Phosphorylation of myosin light chain kinase from vascular smooth muscle by cAMP- and cGMP-dependent protein kinases

Hathaway, David R.; Konicki, Mark V.; Coolican, Sharon A.

doi:10.1016/s0022-2828(85)80098-5

Cited by 54 publications

(28 citation statements)

References 48 publications

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“…Hathaway et al (1985), however, showed that cyclic GMP-dependent protein kinase does not phosphorylate an active site on myosin light-chain kinase nor alter its requirement for calmodulin. Our data indicate that cyclic GMP mediates a reduction of calcium influx and of the intracellular release of calcium, implying that it reduces the intracellular calcium concentration available for contraction.…”

Section: Discussionmentioning

confidence: 96%

“…These in turn have been shown to be associated with reduced phosphorylation of myosin light chains (Rapoport, Draznin & Murad, 1983). Activation of cyclic GMP-dependent protein kinase by cyclic GMP has recently been shown to have no influence on the activity parameters of myosin light chain kinase, however (Hathaway, Konicki & Coolican, 1985). The mode of action of these relaxant agents may thus ge primarily through reduction of the level of activating calcium within the cell with secondary dephosphorylation of myosin light chains.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Endothelium‐derived relaxing factor alters calcium fluxes in rabbit aorta: a cyclic guanosine monophosphate‐mediated effect.

Collins¹,

Griffith²,

Henderson³

et al. 1986

The Journal of Physiology

126

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SUMMARY1. Measurement of tension and 45Ca influx and efflux were used to study the effects of endothelium-derived relaxing factor (EDRF), sodium nitroprusside and 8-bromocyclic guanosine monophosphate (GMP) on contractile responses and calcium movements in aortic ring preparations of the rabbit.2. EDRF activity, induced by stimulating endothelium-containing rings with acetylcholine, was associated with relaxation of noradrenaline-constricted rings and with a marked reduction of noradrenaline-stimulated increase in calcium influx.Sodium nitroprusside and 8-bromo-cyclic GMP had a similar effect in deendothelialized preparations.3. EDRF also inhibited noradrenaline-stimulated calcium efflux. Sodium nitroprusside and 8-bromo-cyclic GMP had a similar effect in de-endothelialized preparations, both in the presence and absence of extracellular calcium.4. The vascular smooth muscle relaxant effect of EDRF and of nitrovasodilators may be effected by a cyclic GMP-mediated reduction of cytosolic calcium, through both inhibition of calcium influx and reduction of intracellular calcium release.

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Section: Discussionmentioning

confidence: 96%

Section: Introductionmentioning

confidence: 99%

Endothelium‐derived relaxing factor alters calcium fluxes in rabbit aorta: a cyclic guanosine monophosphate‐mediated effect.

Collins¹,

Griffith²,

Henderson³

et al. 1986

The Journal of Physiology

126

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“…For example, myosin light-chain kinase has been proposed as a phosphorylation target (21)(22)(23). In fact, myosin light-chain kinase is phosphorylated in vitro by cGMP-dependent protein kinase (24,25), but phosphorylation has no effect on the activity of myosin light-chain kinase. To date, no physiologically relevant target for cGMP-dependent protein kinase has been demonstrated in smooth muscle.…”

Section: Methodsmentioning

confidence: 99%

Guanosine 5'-monophosphate modulates gating of high-conductance Ca2+-activated K+ channels in vascular smooth muscle cells.

Williams

Katz

Roy-Contancin

et al. 1988

Proc. Natl. Acad. Sci. U.S.A.

115

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Ca2+-activated K+ channels (PKCa channels) account for the predominant K+ permeability of many types of smooth muscle cells. When activated, they oppose depolarization due to Na+ and Ca2+ channel activity. Several vasodilatory agents that increase intracellular cGMP levels (e.g., nitroprusside, adenosine, and atrial natriuretic factor) enhance the activity of these high-conductance PKCa channels in on-cell patches of bovine aortic smooth muscle cells. In addition, dibutyryl-cGMP (1.0 mM) causes a similar increase in channel activity. To pursue the mechanism of channel modulation by these agents, a series of guanine and adenine nucleotides were evaluated by using inside-out excised patches. Whereas cAMP, AMP, ADP, and ATP were ineffective, all of the corresponding guanine nucleotides potentiated PKCa channel activity when tested at a high concentration (500 jLM). However, only GMP consistently enhanced channel activity in the 1-100 jiM range by increasing the percent open time and frequency of opening of these channels over a wide range of potentials and Ca2l levels without affecting single-channel conductance. Thus, GMP is a potent modulator of PKCa channels and it, rather than cGMP, may mediate the action of the vasodilators examined in this study.Vascular smooth muscle is relaxed by a number ofendogenous and exogenous agents that cause an increase in intracellular guanosine 3',5'-(cyclic)-monophosphate (cGMP) (see ref. 1 for review). The nitrovasodilators (e.g., nitroprusside) and endothelium-derived relaxing factor (EDRF) activate the soluble form of guanylate cyclase (2-4), whereas atrial natriuretic factor (ANF) and adenosine act through receptors on the cell surface to activate the particulate form of this enzyme (5-8).In spite of extensive studies, the mechanism by which cGMP causes smooth muscle to relax has remained elusive. Mechanisms involving phosphorylation of critical regulatory proteins by cGMP-dependent protein kinase have been proposed, but the identity of a physiologically relevant target for this phosphorylation has not been satisfactorily demonstrated. Others (9, 10) have proposed that members of this group of vasodilators cause relaxation due to hyperpolarization of the smooth muscle cell membrane, but the process by which these agents could affect the muscle cell membrane potential has not been determined. However, in the photo-and olfactory-transduction processes in vertebrates, cGMP alters the transmembrane potential by affecting gating of a population of cationselective channels (11,12).Given the abundance of Ca2+-activated K+ channels (PKCa channels) in various types of smooth muscle (13), a process that increases their activity must oppose depolarizing signals, hyperpolarize the membrane, and promote relaxation. We therefore investigated the possibility that elevation of cytosolic cGMP concentration, elicited by a group of vasodilators, could modulate gating of the high-conductance PKCa channels in bovine aortic smooth muscle. By the use of on-cell patch-clamp techniques, the v...

