Alteration in the myosin phosphorylation pattern of smooth muscle by phorbol ester

1990 ◽  
Vol 259 (5) ◽  
pp. C769-C774 ◽  
Author(s):  
M. Seto ◽  
Y. Sasaki ◽  
Y. Sasaki
Keyword(s):  
Steady State ◽  
Light Chain ◽  
Myosin Light Chain ◽  
High Concentration ◽  
Tension Development ◽  
Aortic Artery ◽  
Phosphorylation Pattern ◽  
Mlc20 Phosphorylation ◽  
High Concentrations ◽  
The Relationship

Pretreatment with a high concentration of phorbol 12-myristate 13-acetate (PMA, 100 nM) increased the degree of tension and extent of myosin light chain (MLC20) phosphorylation in the K(+)-stimulated rabbit aortic artery. Pretreatment with 100 nM PMA did not alter the relationship between MLC20 phosphorylation and the tension seen with K+ stimulation in the initial phase and steady state of contraction. However, a low concentration of PMA (10 nM) potentiated only the MLC20 phosphorylation during the steady state of contraction with no effect on the tension. In contrast, the prostaglandin (PG) F2 alpha-induced tension development and the MLC20 phosphorylation were not affected by PMA pretreatment at both low and high concentrations. The inhibitory action of nifedipine on the K(+)-induced contraction was not affected by pretreatment with 100 nM PMA; the concentration producing half-maximal inhibition of nifedipine for the K(+)-induced contraction (33 nM) was the same as that of the K+ plus 100 nM PMA-induced contraction (32 nM). Our results suggest that PMA may increase the level of myosin light chain kinase-dependent MLC20 phosphorylation and the tension in the K(+)-stimulated artery, an effect which differs from that seen with increases in K+ concentrations. The regulatory mechanism for the contraction involving PGF2 alpha stimulation may differ from that seen in the case of K+ stimulation.

Halothane inhibits agonist-induced potentiation of rMLC phosphorylation in permeabilized airway smooth muscle

1997 ◽  
Vol 273 (1) ◽  
pp. L80-L85 ◽  
Author(s):  
K. A. Jones ◽  
A. Hirasaki ◽  
D. H. Bremerich ◽  
C. Jankowski ◽  
D. O. Warner
Keyword(s):  
Smooth Muscle ◽  
Steady State ◽  
Vascular Smooth Muscle ◽  
Light Chain ◽  
Myosin Light Chain ◽  
Isometric Force ◽  
State Level ◽  
Steady State Level ◽  
Regulatory Myosin Light Chain ◽  
The Relationship

Agonist-induced increases in CA2+ sensitivity are mediated in part by mechanisms that increase phosphorylation of the regulatory myosin light chain (rMLC) at constant cytosolic Ca2+ concentration ([Ca2+]i). The current study tested the hypothesis that halothane inhibits acetylcholine (ACh)-induced potentiation of rMLC phosphorylation in beta-escin-permeabilized canine tracheal smooth muscle. ACh plus GTP significantly potentiated the increase in isometric force and rMLC phosphorylation induced by 0.8 microM free Ca2+. However, whereas the potentiation of isometric force was sustained, the potentiation of rMLC phosphorylation was biphasic, peaking at 0.5 min and then declining by approximately 10 min to a steady-state level significantly above that induced by 0.8 microM free Ca2+ alone. This finding suggests that mechanisms in addition to changes in rMLC phosphorylation may mediate ACh-induced Ca2+ sensitization, as has been reported for vascular smooth muscle. Halothane (0.91 +/- 0.10 mM) significantly inhibited ACh plus GTP-induced potentiation of rMLC phosphorylation and isometric force after 2 (peak rMLC phosphorylation) and 15 (steady-state rMLC phosphorylation) min of stimulation. However, the effect of halothane on the potentiation of isometric force was significantly less than that expected from its effect on rMLC phosphorylation (i.e., halothane changed the relationship between rMLC phosphorylation and isometric force). These results demonstrate that halothane inhibits the ACh-induced increase in Ca2+ sensitivity by inhibiting the membrane receptor-coupled mechanisms that increase rMLC phosphorylation at constant submaximal [Ca2+]i. Possible additional effects of halothane on rMLC phosphorylation-independent mechanisms cannot be ruled out.

