Summary
The actin-binding protein caldesmon (CaD) exists both in smooth muscle (the heavy isoform, h-CaD) and non-muscle cells (the light isoform, l-CaD). In smooth muscles h-CaD binds to myosin and actin simultaneously and modulates the actomyosin interaction. In non-muscle cells l-CaD binds to actin and stabilizes␣the actin stress fibers; it may also mediate the interaction between actin and non-muscle myosins. Both h- and l-CaD are phosphorylated in vivo upon stimulation. The major phosphorylation sites of h-CaD when activated by phorbol ester are the Erk-specific sites, modification of which is attenuated by the MEK inhibitor PD98059. The same sites in l-CaD are also phosphorylated when cells are stimulated to migrate, whereas in dividing cells l-CaD is phosphorylated more extensively, presumably by cdc2 kinase. Both Erk and cdc2 are members of the MAPK family. Thus it appears that CaD is a downstream effector of the Ras signaling pathways. Significantly, the phosphorylatable serine residues shared by both CaD isoforms are in the C-terminal region that also contains the actin-binding sites. Biochemical and structural studies indicated that phosphorylation of CaD at the Erk sites is accompanied by a conformational change that partially dissociates CaD from actin. Such a structural change in h-CaD exposes the myosin-binding sites on the actin surface and allows actomyosin interactions in smooth muscles. In the case of non-muscle cells, the change in l-CaD weakens the stability of the actin filament and facilitates its disassembly. Indeed, the level of l-CaD modification correlates very well in a reciprocal manner with the level of actin stress fibers. Since both cell migration and cell division require dynamic remodeling of actin cytoskeleton that leads to cell shape changes, phosphorylation of CaD may therefore serve as a plausible means to regulate these processes. Thus CaD not only links the smooth muscle contractility and non-muscle motility, but also provides a common mechanism for the regulation of cell migration and cell proliferation.
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Abbreviations
- BPM:
-
benzophenone maleimide
- CaD:
-
caldesmon
- CaM:
-
calmodulin
- EM:
-
electron microscopy
- Erk:
-
extracellular signal-regulated kinase
- FRET:
-
fluorescence resonance energy transfer
- GFP:
-
green fluorescent protein
- h-CaD:
-
smooth muscle caldesmon
- IAEDANS:
-
5-(iodoacetamidoethyl) aminonaphthalene-1-sulfonic acid
- l-CaD:
-
non-muscle caldesmon
- MAPK:
-
mitogen activated protein kinase
- MLCK:
-
myosin light chain kinase
- MS:
-
mass spectrometry
- Pak:
-
p21-activated protein kinase
- PMA:
-
phorbol 12-myristate 13-acetate
- Raf:
-
rat aorta fibroblast cells
- Tm:
-
tropomyosin
References
Wang C.L. (2001). Caldesmon and smooth-muscle regulation. Cell Biochem. Biophys. 35:275–288
Mirzapoiazova T., Kolosova I.A., Romer L., Garcia J.G. and Verin A.D. (2005). The role of caldesmon in the regulation of endothelial cytoskeleton and migration. J. Cell Physiol. 203: 520–528
Yamakita Y., Shigeko Y., Matsumura F. (2001). Inhibition of Arp2/3-dependent actin nucleation by caldesmon. Mol Biol Cell 12:32a
Yamakita Y., Matsumura F. (2002). Caldesmon inhibits cortactin-stabilized, Arp2/3-mediated actin branching. Mol. Biol. Cell 13:316a
Yamashiro S., Chern H., Yamakita Y., Matsumura F. (2001). Mutant Caldesmon lacking cdc2 phosphorylation sites delays M-phase entry and inhibits cytokinesis. Mol. Biol. Cell 12:239–250
Numaguchi Y., Huang S., Polte T.R., Eichler G.S., Wang N., Ingber D.E. (2003). Caldesmon-dependent switching between capillary endothelial cell growth and apoptosis through modulation of cell shape and contractility. Angiogenesis 6:55–64
Cuomo M.E., Knebel A., Platt G., Morrice N., Cohen P., Mittnacht S. (2005). Regulation of microfilament organization by KSHV- cyclin/cdk phosphorylation of caldesmon. J. Biol. Chem. 280:35844–35858
Bretscher A. (1986). Thin filament regulatory proteins of smooth- and non-muscle cells. Nature 321:726–727
Boerner J.L., Danielsen A.J., Lovejoy C.A., Wang Z., Juneja S.C., Faupel-Badger J.M., Darce J.R., Maihle N.J. (2003). Grb2 regulation of the actin-based cytoskeleton is required for ligand-independent EGF receptor-mediated oncogenesis. Oncogene 22:6679–6689
Manes T., Zheng D.Q., Tognin S., Woodard A.S., Marchisio P.C., Languino L.R. (2003). Alpha(v)beta3 integrin expression up-regulates cdc2, which modulates cell migration. J. Cell Biol. 161:817–826
Juliano R. (2003). Movin’ on through with Cdc2. Nat. Cell Biol. 5:589–590
Kordowska J., Hetrick T., Adam L.P. and Wang C.-L.A., Phosphorylated l-caldesmon is involved in disassembly of actin stress fibers and postmitotic spreading. Exp. Cell Res. 312: 95–110, 2006.
