent transcriptional programs for long-term memory and plasticity. Learn Mem. 2010;17:355–363. doi: 10.1101/lm.1766510. [DOI] [PubMed] [Google Scholar]
  28. Handa RK. Characteriza tion and signaling of the AT(4) receptor in human proximal tubule epithelial (HK-2) cells. J Am Soc Nephrol. 2001;12:440–449. doi: 10.1681/ASN.V123440. [DOI] [PubMed] [Google Scholar]
  29. Hinds PW, Mittnacht S, Dulic V, Arnold A, Reed SI, Weinberg RA. Regulation of retinoblastoma protein functions by ectopic expression of human cyclins. Cell. 1992;70:993–1006. doi: 10.1016/0092-8674(92)90249-C. [DOI] [PubMed] [Google Scholar]
  30. Hou J, Cui Z, Xie Z, Xue P, Wu P, Chen X, Li J, Cai T, Yang F. Phosphoproteome analysis of rat L6 myotubes using reversed-phase C18 prefractionation and titanium dioxide enrichment. J Proteome Res. 2010;9:777–788. doi: 10.1021/pr900646k. [DOI] [PubMed] [Google Scholar]
  31. Hsieh-Wilson LC, Benfenati F, Snyder GL, Allen PB, Nairn AC, Greengard P. Phosphorylation of spinophilin modulates its interaction with actin filaments. J Biol Chem. 2003;278:1186–1194. doi: 10.1074/jbc.M205754200. [DOI] [PubMed] [Google Scholar]
  32. Kato J, Matsushime H, Hiebert SW, Ewen ME, Sherr CJ. Direct binding of cyclin D to the retinoblastoma gene product (pRb) and pRb phosphorylation by the cyclin D-dependent kinase CDK4. Genes Dev. 1993;7:331–342. doi: 10.1101/gad.7.3.331. [DOI] [PubMed] [Google Scholar]
  33. Lee J, Albiston AL, Allen AM, Mendelsohn FA, Ping SE, Barrett GL, Murphy M, Morris MJ, McDowall SG, Chai SY. Effect of I.C.V. injection of AT4 receptor ligands, NLE1-angiotensin IV and LVV-hemorphin 7, on spatial learning in rats. Neuroscience. 2004;124:341–349. doi: 10.1016/j.neuroscience.2003.12.006. [DOI] [PubMed] [Google Scholar]
  34. Lew RA, Mustafa T, Ye S, McDowall SG, Chai SY, Albiston AL. Angiotensin AT4 ligands are potent, competitive inhibitors of insulin regulated aminopeptidase (IRAP) J Neurochem. 2003;86:344–350. doi: 10.1046/j.1471-4159.2003.01852.x. [DOI] [PubMed] [Google Scholar]
  35. Li T, Chalifour LE, Paudel HK. Phosphorylation of protein phosphatase 1 by cyclin-dependent protein kinase 5 during nerve growth factor-induced PC12 cell differentiation. J Biol Chem. 2007;282:6619–6628. doi: 10.1074/jbc.M606347200. [DOI] [PubMed] [Google Scholar]
  36. Li YD, Block ER, Patel JM. Activation of multiple signaling modules is critical in angiotensin IV-induced lung endothelial cell proliferation. Am J Physiol Lung Cell Mol Physiol. 2002;283:L707–716. doi: 10.1152/ajplung.00024.2002. [DOI] [PubMed] [Google Scholar]
  37. Liu CWY, Wang RH, Dohadwala M, Schönthal AH, Villa-Moruzzi E, Berndt N. Inhibitory phosphorylation of PP1α catalytic subunit during the G1/S transition. J Biol Chem. 1999;274:29470–29475. doi: 10.1074/jbc.274.41.29470. [DOI] [PubMed] [Google Scholar]
  38. Molina-Pinelo S, Ferrer I, Blanco-Aparicio C, Peregrino S, Pastor MD, Alvarez-Vega J, Suarez R, Verge M, Marin JJ, Hernandez-Losa J. Down-regulation of spinophilin in lung tumours contributes to tumourigenesis. J Pathol. 2011;225:73–82. doi: 10.1002/path.2905. [DOI] [PubMed] [Google Scholar]
  39. Ragusa MJ, Dancheck B, Critton DA, Nairn AC, Page R, Peti W. Spinophilin directs protein phosphatase 1 specificity by blocking substrate binding sites. Nat Struct Mol Biol. 2010;17:459–464. doi: 10.1038/nsmb.1786. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Ruiz-Ortega M, Esteban V, Egido J. The regulation of the inflammatory response through nuclear factor-kappab pathway by angiotensin IV extends the role of the renin angiotensin system in cardiovascular diseases. Trends Cardiovasc Med. 2007;17:19–25. doi: 10.1016/j.tcm.2006.10.003. [DOI] [PubMed] [Google Scholar]
