Journal of Applied Biological Chemistry

The Korean Society for Applied Biological Chemistry (한국응용생명화학회)
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Independent Regulation of Endothelial Nitric Oxide Synthase by Src and Protein Kinase A in Mouse Aorta Endothelial Cells

  • Boo, Yong-Chool (Department of Molecular Medicine, Kyungpook National University School of Medicine)
  • Published : 2005.09.30
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Abstract

Endothelial nitric oxide synthase (eNOS) plays a critical role in vascular biology and pathophysiology. Its activity is regulated by multiple mechanisms such as calcium/calmodulin, protein-protein interactions, sub-cellular locations and phosphorylation at various sites. Phosphorylation of eNOS-Ser1177 (based on mouse sequence) has been identified as an important mechanism of eNOS activation. However, signaling pathway leading to it phosphorylation remains controversial. The regulation of eNOS-Ser1177 phosphorylation by Src and protein kinase A (PKA) was investigated in the present study using cultured mouse aorta endothelial cells. Expression of a constitutively active Src mutant in the cells enhanced phosphorylation of eNOS and protein kinase B (Akt). The Src-stimulated phosphorylation was not attenuated by the expression of a dominant negative PKA regulatory subunit. Neither activation nor inhibition of PKA activity had any significant effect on tyrosine phosphorylation of activation or inactivation site in Src. Based on the results of this study, it is suggested that Src/Akt pathway and PKA signaling may regulate eNOS phosphorylation independently. The existence of multiple mechanisms for eNOS phosphorylation may guarantee endothelial nitric oxide production in various cellular contexts which is essential for maintenance of vascular health.

Keywords

References

  1. Free Radic. Res. v.31 Endogenous nitric oxide synthesis: biological functions and pathophysiology Bredt, D.S. https://doi.org/10.1080/10715769900301161
  2. Biochem. J. v.357 Nitric oxide synthases: structure, function and inhibition Alderton, W.K.;Cooper, C.E.;Knowles, R.G. https://doi.org/10.1042/0264-6021:3570593
  3. Nature v.377 Hypertension in mice lacking the gene for endothelial nitric oxide synthase Huang, P.L.;Huang, Z.;Mashimo, H.;Bloch, K.D.;Moskowitz, M.A.;Bevan, J.A.;Fishman, M.C. https://doi.org/10.1038/377239a0
  4. Proc. Natl. Acad. Sci. USA v.93 Elevated blood pressures in mice lacking endothelial nitric oxide synthase Shesely, E.G.;Maeda, N.;Kim, H.S.;Desai, K.M.;Krege, J.H.;Laubach, V.E.;Sherman, P.A.;Sessa, W.C.;Smithies, O.
  5. J. Biol. Chem. v.275 Characterization of the roles of the 594-645 region in human endothelial nitric-oxide synthase in regulating calmodulin binding and electron transfer Chen, P.F.;Wu, K.K. https://doi.org/10.1074/jbc.275.17.13155
  6. J. Biol. Chem. v.274 Autoinhibition of endothelial nitric-oxide synthase. Identification of an electron transfer control element Nishida, C.R.;Ortiz de Montellano, P.R. https://doi.org/10.1074/jbc.274.21.14692
  7. J. Biol. Chem. v.272 Direct interaction of endothelial nitric-oxide synthase and caveolin-1 inhibits synthase activity Ju, H.;Zou, R.;Venema, V.J.;Venema, R.C. https://doi.org/10.1074/jbc.272.30.18522
  8. J. Biol. Chem. v.272 Reciprocal regulation of endothelial nitric-oxide synthase by $Ca^{2+}$- calmodulin and caveolin Michel, J.B.;Feron, O.;Sacks, D.;Michel, T. https://doi.org/10.1074/jbc.272.25.15583
  9. Nature v.392 Dynamic activation of endothelial nitric oxide synthase by Hsp90 Garcia-Cardena, G.;Fan, R.;Shah, V.;Sorrentino, R.;Cirino, G.;Papapetropoulos, A.;Sessa, W.C. https://doi.org/10.1038/33934
  10. J. Biol. Chem. v.276 Direct interaction between endothelial nitric-oxide synthase and dynamin-2. Implications for nitric-oxide synthase function Cao, S.;Yao, J.;McCabe, T.J.;Yao, Q.;Katusic, Z.S.;Sessa, W.C.;Shah, V. https://doi.org/10.1074/jbc.M006258200
  11. Proc. Natl. Acad. Sci. USA v.99 Functional interaction of endothelial nitric oxide synthase with a voltage-dependent anion channel Sun, J.;Liao, J.K.
