scholarly journals Myosin light chain kinase and Src control membrane dynamics in volume recovery from cell swelling

2011 ◽  
Vol 22 (5) ◽  
pp. 634-650 ◽  
Author(s):  
Elisabeth T. Barfod ◽  
Ann L. Moore ◽  
Benjamin G. Van de Graaf ◽  
Steven D. Lidofsky
Keyword(s):  
Cell Volume ◽  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Myosin Ii ◽  
Membrane Dynamics ◽  
Cell Swelling ◽  
Membrane Turnover ◽  
Osmotic Swelling ◽  
Volume Recovery

 The expansion of the plasma membrane, which occurs during osmotic swelling of epithelia, must be retrieved for volume recovery, but the mechanisms are unknown. Here we have identified myosin light chain kinase (MLCK) as a regulator of membrane internalization in response to osmotic swelling in a model liver cell line. On hypotonic exposure, we found that there was time-dependent phosphorylation of the MLCK substrate myosin II regulatory light chain. At the sides of the cell, MLCK and myosin II localized to swelling-induced membrane blebs with actin just before retraction, and MLCK inhibition led to persistent blebbing and attenuated cell volume recovery. At the base of the cell, MLCK also localized to dynamic actin-coated rings and patches upon swelling, which were associated with uptake of the membrane marker FM4-64X, consistent with sites of membrane internalization. Hypotonic exposure evoked increased biochemical association of the cell volume regulator Src with MLCK and with the endocytosis regulators cortactin and dynamin, which colocalized within these structures. Inhibition of either Src or MLCK led to altered patch and ring lifetimes, consistent with the concept that Src and MLCK form a swelling-induced protein complex that regulates volume recovery through membrane turnover and compensatory endocytosis under osmotic stress.

scholarly journals Myosin light chain kinase-regulated endothelial cell contraction: the relationship between isometric tension, actin polymerization, and myosin phosphorylation.

1995 ◽  
Vol 130 (3) ◽  
pp. 613-627 ◽  
Author(s):  
Z M Goeckeler ◽  
R B Wysolmerski
Keyword(s):  
Endothelial Cell ◽  
Light Chain ◽  
Isometric Contraction ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Myosin Ii ◽  
Isometric Tension ◽  
Stress Fibers ◽  
Light Chains ◽  
Myosin Light Chains

The phosphorylation of regulatory myosin light chains by the Ca2+/calmodulin-dependent enzyme myosin light chain kinase (MLCK) has been shown to be essential and sufficient for initiation of endothelial cell retraction in saponin permeabilized monolayers (Wysolmerski, R. B. and D. Lagunoff. 1990. Proc. Natl. Acad. Sci. USA. 87:16-20). We now report the effects of thrombin stimulation on human umbilical vein endothelial cell (HUVE) actin, myosin II and the functional correlate of the activated actomyosin based contractile system, isometric tension development. Using a newly designed isometric tension apparatus, we recorded quantitative changes in isometric tension from paired monolayers. Thrombin stimulation results in a rapid sustained isometric contraction that increases 2- to 2.5-fold within 5 min and remains elevated for at least 60 min. The phosphorylatable myosin light chains from HUVE were found to exist as two isoforms, differing in their molecular weights and isoelectric points. Resting isometric tension is associated with a basal phosphorylation of 0.54 mol PO4/mol myosin light chain. After thrombin treatment, phosphorylation rapidly increases to 1.61 mol PO4/mol myosin light chain within 60 s and remains elevated for the duration of the experiment. Myosin light chain phosphorylation precedes the development of isometric tension and maximal phosphorylation is maintained during the sustained phase of isometric contraction. Tryptic phosphopeptide maps from both control and thrombin-stimulated cultures resolve both monophosphorylated Ser-19 and diphosphorylated Ser-19/Thr-18 peptides indicative of MLCK activation. Changes in the polymerization of actin and association of myosin II correlate temporally with the phosphorylation of myosin II and development of isometric tension. Activation results in a 57% increase in F-actin content within 90 s and 90% of the soluble myosin II associates with the reorganizing F-actin. Furthermore, the disposition of actin and myosin II undergoes striking reorganization. F-actin initially forms a fine network of filaments that fills the cytoplasm and then reorganizes into prominent stress fibers. Myosin II rapidly forms discrete aggregates associated with the actin network and by 2.5 min assumes a distinct periodic distribution along the stress fibers.