Marston S.B. and Huber P.A.J., Caldesmon. In: Bárány M. (Ed), Biochemistry of Smooth Muscle Contraction. Academic Press, Inc., San Diego, CA, 1996, pp. 77–90.
Matsumura F. and Yamashiro S. (1993). Caldesmon. Curr. Opin. Cell Biol. 5:70–76
Humphrey M.B., Herrera-Sosa H., Gonzalez G., Lee R., Bryan J. (1992). Cloning of cDNAs encoding human caldesmons. Gene 112:197–204
Mabuchi K., Li Y., Tao T. and Wang C.L. (1996). Immunocytochemical localization of caldesmon and calponin in chicken gizzard smooth muscle. J. Muscle Res. Cell Motil. 17:243–260
Bretscher A. and Lynch W. (1985). Identification and localization of immunoreactive forms of caldesmon in smooth and nonmuscle cells: a comparison with the distributions of tropomyosin and alpha-actinin. J. Cell Biol. 100:1656–1663
Warren K.S., Lin J.L., Wamboldt D.D. and Lin J.J. (1994). Overexpression of human fibroblast caldesmon fragment containing actin-, Ca++/calmodulin-, and tropomyosin-binding domains stabilizes endogenous tropomyosin and microfilaments. J. Cell Biol. 125:359–368
Pittenger M.F., Kistler A. and Helfman D.M. (1995). Alternatively spliced exons of the beta tropomyosin gene exhibit different affinities for F-actin and effects with nonmuscle caldesmon. J. Cell Sci. 108:3253–3265
Owada M.K., Hakura A., Iida K., Yahara I., Sobue K. and Kakiuchi S. (1984). Occurrence of caldesmon (a calmodulin-binding protein) in cultured cells: comparison of normal and transformed cells. Proc. Natl. Acad. Sci. USA 81:3133–3137
Sobue K., Muramoto Y., Fujita M. and Kakiuchi S. (1981). Purification of a calmodulin-binding protein from chicken gizzard that interacts with F-actin. Proc. Natl. Acad. Sci. USA 78:5652–5655
Marston S.B. and Lehman W. (1985). Caldesmon is a Ca2+-regulatory component of native smooth-muscle thin filaments. Biochem. J. 231:517–522
Ngai P.K. and Walsh M.P. (1984). Inhibition of smooth muscle actin-activated myosin Mg2+-ATPase activity by caldesmon. J. Biol. Chem. 259:13656–13659
Horiuchi K.Y., Miyata H. and Chacko S. (1986). Modulation of smooth muscle actomyosin ATPase by thin filament associated proteins. Biochem. Biophys. Res. Commun. 136:962–968
Smith C.W. and Marston S.B. (1985). Disassembly and reconstitution of the Ca2+-sensitive thin filaments of vascular smooth muscle. FEBS Lett. 184:115–119
Bartegi A., Fattoum A., Derancourt J. and Kassab R. (1990). Characterization of the carboxyl-terminal 10-kDa cyanogen bromide fragment of caldesmon as an actin–calmodulin-binding region. J. Biol. Chem. 265:15231–15238
Fujii T., Imai M., Rosenfeld G.C. and Bryan J. (1987). Domain mapping of chicken gizzard caldesmon. J. Biol. Chem. 262:2757–2763
Riseman V.M., Lynch W.P., Nefsky B. and Bretscher A. (1989). The calmodulin and F-actin binding sites of smooth muscle caldesmon lie in the carboxyl-terminal domain whereas the molecular weight heterogeneity lies in the middle of the molecule. J. Biol. Chem. 264:2869–2875
Szpacenko, A. and Dabrowska, R. (1986). Functional domain of caldesmon. FEBS Lett. 202:182–186
Wang C.L., Wang L.W., Xu S.A., Lu R.C., Saavedra-Alanis V. and Bryan J. (1991). Localization of the calmodulin- and the actin-binding sites of caldesmon. J. Biol. Chem. 266:9166–9172
Wang Z. and Chacko S. (1996). Mutagenesis analysis of functionally important domains within the C-terminal end of smooth muscle caldesmon. J. Biol. Chem. 271:25707–25714
Velaz L., Ingraham R.H. and Chalovich J.M. (1990). Dissociation of the effect of caldesmon on the ATPase activity and on the binding of smooth heavy meromyosin to actin by partial digestion of caldesmon. J. Biol. Chem. 265:2929–2934