Hypoxia-induced pulmonary arterial contraction appears to be dependent on myosin light chain phosphorylation

1996 ◽  
Vol 271 (5) ◽  
pp. L768-L774 ◽  
Author(s):  
Y. Zhao ◽  
R. A. Rhoades ◽  
C. S. Packer
Keyword(s):  
Smooth Muscle ◽  
Muscle Contraction ◽  
Light Chain ◽  
Myosin Light Chain ◽  
Kinase Inhibitor ◽  
Tension Development ◽  
Arterial Contraction ◽  
Pulmonary Arterial ◽  
Mlc20 Phosphorylation ◽  
Arterial Muscle

The signal transduction pathway of hypoxic pulmonary arterial contraction has not been elucidated. Phosphorylation of the 20-kDa myosin light chain (MLC20) is thought to be essential for vascular muscle contraction. However, there are reports that smooth muscle will contract in response to nonphysiological stimuli such as phorbol esters without the involvement of MLC20 phosphorylation. The purpose of this study was to determine if hypoxia-induced pulmonary arterial contraction is dependent on MLC20 phosphorylation. Isolated rat pulmonary and carotid (for comparative purposes) arterial strips were contracted with 80 mM KCl to establish maximum active tension in response to membrane depolarization. The strips were then stimulated with one of the following: 30 mM KCl, 1 microM phenylephrine, 0.01 microM angiotensin II, 1 microM phorbol 12-myristate 13-acetate (PMA), or hypoxia (95% N2-5% CO2). In some experiments ML-9, a myosin light chain kinase inhibitor, or calphostin C, a protein kinase C (PKC) inhibitor, was introduced into the bath before hypoxia. Isometric tension was recorded as a function of time. Muscle strips were freeze-clamped (liquid N2) at various time points during the course of responses to the various stimuli. MLC20 phosphorylation levels were measured by ureaglycerol gel electrophoresis followed by Western blot procedure. Results show that increased MLC20 phosphorylation correlates with initiation of pulmonary arterial smooth muscle contraction in response to all agonists with the exception of PMA, a known activator of PKC. The MLC20 phosphorylation levels correlate with tension development in response to hypoxia, and ML-9 abolished the hypoxic contractions. In contrast, hypoxia relaxed carotid arterial muscle, and there was a corresponding decrease in the MLC20 phosphorylation level. In conclusion, hypoxia appears to result in MLC20 phosphorylation-mediated contraction in conduit pulmonary arterial muscle and in MLC20 dephosphorylation-mediated relaxation in systemic arterial muscle.

scholarly journals FFAR1 activation attenuates histamine-induced myosin light chain phosphorylation and cortical tension development in human airway smooth muscle cells

2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Shengjie Xu ◽  
Anthony Schwab ◽  
Nikhil Karmacharya ◽  
Gaoyuan Cao ◽  
Joanna Woo ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Light Chain ◽  
Myosin Light Chain ◽  
Airway Hyperreactivity ◽  
Human Airway Smooth Muscle ◽  
Tension Development ◽  
Cortical Tension ◽  
Human Airway ◽  
Mlc Phosphorylation