  12. Faseb. J. v.15 NOSIP, a novel modulator of endothelial nitric oxide synthase activity Dedio, J.;Konig, P.;Wohlfart, P.;Schroeder, C.;Kummer, W.;Muller-Esterl, W. https://doi.org/10.1096/fj.00-0078com
  13. J. Biol. Chem. v.269 Identification of covalently bound amino-terminal myristic acid in endothelial nitric oxide synthase Liu, J.;Sessa, W.C.
  14. Circ. Res. v.72 Mutation of N-myristoylation site converts endothelial cell nitric oxide synthase from a membrane to a cytosolic protein Sessa, W.C.;Barber, C.M.;Lynch, K.R. https://doi.org/10.1161/01.RES.72.4.921
  15. J. Biol. Chem. v.277 Subcellular targeting and agonist-induced site-specific phosphorylation of endothelial nitric-oxide synthase Gonzalez, E.;Kou, R.;Lin, A.J.;Golan, D.E.;Michel, T. https://doi.org/10.1074/jbc.M207299200
  16. J. Biol. Chem. v.279 Targeting of endothelial nitric-oxide synthase to the cytoplasmic face of the Golgi complex or plasma membrane regulates Akt- versus calcium-dependent mechanisms for nitric oxide release Fulton, D.;Babbitt, R.;Zoellner, S.;Fontana, J.;Acevedo, L.;McCabe, T.J.;Iwakiri, Y.;Sessa, W.C. https://doi.org/10.1074/jbc.M402155200
  17. Nature v.399 Regulation of endothelium-derived nitric oxide production by the protein kinase Akt Fulton, D.;Gratton, J.P.;McCabe, T.J.;Fontana, J.;Fujio, Y.;Walsh, K.;Franke, T.F.;Papapetropoulos, A.;Sessa, W.C. https://doi.org/10.1038/21218
  18. Nature v.399 Activation of nitric oxide synthase in endothelial cells by Akt- dependent phosphorylation Dimmeler, S.;Fleming, I.;Fisslthaler, B.;Hermann, C.;Busse, R.;Zeiher, A.M. https://doi.org/10.1038/21224
  19. J. Biol. Chem. v.277 Identification of regulatory sites of phosphorylation of the bovine endothelial nitric-oxide synthase at serine 617 and serine 635 Michell, B.J.;Harris, M.B.;Chen, Z.P.;Ju, H.;Venema, V.J.;Blackstone, M.A.;Huang, W.;Venema, R.C.;Kemp, B.E. https://doi.org/10.1074/jbc.M205144200
  20. J. Biol. Chem. v.276 Coordinated control of endothelial nitric-oxide synthase phosphorylation by protein kinase C and the cAMP-dependent protein kinase Michell, B.J.;Chen, Z.;Tiganis, T.;Stapleton, D.;Katsis, F.;Power, D.A.;Sim, A.T.;Kemp, B.E. https://doi.org/10.1074/jbc.C100122200
  21. Am. J. Physiol. Cell Physiol. v.285 Flow-dependent regulation of endothelial nitric oxide synthase: role of protein kinases Boo, Y.C.;Jo, H. https://doi.org/10.1152/ajpcell.00122.2003
  22. Circ. Res. v.93 Ligand-independent activation of vascular endothelial growth factor receptor 2 by fluid shear stress regulates activation of endothelial nitric oxide synthase Jin, Z.G.;Ueba, H.;Tanimoto, T.;Lungu, A.O.;Frame, M.D.;Berk, B.C. https://doi.org/10.1161/01.RES.0000089257.94002.96
  23. J. Biol. Chem. v.280 Flow shear stress stimulates Gab1 tyrosine phosphorylation to mediate protein kinase B and endothelial nitric-oxide synthase activation in endothelial cells Jin, Z.G.;Wong, C.;Wu, J.;Berk, B.C. https://doi.org/10.1074/jbc.M500294200
  24. J. Biol. Chem. v.277 Shear stress stimulates phosphorylation of endothelial nitric-oxide synthase at Ser1179 by Akt-independent mechanisms: role of protein kinase A Boo, Y.C.;Sorescu, G.;Boyd, N.;Shiojima, I.;Walsh, K.;Du, J.;Jo, H. https://doi.org/10.1074/jbc.M108789200