scholarly journals Slit and Netrin-1 guide cranial motor axon pathfinding via Rho-kinase, myosin light chain kinase and myosin II

2010 ◽  
Vol 5 (1) ◽  
pp. 16 ◽  
Author(s):  
Ailish Murray ◽  
Arifa Naeem ◽  
Sarah H Barnes ◽  
Uwe Drescher ◽  
Sarah Guthrie
Keyword(s):  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Myosin Ii ◽  
Rho Kinase ◽  
Motor Axon ◽  
Axon Pathfinding

scholarly journals Non-muscle Myosin II and Myosin Light Chain Kinase Are Downstream Targets for Vasopressin Signaling in the Renal Collecting Duct

2004 ◽  
Vol 279 (47) ◽  
pp. 49026-49035 ◽  
Author(s):  
Chung-Lin Chou ◽  
Birgitte M. Christensen ◽  
Sebastian Frische ◽  
Henrik Vorum ◽  
Ravi A. Desai ◽  
...  
Keyword(s):  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Collecting Duct ◽  
Myosin Ii ◽  
Downstream Targets ◽  
Renal Collecting Duct ◽  
Muscle Myosin

scholarly journals Localization and Activity of Myosin Light Chain Kinase Isoforms during the Cell Cycle

2000 ◽  
Vol 151 (3) ◽  
pp. 697-708 ◽  
Author(s):  
Angela Poperechnaya ◽  
Olga Varlamova ◽  
Pei-ju Lin ◽  
James T. Stull ◽  
Anne R. Bresnick
Keyword(s):  
Molecular Weight ◽  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Fluorescent Protein ◽  
Temporal Dynamics ◽  
Myosin Ii ◽  
Amino Acid Sequences ◽  
Relative Localization ◽  
Nonmuscle Cells

Phosphorylation on Ser 19 of the myosin II regulatory light chain by myosin light chain kinase (MLCK) regulates actomyosin contractility in smooth muscle and vertebrate nonmuscle cells. The smooth/nonmuscle MLCK gene locus produces two kinases, a high molecular weight isoform (long MLCK) and a low molecular weight isoform (short MLCK), that are differentially expressed in smooth and nonmuscle tissues. To study the relative localization of the MLCK isoforms in cultured nonmuscle cells and to determine the spatial and temporal dynamics of MLCK localization during mitosis, we constructed green fluorescent protein fusions of the long and short MLCKs. In interphase cells, localization of the long MLCK to stress fibers is mediated by five DXRXXL motifs, which span the junction of the NH2-terminal extension and the short MLCK. In contrast, localization of the long MLCK to the cleavage furrow in dividing cells requires the five DXRXXL motifs as well as additional amino acid sequences present in the NH2-terminal extension. Thus, it appears that nonmuscle cells utilize different mechanisms for targeting the long MLCK to actomyosin structures during interphase and mitosis. Further studies have shown that the long MLCK has twofold lower kinase activity in early mitosis than in interphase or in the early stages of postmitotic spreading. These findings suggest a model in which MLCK and the myosin II phosphatase (Totsukawa, G., Y. Yamakita, S. Yamashiro, H. Hosoya, D.J. Hartshorne, and F. Matsumura. 1999. J. Cell Biol. 144:735–744) act cooperatively to regulate the level of Ser 19–phosphorylated myosin II during mitosis and initiate cytokinesis through the activation of myosin II motor activity.

Na+-K+-2Cl−cotransport in Ehrlich cells: regulation by protein phosphatases and kinases

1998 ◽  
Vol 275 (1) ◽  
pp. C239-C250 ◽  
Author(s):  
Thomas Krarup ◽  
Lene D. Jakobsen ◽  
Bo S. Jensen ◽  
Else K. Hoffmann
Keyword(s):  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Cell Swelling ◽  
Cell Shrinkage ◽  
Volume Increase ◽  
Ehrlich Cells ◽  
Dependent Inhibition ◽  
Cotransport System