Abstract Background Activation of free fatty acid receptors (FFAR1 and FFAR4) which are G protein-coupled receptors (GPCRs) with established (patho)physiological roles in a variety of obesity-related disorders, induce human airway smooth muscle (HASM) cell proliferation and shortening. We reported amplified agonist-induced cell shortening in HASM cells obtained from obese lung donors. We hypothesized that FFAR1 modulate excitation–contraction (EC) coupling in HASM cells and play a role in obesity-associated airway hyperresponsiveness. Methods In HASM cells pre-treated (30 min) with FFAR1 agonists TAK875 and GW9508, we measured histamine-induced Ca2+ mobilization, myosin light chain (MLC) phosphorylation, and cortical tension development with magnetic twisting cytometry (MTC). Phosphorylation of MLC phosphatase and Akt also were determined in the presence of the FFAR1 agonists or vehicle. In addition, the effects of TAK875 on MLC phosphorylation were measured in HASM cells desensitized to β2AR agonists by overnight salmeterol treatment. The inhibitory effect of TAK875 on MLC phosphorylation was compared between HASM cells from age and sex-matched non-obese and obese human lung donors. The mean measurements were compared using One-Way ANOVA with Dunnett’s test for multiple group comparisons or Student’s t-test two-group comparison. For cortical tension measurements by magnetic twisted cytometry, mixed effect model using SAS V.9.2 was applied. Means were considered significant when p ≤ 0.05. Results Unexpectedly, we found that TAK875, a synthetic FFAR1 agonist, attenuated histamine-induced MLC phosphorylation and cortical tension development in HASM cells. These physiological outcomes were unassociated with changes in histamine-evoked Ca2+ flux, protein kinase B (AKT) activation, or MLC phosphatase inhibition. Of note, TAK875-mediated inhibition of MLC phosphorylation was maintained in β2AR-desensitized HASM cells and across obese and non-obese donor-derived HASM cells. Conclusions Taken together, our findings identified the FFAR1 agonist TAK875 as a novel bronchoprotective agent that warrants further investigation to treat difficult-to-control asthma and/or airway hyperreactivity in obesity.

scholarly journals Increased Phosphorylation of Myosin Light Chain Prevents in Vitro Decidualization

Endocrinology ◽  
2007 ◽  
Vol 148 (7) ◽  
pp. 3176-3184 ◽  
Author(s):  
Ivanna Ihnatovych ◽  
WenYang Hu ◽  
Jody L. Martin ◽  
Asgerally T. Fazleabas ◽  
Primal de Lanerolle ◽  
...  
Keyword(s):  
Light Chain ◽  
Myosin Light Chain ◽  
Smooth Muscle Actin ◽  
Myosin Ii ◽  
Active Form ◽  
Adenoviral Infection ◽  
Decidual Cells ◽  
Cytoskeletal Dynamics ◽  
Mlc20 Phosphorylation

Differentiation of stromal cells into decidual cells, which is critical to successful pregnancy, represents a complex transformation requiring changes in cytoskeletal architecture. We demonstrate that in vitro differentiation of human uterine fibroblasts into decidual cells includes down-regulation of α-smooth muscle actin and β-tubulin, phosphorylation of focal adhesion kinase, and redistribution of vinculin. This is accompanied by varied adhesion to fibronectin and a modified ability to migrate. Cytoskeletal organization is determined primarily by actin-myosin II interactions governed by the phosphorylation of myosin light chain (MLC20). Decidualization induced by cAMP [with estradiol-17β (E) and medroxyprogesterone acetate (P)] results in a 40% decrease in MLC20 phosphorylation and a 55% decline in the long (214 kDa) form of myosin light-chain kinase (MLCK). Destabilization of the cytoskeleton by inhibitors of MLCK (ML-7) or myosin II ATPase (blebbistatin) accelerates decidualization induced by cAMP (with E and P) but inhibits decidualization induced by IL-1β (with E and P). Adenoviral infection of human uterine fibroblast cells with a constitutively active form of MLCK followed by decidualization stimuli leads to a 30% increase in MLC20 phosphorylation and prevents decidualization. These data provide evidence that the regulation of cytoskeletal dynamics by MLC20 phosphorylation is critical for decidualization.

Expression of the catalytic domain of myosin light chain kinase increases paracellular permeability

1996 ◽  
Vol 271 (5) ◽  
pp. C1678-C1684 ◽  
Author(s):  
G. Hecht ◽  
L. Pestic ◽  
G. Nikcevic ◽  
A. Koutsouris ◽  
J. Tripuraneni ◽  
...  
Keyword(s):  
Tight Junction ◽  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Catalytic Domain ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Epithelial Tight Junction ◽  
Tight Junction Permeability ◽  
Mlc20 Phosphorylation