Wang Z., Jiang H., Yang Z.Q. and Chacko S. (1997). Both N-terminal myosin-binding and C-terminal actin-binding sites on smooth muscle caldesmon are required for caldesmon-mediated inhibition of actin filament velocity. Proc. Natl. Acad. Sci. USA 94:11899–11904
Li Y., Zhuang S., Guo H., Mabuchi K., Lu R.C. and Wang C.A. (2000). The major myosin-binding site of caldesmon resides near its N-terminal extreme. J. Biol. Chem. 275:10989–10994
Lee Y.H., Gallant C., Guo H., Li Y., Wang C.A. and Morgan K.G. (2000). Regulation of vascular smooth muscle tone by N-terminal region of caldesmon. Possible role of tethering actin to myosin. J. Biol. Chem. 275:3213–3220
Zhan Q.Q., Wong S.S. and Wang C.L. (1991). A calmodulin-binding peptide of caldesmon. J. Biol. Chem. 266:21810–21814
Katsuyama H., Wang C.L. and Morgan K.G. (1992). Regulation of vascular smooth muscle tone by caldesmon. J. Biol. Chem. 267:14555–14558
Adam L.P., Haeberle J.R. and Hathaway D.R. (1989). Phosphorylation of caldesmon in arterial smooth muscle. J. Biol. Chem. 264:7698–7703
Adam L.P., Gapinski C.J. and Hathaway D.R. (1992). Phosphorylation sequences in h-caldesmon from phorbol ester-stimulated canine aortas. FEBS Lett. 302:223–226
Childs T.J., Watson M.H., Sanghera J.S., Campbell D.L., Pelech S.L. and Mak A.S. (1992). Phosphorylation of smooth muscle caldesmon by mitogen-activated protein (MAP) kinase and expression of MAP kinase in differentiated smooth muscle cells. J. Biol. Chem. 267:22853–22859
D’Angelo G., Graceffa P., Wang C.A., Wrangle J. and Adam L.P. (1999). Mammal-specific, ERK-dependent, caldesmon phosphorylation in smooth muscle. Quantitation using novel anti-phosphopeptide antibodies. J. Biol. Chem. 274:30115–30121
Adam L.P. and Hathaway D.R. (1993). Identification of mitogen-activated protein kinase phosphorylation sequences in mammalian h-Caldesmon. FEBS Lett. 322:56–60
Yamashiro S., Yamakita Y., Yoshida K., Takiguchi K. and Matsumura F (1995). Characterization of the COOH terminus of non-muscle caldesmon mutants lacking mitosis-specific phosphorylation sites. J. Biol. Chem. 270:4023–4230
Huang R., Li L., Guo H. and Wang C.-L.A. (2003). Caldesmon binding to actin is regulated by calmodulin and phosphorylation via different mechanisms. Biochemistry 42:2513–2523
Foster D.B., Huang R., Hatch V., Craig R., Graceffa P., Lehman W. and Wang C.-L.A. (2004). Modes of caldesmon binding to actin: sites of caldesmon contact and modulation of interactions by phosphorylation. J. Biol. Chem. 279:53387–53394
Lehman W., Vibert P. and Craig R. (1997). Visualization of caldesmon on smooth muscle thin filaments. J. Mol. Biol. 274:310–317
Nixon G.F., Iizuka K., Haystead C.M., Haystead T.A., Somlyo A.P. and Somlyo A.V. (1995). Phosphorylation of caldesmon by mitogen-activated protein kinase with no effect on Ca2+ sensitivity in rabbit smooth muscle. J. Physiol. 487:283–289
Yamashiro S., Yamakita Y., Ishikawa R. and Matsumura F. (1990). Mitosis-specific phosphorylation causes 83K non-muscle caldesmon to dissociate from microfilaments. Nature 344:675–678
Yamashiro S., Yamakita Y., Hosoya H. and Matsumura F. (1991). Phosphorylation of non-muscle caldesmon by p34cdc2 kinase during mitosis. Nature 349:169–172
Hosoya N., Hosoya H., Yamashiro S., Mohri H. and Matsumura F. (1993). Localization of caldesmon and its dephosphorylation during cell division. J. Cell Biol. 121:1075–1082
Li Y., Wessels D., Wang T., Lin J.L., Soll D.R. and Lin J.J. (2003). Regulation of caldesmon activity by Cdc2 kinase plays an important role in maintaining membrane cortex integrity during cell division. Cell Mol. Life Sci. 60:198–211
Franklin M.T., Wang C.L. and Adam L.P. (1997). Stretch-dependent activation and desensitization of mitogen-activated protein kinase in carotid arteries. Am. J. Physiol. 273:C1819–C1827