  41. Sarrouilhe D, Di Tommaso A, Metaye T, Ladeveze V. Spinophilin: from partners to functions. Biochimie. 2006;88:1099–1113. doi: 10.1016/j.biochi.2006.04.010. [DOI] [PubMed] [Google Scholar]
  42. Satoh A, Nakanishi H, Obaishi H, Wada M, Takahashi K, Satoh K, Hirao K, Nishioka H, Hata Y, Mizoguchi A. Neurabin-II/spinophilin. J Biol Chem. 1998;273:3470–3475. doi: 10.1074/jbc.273.6.3470. [DOI] [PubMed] [Google Scholar]
  43. Shi Y. Serine/threonine phosphatases: mechani sm through structure. Cell. 2009;139:468–484. doi: 10.1016/j.cell.2009.10.006. [DOI] [PubMed] [Google Scholar]
  44. Shibasaki Y, Mori Y, Tsutumi Y, Masaki H, Sakamoto K, Murasawa S, Maruyama K, Moriguchi Y, Tanaka Y, Iwasaka T, et al. Differential kinetics of circulating angiotensin IV and II after treatment with angiotensin II type 1 receptor antagonist and their plasma levels in patients with chronic renal failure. Clin Nephrol. 1999;51:83–91. [PubMed] [Google Scholar]
  45. Smith FD, Oxford GS, Milgram SL. As sociation of the D2 dopamine receptor third cytoplasmic loop with spinophilin, a protein phosphatase-1-interacting protein. J Biol Chem. 1999;274:19894–19900. doi: 10.1074/jbc.274.28.19894. [DOI] [PubMed] [Google Scholar]
  46. Takai A, Bialojan C, Troschka M, Rüegg J. Smooth muscle myosin phosphatase inhibition and force enhancement by black sponge toxin. FEBS Lett. 1987;217:81–84. doi: 10.1016/0014-5793(87)81247-4. [DOI] [PubMed] [Google Scholar]
  47. Terry-Lorenzo RT, Carmody LC, Voltz JW, Connor JH, Li S, Smith FD, Milgram SL, Colbran RJ, Shenolikar S. The neuronal actin-binding proteins, neurabin I and neurabin II, recruit specific isoforms of protein phosphatase-1 catalytic subunits. J Biol Chem. 2002;277:27716–27724. doi: 10.1074/jbc.M203365200. [DOI] [PubMed] [Google Scholar]
  48. Tsukada M, Prokscha A, Oldekamp J, Eichele G. Identification of neurabin II as a novel doublecortin interacting protein. Mech Dev. 2003;120:1033–1043. doi: 10.1016/S0925-4773(03)00177-1. [DOI] [PubMed] [Google Scholar]
  49. Vivo M, Calogero RA, Sansone F, Calabro V, Parisi T, Borrelli L, Saviozzi S, La Mantia G. The human tumor suppressor arf interacts with spinophilin/neurabin II, a type 1 proteinphosphatase-binding protein. J Biol Chem. 2001;276:14161–14169. doi: 10.1074/jbc.M006845200. [DOI] [PubMed] [Google Scholar]
  50. Wang P, Bai Y, Song B, Wang Y, Liu D, Lai Y, Bi X, Yuan Z. PP1A-mediated dephosphorylation positively regulates YAP2 activity. PloS One. 2011;6:e24288. doi: 10.1371/journal.pone.0024288. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Wright JW, Stubley L, Pederson ES, Kramar EA, Hanesworth JM, Harding JW. Contributions of the brain angiotensin IV-AT4 receptor subtype system to spatial learning. J Neurosci. 1999;19:3952–3961. doi: 10.1523/JNEUROSCI.19-10-03952.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Yan Y, Mumby MC. Distinct Roles for PP1 and P P2A in Phosphorylation of the Retinoblastoma Protein PP2A REGULATES THE ACTIVITIES OF G1 CYCLIN-DEPENDENT KINASES. J Biol Chem. 1999;274:31917–31924. doi: 10.1074/jbc.274.45.31917. [DOI] [PubMed] [Google Scholar]

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