Contractile events resulting from phosphorylation of the 20-kDa myosin light chain (MLC20) have been implicated in the regulation of epithelial tight junction permeability. To address this question, Madin-Darby canine kidney cells were transfected with a murine leukemia retroviral vector containing DNA encoding either the catalytic domain of myosin light chain kinase (tMK) or the beta-galactosidase gene (beta-gal). Autoradiograms of sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of myosin immunoprecipitated from 32Pi-labeled transfected cells demonstrated that MLC20 phosphorylation was increased 3.1 +/- 0.9-fold in cells expressing tMK compared with cells expressing beta-gal. Phosphopeptide mapping confirmed that myosin light chain kinase was responsible for the increased MLC20 phosphorylation. Transepithelial electrical resistance, a measurement of barrier function, of tMK cell monolayers was consistently < 10% (123 +/- 20 omega.cm2) of that of monolayers comprised of wild-type cells (1,456 +/- 178 omega.cm2) or cells expressing beta-gal (1,452 +/- 174 omega.cm2). Dual 22Na+ and [3H]mannitol flux studies indicated that the decrease in resistance in tMK cells was attributable to increased paracellular flow. These data support the idea that MLC20 phosphorylation by myosin light chain kinase is involved in regulating epithelial tight junction permeability.

Role of myosin phosphorylation in contractility of a platelet aggregate

1985 ◽  
Vol 249 (3) ◽  
pp. C297-C303 ◽  
Author(s):  
M. E. Bromberg ◽  
R. W. Sevy ◽  
J. L. Daniel ◽  
L. Salganicoff
Keyword(s):  
Steady State ◽  
Light Chain ◽  
Myosin Phosphorylation ◽  
Nonmuscle Cells ◽  
Platelet Aggregate ◽  
Steady State Levels ◽  
State Contraction ◽  
The Relationship ◽  
Dose Dependent

The relationship between tension and myosin 20,000-Da light chain phosphorylation in intact nonmuscle cells was investigated using a preparation of thrombin-activated, irreversibly aggregated platelets known as the platelet strip. Steady-state levels of tension generated by the platelet strip were found to be linearly related to the level of myosin phosphorylation. This relationship was observed during dose-dependent relaxation induced by the adenylate cyclase activators prostaglandin (PG) E1 and PGI2, and during contraction induced by ADP, epinephrine, and the prostaglandin endoperoxide analogue U-46619, which did not appreciably alter the basal level of adenosine 3',5'-cyclic monophosphate in the preparation. The fully relaxed platelet strip, in the absence of external Ca2+, was associated with a level of 12% light chain phosphorylation, which increased to 72% on maximal contraction. During both relaxation and contraction, changes in myosin phosphorylation were also found to precede or coincide with tension changes. Furthermore, steady-state contraction induced by ADP was associated with a maintained elevation in the level of myosin phosphorylation. These results support the concept that myosin phosphorylation is an important regulatory mechanism for contractility in platelets.

scholarly journals Separate upstream and convergent downstream pathways of G-protein- and phorbol ester-mediated Ca2+ sensitization of myosin light chain phosphorylation in smooth muscle

1996 ◽  
Vol 318 (2) ◽  
pp. 469-475 ◽  
Author(s):  
Peter E. JENSEN ◽  
Ming C. GONG ◽  
Avril V. SOMLYO ◽  
Andrew P. SOMLYO
Keyword(s):  
Smooth Muscle ◽  
G Protein ◽  
Phorbol Ester ◽  
Myosin Light Chain ◽  
Down Regulation ◽  
Kinase C ◽  
Rabbit Portal Vein ◽  
Mlc20 Phosphorylation ◽  
The Relationship ◽  
G Protein Coupled