  25. Am. J. Physiol. Heart Circ. Physiol. v.283 Shear stress stimulates phosphorylation of eNOS at Ser(635) by a protein kinase A-dependent mechanism Boo, Y.C.;Hwang, J.;Sykes, M.;Michell, B.J.;Kemp, B.E.;Lum, H.;Jo, H. https://doi.org/10.1152/ajpheart.00214.2002
  26. J. Exp. Med. v.193 Activation of the COOH-terminal Src kinase (Csk) by cAMP-dependent protein kinase inhibits signaling through the T cell receptor Vang, T.;Torgersen, K.M.;Sundvold, V.;Saxena, M.;Levy, F.O.;Skalhegg, B.S.;Hansson, V.;Mustelin, T.;Tasken, K. https://doi.org/10.1084/jem.193.4.497
  27. Mol. Cell v.9 PKA phosphorylation of Src mediates cAMP's inhibition of cell growth via Rap1 Schmitt, J.M.;Stork, P.J. https://doi.org/10.1016/S1097-2765(01)00432-4
  28. J. Biol. Chem. v.278 Protein kinase A intersects SRC signaling in membrane microdomains Abrahamsen, H.;Vang, T.;Tasken, K. https://doi.org/10.1074/jbc.M211426200
  29. J. Biol. Chem. v.278 Oscillatory shear stress stimulates endothelial production of $O_2^-$ from $p47^{phox}$- dependent NAD(P)H oxidases, leading to monocyte adhesion Hwang, J.;Saha, A.;Boo, Y.C.;Sorescu, G.P.;McNally, J.S.;Holland, S.M.;Dikalov, S.;Giddens, D.P.;Griendling, K.K.;Harrison, D.G.;Jo, H. https://doi.org/10.1074/jbc.M305150200
  30. Proc. Natl. Acad Sci. USA v.89 Mutations in the catalytic subunit of cAMP-dependent protein kinase result in unregulated biological activity Orellana, S.A.;McKnight, G.S.
  31. J. Biol. Chem. v.262 Inhibition of intracellular cAMP-dependent protein kinase using mutant genes of the regulatory type I subunit Clegg, C.H.;Correll, L.A.;Cadd, G.G.;McKnight, G.S.
  32. EMBO J. v.14 Mutational analysis of the Src SH3 domain: the same residues of the ligand binding surface are important for intraand intermolecular interactions Erpel, T.;Superti-Furga, G.;Courtneidge, S.
  33. Free Radic. Biol. Med. v.35 Endothelial NO synthase phosphorylated at SER635 produces NO without requiring intracellular calcium increase Boo, Y.C.;Sorescu, G.P.;Bauer, P.M.;Fulton, D.;Kemp, B.E.;Harrison, D.G.;Sessa, W.C.;Jo, H. https://doi.org/10.1016/S0891-5849(03)00397-6
  34. Biochemistry (Mosc) v.65 Kinases of the Src family: structure and functions Tatosyan, A.G.;Mizenina, O.A.
  35. J. Biol. Chem. v.271 Phosphorylation of caveolin by Src tyrosine kinases - $The\;{\alpha}$ - isoform of caveolin is selectively phosphorylated by v-Src in vivo Li, S.W.;Seitz, R.;Lisanti, M.P. https://doi.org/10.1074/jbc.271.7.3863
  36. Curr. Opin. Neurobiol. v.7 PKA isoforms, neural pathways, and behaviour: making the connection Brandon, E.P.;Idzerda, R.L.;McKnight, G.S. https://doi.org/10.1016/S0959-4388(97)80069-4
  37. J. Vasc. Res. v.34 Calcium-dependent and calcium-independent activation of the endothelial NO synthase Fleming, I.;Bauersachs, J.;Busse, R. https://doi.org/10.1159/000159220
  38. FEBS Lett. v.477 Phosphorylation of the endothelial nitric oxide synthase at ser-1177 is required for VEGF-induced endothelial cell migration Dimmeler, S.;Dernbach, E.;Zeiher, A.M. https://doi.org/10.1016/S0014-5793(00)01657-4
  39. J. Biol. Chem. v.275 Endothelial nitric-oxide synthase (type III) is activated and becomes calcium independent upon phosphorylation by cyclic nucleotide-dependent protein kinases Butt, E.;Bernhardt, M.;Smolenski, A.;Kotsonis, P.;Frohlich, L.G.;Sickmann, A.;Meyer, H.E.;Lohmann, S.M.;Schmidt, H.H. https://doi.org/10.1074/jbc.275.7.5179