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“…Not surprisingly, it had been proposed that the cGMP-activated kinase (PKG) phosphorylates MLCK and accounts for the Ca 2ϩ desensitizing effect of cGMP. Direct phosphorylation of MLCK by PKG does indeed occur in vitro but not at site A; furthermore, this phosphorylation does not modify the activity of MLCK (21,22). Thus from the biochemical studies, it seems that cGMP is not capable of affecting MLCK activity through its protein kinase PKG.…”

Section: Cgmp Activates Myosin Light Chain Phosphatasementioning

confidence: 99%

Cyclic GMP Causes Ca2+ Desensitization in Vascular Smooth Muscle by Activating the Myosin Light Chain Phosphatase

Lee

Kitazawa

1997

Journal of Biological Chemistry

235

256

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Using permeabilized, arterial smooth muscle strips where membrane-associated pathways remain intact but intracellular Ca 2؉ stores are depleted, we investigated mechanism(s) for the Ca 2؉ desensitization of contractile force by cGMP. The nonhydrolyzable analog 8-bromo-cGMP, when applied to these strips with submaximal Ca 2؉ levels clamped, dramatically and reversibly reduced the steady state levels of phosphorylation at 20-kDa myosin light chain and contractile force, with a nanomolar concentration required to obtain 50% reduction. Supramaximal concentrations of 8-bromocGMP (10 M), however, did not change the steady state relationship between phosphorylation and force. When light chain phosphatase activity was blocked at pCa 6.7, 10 M 8-bromo-cGMP did not affect the rates of rise of light chain phosphorylation and contractile force. When light chain kinase activity was blocked, 10 M 8-bromocGMP significantly accelerated light chain dephosphorylation and force relaxation from the maximal contraction steady state. The light chain phosphorylation time course of a pCa 6.0-induced contraction in the presence of 8-bromo-cGMP exhibited kinetics that are predictable from a mathematical model in which only light chain phosphatase activity is increased. The results of this study strongly suggest that cGMP indirectly activates light chain phosphatase, the first proposed mechanism for cGMP-induced Ca 2؉ desensitization in vasodilatation.The primary mechanisms that regulate smooth muscle contraction and relaxation are, respectively, phosphorylation of the regulatory 20-kDa myosin light chain (MLC 20 ) 1 at Ser-19 by MLC 20 kinase (MLCK) and its dephosphorylation by MLC 20 phosphatase (MLCP) (1, 2). Typically, intracellular Ca 2ϩ levels modulate the MLCK to MLCP activity ratio and ultimately the degree of contractile force because MLCK activity depends on the amount of the Ca 2ϩ -calmodulin complex, which itself hinges on cytosolic Ca 2ϩ levels. In many cases, however, the sensitivity of force to Ca 2ϩ can be changed by physiological modulation of the Ca 2ϩ dependence of MLC 20 phosphorylation (3, 4) or by other mechanisms such as thin filament disinhibition (5, 6).The nucleotide cyclic GMP has emerged as a potent, physiological second messenger involved in both vasodilator action and failure of vasoconstrictor activity. The neighboring endothelium, endogenous circulating hormones, as well as clinically administered nitrovasodilators all function to widen the lumen of vessels by stimulating guanylyl cyclase in vascular smooth muscle to produce cGMP (7). To date, cGMP has been implicated in lowering intracellular Ca 2ϩ (8, 9) and in decreasing sensitivity of the contractile force to Ca 2ϩ (10,11). A myriad of Ca 2ϩ lowering mechanisms have since been reportedly shown (see Ref. 9 for references): increased Ca 2ϩ sequestration, increased Ca 2ϩ efflux, decreased Ca 2ϩ influx through decreased Ca 2ϩ channel activity and through hyperpolarization via increased K ϩ channel activity, and decreased Ca 2ϩ release through antagonism of ...

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Phosphorylation of myosin light chain kinase from vascular smooth muscle by cAMP- and cGMP-dependent protein kinases

Cited by 54 publications

References 48 publications

Endothelium‐derived relaxing factor alters calcium fluxes in rabbit aorta: a cyclic guanosine monophosphate‐mediated effect.

Endothelium‐derived relaxing factor alters calcium fluxes in rabbit aorta: a cyclic guanosine monophosphate‐mediated effect.

Guanosine 5'-monophosphate modulates gating of high-conductance Ca2+-activated K+ channels in vascular smooth muscle cells.

Cyclic GMP Causes Ca2+ Desensitization in Vascular Smooth Muscle by Activating the Myosin Light Chain Phosphatase

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