Mammal-specific, ERK-dependent, Caldesmon Phosphorylation in Smooth Muscle

D’Angelo, Gerard; Graceffa, Philip; Wang, C.-L. Albert; Wrangle, John; Adam, Leonard P.

doi:10.1074/jbc.274.42.30115

Cited by 69 publications

(64 citation statements)

References 26 publications

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“…In cultured smooth muscle cells where dedifferentiation renders expression switchover of many contractile proteins to the non-muscle isoforms, phosphorylation of l-CaD was also detected upon serum stimulation [41]. Such an increase in phosphorylation was markedly reduced by the MEK inhibitor PD98059, but not by the p38 MAPK inhibitor SB203580 [41]. These results strongly point to extracellular signal-regulated kinase (Erk)-induced phosphorylation of l-CaD during cell proliferation.…”

supporting

confidence: 47%

“…The kinase responsible for the in vivo phosphorylation has been identified to be mitogen activated protein kinase (MAPK; see [39,40]). In cultured smooth muscle cells where dedifferentiation renders expression switchover of many contractile proteins to the non-muscle isoforms, phosphorylation of l-CaD was also detected upon serum stimulation [41]. Such an increase in phosphorylation was markedly reduced by the MEK inhibitor PD98059, but not by the p38 MAPK inhibitor SB203580 [41].…”

mentioning

confidence: 84%

“…These results strongly point to extracellular signal-regulated kinase (Erk)-induced phosphorylation of l-CaD during cell proliferation. It has been demonstrated that the sites of phosphorylation are at Ser759 and Ser789 in the mammalian h-CaD sequence [42], which correspond to Ser497 and Ser527 in the l-CaD sequence [41,43].…”

mentioning

confidence: 99%

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Phosphorylation of caldesmon during smooth muscle contraction and cell migration or proliferation

2006

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SummaryThe 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.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 Remodeling of actin cytoskeleton plays a central role in a variety of cellular processes that involve shape change and movement. Malfunction of these processes could lead to pathological consequences, but how actin-mediated motility is regulated is only beginning to be understood. With recent

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Phosphorylation of caldesmon during smooth muscle contraction and cell migration or proliferation

2006

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“…This activity is likely to correspond to a caldesmon kinase activity observed in G 0 /G 1 transition (Watson et al, 1993). Indeed, a recent report has shown a rapid increase in MAPK-dependent phosphorylation of caldesmon upon serum stimulation of cultured smooth muscle cells (D'Angelo et al, 1999). Because the sites of MAPK phosphorylation are a subset of the cdc2 kinase sites (Childs et al, 1992;Adam and Hathaway, 1993;Redwood et al, 1993), such phosphorylation would inhibit, to a certain extent, the activities of wild-type caldesmon, however the mutant caldesmon would not be affected.…”

Section: Discussionmentioning

confidence: 99%

Mutant Caldesmon Lacking cdc2 Phosphorylation Sites Delays M-Phase Entry and Inhibits Cytokinesis

Yamashiro

Chern

Yamakita

et al. 2001

MBoC

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Caldesmon is phosphorylated by cdc2 kinase during mitosis, resulting in the dissociation of caldesmon from microfilaments. To understand the physiological significance of phosphorylation, we generated a caldesmon mutant replacing all seven cdc2 phosphorylation sites with Ala, and examined effects of expression of the caldesmon mutant on M-phase progression. We found that microinjection of mutant caldesmon effectively blocked early cell division of Xenopus embryos. Similar, though less effective, inhibition of cytokinesis was observed with Chinese hamster ovary (CHO) cells microinjected with 7th mutant. When mutant caldesmon was introduced into CHO cells either by protein microinjection or by inducible expression, delay of M-phase entry was observed. Finally, we found that 7th mutant inhibited the disassembly of microfilaments during mitosis. Wild-type caldesmon, on the other hand, was much less potent in producing these three effects. Because mutant caldesmon did not inhibit cyclin B/cdc2 kinase activity, our results suggest that alterations in microfilament assembly caused by caldesmon phosphorylation are important for M-phase progression.

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“…159,160 Phosphorylation of CaD was also detected in cultured smooth muscle cells upon serum stimulation. 161 Such phosphorylation was markedly reduced by the MEK inhibitor PD98059, but not by the p38 MAPK inhibitor SB203580, 161 thus strongly pointing to ERK-induced phosphorylation during cell proliferation. With the use of mass spectrometry it was shown that phosphorylation at Ser759 and Ser789 of mammalian smooth muscle CaD (or Ser497 and Ser527 in nonmuscle CaD; Fig.…”

Section: Phosphorylation As a Regulatory Mechanismmentioning

confidence: 99%

Caldesmon and the Regulation of Cytoskeletal Functions

Wang

2008

Advances in Experimental Medicine and Biology

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Caldesmon (CaD) is an extraordinary actin-binding protein, because in addition to actin, it also binds myosin, calmodulin and tropomyosin. As a component of the smooth muscle and nonmuscle contractile apparatus CaD inhibits the actomyosin ATPase activity and its inhibitory action is modulated by both Ca 2+ and phosphorylation. The multiplicity of binding partners and diverse biochemical properties suggest CaD is a potent and versatile regulatory protein both in contractility and cell motility. However, after decades of investigation in numerous laboratories, hard evidence is still lacking to unequivocally identify its in vivo functions, although indirect evidence is mounting to support an important role in connection with the actin cytoskeleton. This chapter reviews the highlights of the past findings and summarizes the current views on this protein, with emphasis of its interaction with tropomyosin.

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Mammal-specific, ERK-dependent, Caldesmon Phosphorylation in Smooth Muscle

Abstract: Caldesmon is an actin-, tropomyosin-, myosin-, and calmodulin-binding protein that is alternatively spliced to yield either a 93-kDa protein (high molecular mass caldesmon (h-CaD)

Cited by 69 publications

References 26 publications

Phosphorylation of caldesmon during smooth muscle contraction and cell migration or proliferation

Phosphorylation of caldesmon during smooth muscle contraction and cell migration or proliferation

Mutant Caldesmon Lacking cdc2 Phosphorylation Sites Delays M-Phase Entry and Inhibits Cytokinesis

Caldesmon and the Regulation of Cytoskeletal Functions

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