To identify protein kinases (PK) and phosphatases (PP) involved in regulation of the Na+-K+-2Cl−cotransporter in Ehrlich cells, the effect of various PK and PP inhibitors was examined. The PP-1, PP-2A, and PP-3 inhibitor calyculin A (Cal-A) was a potent activator of Na+-K+-2Cl−cotransport (EC50 = 35 nM). Activation by Cal-A was rapid (<1 min) but transient. Inactivation is probably due to a 10% cell swelling and/or the concurrent increase in intracellular Cl− concentration. Cell shrinkage also activates the Na+-K+-2Cl−cotransport system. Combining cell shrinkage with Cal-A treatment prolonged the cotransport activation compared with stimulation with Cal-A alone, suggesting PK stimulation by cell shrinkage. Shrinkage-induced cotransport activation was pH and Ca2+/calmodulin dependent. Inhibition of myosin light chain kinase by ML-7 and ML-9 or of PKA by H-89 and KT-5720 inhibited cotransport activity induced by Cal-A and by cell shrinkage, with IC50 values similar to reported inhibition constants of the respective kinases in vitro. Cell shrinkage increased the ML-7-sensitive cotransport activity, whereas the H-89-sensitive activity was unchanged, suggesting that myosin light chain kinase is a modulator of the Na+-K+-2Cl−cotransport activity during regulatory volume increase.

scholarly journals Distinct Roles of Smooth Muscle and Non-muscle Myosin Light Chain-Mediated Smooth Muscle Contraction

2020 ◽  
Vol 11 ◽  
Author(s):  
Jie Sun ◽  
Yan-Ning Qiao ◽  
Tao Tao ◽  
Wei Zhao ◽  
Li-Sha Wei ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Initial Phase ◽  
Myosin Light Chain ◽  
Myosin Ii ◽  
Smooth Muscle Contraction ◽  
Myosin Regulatory Light Chain ◽  
Gene Encoding ◽  
Muscle Myosin

Both smooth muscle (SM) and non-muscle (NM) myosin II are expressed in hollow organs such as the bladder and uterus, but their respective roles in contraction and corresponding physiological functions remain to be determined. In this report, we assessed their roles by analyzing mice deficient of Myl9, a gene encoding the SM myosin regulatory light chain (SM RLC). We find that global Myl9-deficient bladders contracted with an apparent sustained phase, despite no initial phase. This sustained contraction was mediated by NM myosin RLC (NM RLC) phosphorylation by myosin light chain kinase (MLCK). NM myosin II was expressed abundantly in the uterus and young mice bladders, of which the force was accordingly sensitive to NM myosin inhibition. Our findings reveal distinct roles of SM RLC and NM RLC in SM contraction.

scholarly journals A Global, Myosin Light Chain Kinase-dependent Increase in Myosin II Contractility Accompanies the Metaphase–Anaphase Transition in Sea Urchin Eggs

2006 ◽  
Vol 17 (9) ◽  
pp. 4093-4104 ◽  
Author(s):  
Amy Lucero ◽  
Christianna Stack ◽  
Anne R. Bresnick ◽  
Charles B. Shuster
Keyword(s):  
Sea Urchin ◽  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Myosin Ii ◽  
Rho Kinase ◽  
Specific Cell ◽  
Spatial Cues ◽  
Sea Urchin Eggs ◽  
Cortical Contractility

Myosin II is the force-generating motor for cytokinesis, and although it is accepted that myosin contractility is greatest at the cell equator, the temporal and spatial cues that direct equatorial contractility are not known. Dividing sea urchin eggs were placed under compression to study myosin II-based contractile dynamics, and cells manipulated in this manner underwent an abrupt, global increase in cortical contractility concomitant with the metaphase–anaphase transition, followed by a brief relaxation and the onset of furrowing. Prefurrow cortical contractility both preceded and was independent of astral microtubule elongation, suggesting that the initial activation of myosin II preceded cleavage plane specification. The initial rise in contractility required myosin light chain kinase but not Rho-kinase, but both signaling pathways were required for successful cytokinesis. Last, mobilization of intracellular calcium during metaphase induced a contractile response, suggesting that calcium transients may be partially responsible for the timing of this initial contractile event. Together, these findings suggest that myosin II-based contractility is initiated at the metaphase–anaphase transition by Ca2+-dependent myosin light chain kinase (MLCK) activity and is maintained through cytokinesis by both MLCK- and Rho-dependent signaling. Moreover, the signals that initiate myosin II contractility respond to specific cell cycle transitions independently of the microtubule-dependent cleavage stimulus.

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