Yamboliev I.A. and Gerthoffer W.T. (2001). Modulatory role of ERK MAPK-caldesmon pathway in PDGF-stimulated migration of cultured pulmonary artery SMCs. Am. J. Physiol. Cell Physiol. 280:C1680–C1688, 2001
Goncharova E.A., Vorotnikov A.V., Gracheva E.O., Albert W.C., Panettieri R.A. Jr., Stepanova V.V. and Tkachuk V.A. (2002). Activation of p38 MAP-kinase and caldesmon phosphorylation are essential for urokinase-induced human smooth muscle cell migration. Biol. Chem. 383:115–126
Liu X., Yan S., Zhou T., Terada Y. and Erikson R.L. (2004). The MAP kinase pathway is required for entry into mitosis and cell survival. Oncogene 23:763–776
Van Eyk J.E., Arrell D.K., Foster D.B., Strauss J.D., Heinonen T.Y., Furmaniak-Kazmierczak E., Cote G.P. and Mak A.S. (1998). Different molecular mechanisms for Rho family GTPase-dependent, Ca2+-independent contraction of smooth muscle. J. Biol. Chem. 273: 23433–23439
Guo H., Bryan J. and Wang C.L. (1999). A note on the caldesmon sequence. J. Muscle Res. Cell Motil. 20:725–726
McFawn P.K., Shen L., Vincent S.G., Mak A., Van Eyk J.E. and Fisher J.T. (2003). Calcium-independent contraction and sensitization of airway smooth muscle by p21-activated protein kinase. Am. J. Physiol. Lung Cell Mol. Physiol. 284:L863–L870
Vidal C., Geny B., Melle J., Jandrot-Perrus M. and Fontenay-Roupie M. (2002). Cdc42/Rac1-dependent activation of the p21-activated kinase (PAK) regulates human platelet lamellipodia spreading: implication of the cortical-actin binding protein cortactin. Blood 100: 4462–4469
Helfman D.M. et al. (1999) Caldesmon inhibits nonmuscle cell contractility and interferes with the formation of focal adhesions. Mol. Biol. Cell 10: 3097–3112
Boerner J.L., McManus M.J., Martin G.S., Maihle N.J. (2000). Ras-independent oncogenic transformation by an EGF-receptor mutant. J. Cell Sci. 113: 935–942
Beningo K.A., Dembo M., Kaverina I., Small J.V., Wang Y.L. (2001). Nascent focal adhesions are responsible for the generation of strong propulsive forces in migrating fibroblasts. J. Cell Biol. 153: 881–888
Geiger B., Bershadsky A. (2001). Assembly and mechanosensory function of focal contacts. Curr. Opin. Cell Biol. 13: 584–592
Kaverina I., Krylyshkina O., Small J.V. (2002). Regulation of substrate adhesion dynamics during cell motility. Int. J. Biochem. Cell Biol. 34: 746–761
Zamir E. et al. (2000) Dynamics and segregation of cell-matrix adhesions in cultured fibroblasts. Nat. Cell Biol. 2: 191–196
Cramer L.P., Mitchison T.J. (1995). Myosin is involved in postmitotic cell spreading. J. Cell Biol. 131: 179–189
Verkhovsky A.B., Svitkina T.M. and Borisy G.G. (1995). Myosin II filament assemblies in the active lamella of fibroblasts: their morphogenesis and role in the formation of actin filament bundles. J. Cell Biol. 131: 989–1002
Conrad P.A., Giuliano K.A., Fisher G., Collins K., Matsudaira P.T. and Taylor D.L. (1993). Relative distribution of actin, myosin I, and myosin II during the wound healing response of fibroblasts. J. Cell Biol. 120: 1381–1391
Burridge K. and Chrzanowska-Wodnicka M. (1996). Focal adhesions, contractility, and signaling. Annu. Rev. Cell Dev. Biol. 12: 463–518
Acknowledgement
This work was supported by a grant from NIH to C.-L. A Wang (PO1-AR41637).
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Kordowska, J., Huang, R. & Wang, CL.A. Phosphorylation of caldesmon during smooth muscle contraction and cell migration or proliferation. J Biomed Sci 13, 159–172 (2006). https://doi.org/10.1007/s11373-005-9060-8
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DOI: https://doi.org/10.1007/s11373-005-9060-8