The effect of phorbol ester-induced down-regulation of protein kinase C (PKC) on diacylglycerol (sn-1,2-dioctanoylglycerol, diC8)- and G-protein-coupled Ca2+ sensitization and on the relationship between phosphorylation of the regulatory myosin light chains (MLC20) and force during Ca2+ sensitization were investigated in rabbit portal vein (PV), femoral artery (FA) and ileum smooth muscle. The effects of phorbol dibutyrate (PDBu), guanosine 5´-[γ-thio]triphosphate (GTP[S]) and agonists on the membrane versus cytosolic distribution of PKC isoenzymes were also determined. Down-regulation of PKC abolished Ca2+ sensitization of force and the accompanying increases in MLC20 phosphorylation induced by PDBu, as well as Ca2+ sensitization of force by diC8, but not that by GTP[S], aluminum fluoride (AlF4-) or agonists (phenylephrine, endothelin or carbachol). Down-regulation also inhibited the PDBu-, but not the GTP[S]-induced increase in force under Ca2+-free conditions. In ileum, PDBu translocated PKCs α, β1, β2, ϵ and θ to the membrane fraction, and GTP[S] caused a small translocation of PKC-ϵ. Carbachol- and GTP[S]-induced Ca2+ sensitization remained unaffected in down-regulated ileum in which no cytosolic PKC-ϵ was detectable. We conclude that, although both phorbol ester-induced and G-protein-coupled Ca2+ sensitization of force are mediated by increased MLC20 phosphorylation, it is likely that PKCs α, β1, β2, ϵ and θ do not play an essential role in, although they may contribute to, the G-protein-coupled mechanism.

Trace Elements Geochemistry of Soil-Plant System in Puxiong Lead-Zinc Mining Area, Yunnan, China

2013 ◽  
Vol 726-731 ◽  
pp. 239-244
Author(s):  
Juan Wang ◽  
Zheng Hai Wang
Keyword(s):  
Trace Elements ◽  
Correlation Coefficients ◽  
Mining Area ◽  
High Concentration ◽  
Transition Area ◽  
Lead Zinc ◽  
Measured Ratio ◽  
Concentration Factors ◽  
High Concentrations ◽  
The Relationship

This study aims to obtain an overview of trace elements concentrations in rocks, soils and plants from Puxiong lead-zinc mining area in Yunnan, China, and analyze the connection as well. Concentrations of Ba, Cd, Co, Cu, Mn, Pb, Zn, As, Sb, Hg, Sn, Ni and Sr among soils, rocks and three dominant plants in mining area, transition area and background were measured. Ratio parameter, bioconcentration factors (BF) and Pearson's correlation coefficients were used to analyze the relationship between these elements in different feature. The results indicate that rocks, soils and plants samples are all characterized by high concentrations of Pb, Zn and other associated elements in Puxiong lead-zinc mining area. Geochemistry of trace elements concentrations from rocks to soils, then to plants presents obvious characters of inheritance and variability. Whats more, three dominant plants are same in high concentration of Mn and low concentration of Cd, Hg, Co, but are huge different in bio-concentration factors.

Blebbistatin, a myosin II inhibitor, suppresses contraction and disrupts contractile filaments organization of skinned taenia cecum from guinea pig

2010 ◽  
Vol 298 (5) ◽  
pp. C1118-C1126 ◽  
Author(s):  
Masaru Watanabe ◽  
Masatoshi Yumoto ◽  
Hideyuki Tanaka ◽  
Hon Hui Wang ◽  
Takeshi Katayama ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Guinea Pig ◽  
Muscle Contraction ◽  
Light Chain ◽  
Myosin Light Chain ◽  
Myosin Ii ◽  
Smooth Muscle Contraction ◽  
Myosin Light Chain Phosphorylation ◽  
Thin Filaments ◽  
Tension Development

To explore the precise mechanisms of the inhibitory effects of blebbistatin, a potent inhibitor of myosin II, on smooth muscle contraction, we studied the blebbistatin effects on the mechanical properties and the structure of contractile filaments of skinned (cell membrane permeabilized) preparations from guinea pig taenia cecum. Blebbistatin at 10 μM or higher suppressed Ca2+-induced tension development at any given Ca2+ concentration but had little effects on the Ca2+-induced myosin light chain phosphorylation. Blebbistatin also suppressed the 10 and 2.75 mM Mg2+-induced, “myosin light chain phosphorylation-independent” tension development at more than 10 μM. Furthermore, blebbistatin induced conformational change of smooth muscle myosin (SMM) and disrupted arrangement of SMM and thin filaments, resulting in inhibition of actin-SMM interaction irrespective of activation with Ca2+. In addition, blebbistatin partially inhibited Mg2+-ATPase activity of native actomyosin from guinea pig taenia cecum at around 10 μM. These results suggested that blebbistatin suppressed skinned smooth muscle contraction through disruption of structure of SMM by the agent.

Sign in / Sign up

Export Citation Format

Share Document