  40. FEBS Lett. v.443 AMP-activated protein kinase phosphorylation of endothelial NO synthase Chen, Z.P.;Mitchelhill, K.I.;Michell, B.J.;Stapleton, D.;Rodriguez-Crespo, I.;Witters, L.A.;Power, D.A.;Ortiz de Montellano, P.R.;Kemp, B.E. https://doi.org/10.1016/S0014-5793(98)01705-0
  41. Circ. Res. v.88 Phosphorylation of Thr(495) regulates $Ca(^{2+})$/calmodulin-dependent endothelial nitric oxide synthase activity Fleming, I.;Fisslthaler, B.;Dimmeler, S.;Kemp, B.E.;Busse, R. https://doi.org/10.1161/hh1101.092677
  42. Circ. Res. v.78 Intracellular pH and tyrosine phosphorylation but not calcium determine shear stress-induced nitric oxide production in native endothelial cells Ayajiki, K.;Kindermann, M.;Hecker, M.;Fleming, I.;Busse, R. https://doi.org/10.1161/01.RES.78.5.750
  43. Circ. Res. v.79 Phosphorylation of endothelial nitric oxide synthase in response to fluid shear stress Corson, M.A.;James, N.L.;Latta, S.E.;Nerem, R.M.;Berk, B.C.;Harrison, D.G. https://doi.org/10.1161/01.RES.79.5.984
  44. J. Biol. Chem. v.271 Endothelial nitric oxide synthase is regulated by tyrosine phosphorylation and interacts with caveolin-1 Garcia-Cardena, G.;Fan, R.;Stern, D.F.;Liu, J.;Sessa, W.C. https://doi.org/10.1074/jbc.271.44.27237
  45. Circ. Res. v.82 $Ca^{2+}$-independent activation of the endothelial nitric oxide synthase in response to tyrosine phosphatase inhibitors and fluid shear stress Fleming, I.;Bauersachs, J.;Fisslthaler, B.;Busse, R. https://doi.org/10.1161/01.RES.82.6.686
  46. J. Biol. Chem. v.277 Hydrogen peroxide activates endothelial nitric-oxide synthase through coordinated phosphorylation and dephosphorylation via a phosphoinositide 3-kinase-dependent signaling pathway Thomas, S.R.;Chen, K.;Keaney, J.F. Jr. https://doi.org/10.1074/jbc.M109107200
  47. Circ. Res. v.87 Membrane estrogen receptor engagement activates endothelial nitric oxide synthase via the PI3-kinase-Akt pathway in human endothelial cells Haynes, M.P.;Sinha, D.;Russell, K.S.;Collinge, M.;Fulton, D.;Morales-Ruiz, M.;Sessa, W.C.;Bender, J.R. https://doi.org/10.1161/01.RES.87.8.677
  48. J. Biol. Chem. v.278 Src kinase mediates phosphatidylinositol 3-kinase/ Akt-dependent rapid endothelial nitric-oxide synthase activation by estrogen Haynes, M.P.;Li, L.;Sinha, D.;Russell, K.S.;Hisamoto, K.;Baron, R.;Collinge, M.;Sessa, W.C.;Bender, J.R. https://doi.org/10.1074/jbc.M210828200
  49. J. Biol. Chem. v.277 Transactivation of vascular endothelial growth factor (VEGF) receptor Flk-1/KDR is involved in sphingosine 1-phosphate-stimulated phosphorylation of Akt and endothelial nitric-oxide synthase (eNOS) Tanimoto, T.;Jin, Z.G.;Berk, B.C. https://doi.org/10.1074/jbc.M204764200
  50. J. Biol. Chem. v.276 Regulation of Akt/PKB activation by tyrosine phosphorylation Chen, R.;Kim, O.;Yang, J.;Sato, K.;Eisenmann, K.M.;McCarthy, J.;Chen, H.;Qiu, Y. https://doi.org/10.1074/jbc.C100271200
  51. J. Mol. Cell. Cardiol. v.37 The role of phosphoinositide-3 kinase and PTEN in cardiovascular physiology and disease Oudit, G.Y.;Sun, H.;Kerfant, B.G.;Crackower, M.A.;Penninger, J.M.;Backx, P.H. https://doi.org/10.1016/j.yjmcc.2004.05.015
  52. Am. J. Physiol. v.275 Phosphatidylinositol 3-kinase gamma mediates shear stress-dependent activation of JNK in endothelial cells Go, Y.M.;Park, H.;Maland, M.C.;Darley-Usmar, V.M.;Stoyanov, B.;Wetzker, R.;Jo, H.