Actin isoform compartments in chicken gizzard smooth muscle cells

1994 ◽  
Vol 107 (3) ◽  
pp. 445-455 ◽  
Author(s):  
A.J. North ◽  
M. Gimona ◽  
Z. Lando ◽  
J.V. Small
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Contractile Apparatus ◽  
Thin Filaments ◽  
Chicken Gizzard ◽  
Cytoplasmic Actin ◽  
Dense Bodies ◽  
Beta Actin ◽  
Double Label

Differentiated smooth muscle cells typically contain a mixture of muscle (alpha and gamma) and cytoplasmic (beta and gamma) actin isoforms. Of the cytoplasmic actins the beta-isoform is the more dominant, making up from 10% to 30% of the total actin complement. Employing an antibody raised against the N-terminal peptide specific to beta-actin, which labels only the beta-isoform on two-dimensional gel immunoblots, we have shown that this isoform has a restricted localisation in smooth muscle. Using double-label immunofluorescence and immunoelectron microscopy of ultrathin sections of chicken gizzard, beta-actin was localised in the dense bodies and in longitudinal channels linking consecutive dense bodies that were also occupied by desmin. It was additionally found in the membrane-associated dense plaques, but was excluded from the actomyosin-containing regions of the contractile apparatus. Taken together with earlier results these findings identify a cytoskeletal compartment containing intermediate filaments, cytoplasmic actin and the actin cross-linking protein filamin. Using an antibody specific only for muscle actin, labelling was found generally around the myosin filaments of the contractile apparatus, but was absent from the core of the dense bodies that contained beta-actin. Thus, if dense bodies act as dual-purpose anchorage sites, for the cytoskeletal actin and the contractile actin, the thin filaments of the contractile apparatus must be anchored at the periphery of the dense bodies. A model of the structural organisation of the cell is presented and the possible roles of the cytoskeleton are discussed.

scholarly journals Immunoelectron microscope studies of membrane-microfilament interactions: distributions of alpha-actinin, tropomyosin, and vinculin in intestinal epithelial brush border and chicken gizzard smooth muscle cells.

1981 ◽  
Vol 91 (3) ◽  
pp. 614-628 ◽  
Author(s):  
B Geiger ◽  
A H Dutton ◽  
K T Tokuyasu ◽  
S J Singer
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Brush Border ◽  
Muscle Cells ◽  
Ultrastructural Localization ◽  
Cytoskeletal Proteins ◽  
Junctional Complex ◽  
Intestinal Epithelial ◽  
Chicken Gizzard ◽  
Dense Bodies

The ultrastructural localization of three cytoskeletal proteins, alpha-actinin, tropomyosin, and vinculin, in the brush border of epithelial cells of chicken small intestine and the smooth muscle cells of chicken gizzard was studied by immunofluorescence and immunonelectron microscope labeling of frozen sections of lightly fixed, intact tissues. In the immunoelectron microscope studies, a recently described new type of electron-dense antibody conjugate, imposil-antibody, has been successfully used, along with ferritin-antibody conjugates, in single and double immunolabeling experiments. In the intestinal brush border shows that vinvulin is sharply confined to the junctional complex close to the membrane region of the zonula adherens, in distinct contrast to the more diffuse distributions of the other two proteins. In the smooth muscle cells, the labeling patterns show that vinculin is sharply confined to the membrane-associated dense plaques, closer to the membrane than the alpha-Actinin is also present in the cytoplastic dense bodies, from which vinculin is absent. Tropomyosin is present diffusely distributed in the cytoplasm, but absent from both dense plaques and dense bodies. These findings with the muscle cells demonstrate, therefore, that the dense plaques and dense bodies are chemically and structurally distinct entities. The results with both tissues, along with those in previous papers (Geiger, 1979, Cell. 18:193-205.; Geiger et al., 1980, Proc. Natl. Acad. Sci. U. S. A. 77:4127-4131), suggest that vinculin may play an important and widespread role in the linkage of actin-containing microfilament bundles to membranes.

Calponin is localised in both the contractile apparatus and the cytoskeleton of smooth muscle cells

1994 ◽  
Vol 107 (3) ◽  
pp. 437-444 ◽  
Author(s):  
A.J. North ◽  
M. Gimona ◽  
R.A. Cross ◽  
J.V. Small
Keyword(s):  
Smooth Muscle ◽  
High Resolution ◽  
Cell Surface ◽  
Thin Filament ◽  
Receptor Protein ◽  
Contractile Apparatus ◽  
Thin Filaments ◽  
Cytoplasmic Actin ◽  
Dense Bodies ◽  
Dual Label

Calponin and caldesmon are two thin filament-binding proteins found in smooth muscle that have both been attributed a role in modulating the interaction of actin and myosin. Using high-resolution dual-label immunocytochemistry we have determined the distribution of calponin relative to the contractile and cytoskeletal compartments of the smooth muscle cell. We show, using chicken gizzard smooth muscle, that calponin occurs in the cytoskeleton, with beta-cytoplasmic actin, filamin and desmin, as well as in the contractile apparatus, with myosin and caldesmon. According to the observed labelling intensities, calponin was more concentrated in the cytoskeleton and it was additionally localised in the cytoplasmic dense bodies as well as in the adhesion plaques at the cell surface, which both harbour the beta-cytoplasmic isoform of actin. It is probable that these results explain earlier conflicting reports on the composition of smooth muscle thin filaments and suggest that calponin, together with a Ca(2+)-receptor protein, could just as likely serve a role in the cytoskeleton of smooth muscle as in the contractile apparatus.

Models of contractile units and their assembly in smooth muscle

2005 ◽  
Vol 83 (10) ◽  
pp. 825-831 ◽  
Author(s):  
Farah Ali ◽  
Peter D Paré ◽  
Chun Y Seow
Keyword(s):  
Smooth Muscle ◽  
Striated Muscle ◽  
Dense Body ◽  
Unit Length ◽  
Muscle Length ◽  
Thick Filament ◽  
Contractile Apparatus ◽  
Thin Filaments ◽  
Dense Bodies ◽  
In Series

It is believed that the contractile filaments in smooth muscle are organized into arrays of contractile units (similar to the sarcomeric structure in striated muscle), and that such an organization is crucial for transforming the mechanical activities of actomyosin interaction into cell shortening and force generation. Details of the filament organization, however, are still poorly understood. Several models of contractile filament architecture are discussed here. To account for the linear relationship observed between the force generated by a smooth muscle and the muscle length at the plateau of an isotonic contraction, a model of contractile unit is proposed. The model consists of 2 dense bodies with actin (thin) filaments attached, and a myosin (thick) filament lying between the parallel thin filaments. In addition, the thick filament is assumed to span the whole contractile unit length, from dense body to dense body, so that when the contractile unit shortens, the amount of overlap between the thick and thin filaments (i.e., the distance between the dense bodies) decreases in exact proportion to the amount of shortening. Assembly of the contractile units into functional contractile apparatus is assumed to involve a group of cells that form a mechanical syncytium. The contractile apparatus is assumed malleable in that the number of contractile units in series and in parallel can be altered to accommodate strains on the muscle and to maintain the muscle's optimal mechanical function.Key words: contraction model, ultrastructure, length adaptation, plasticity.

scholarly journals Intermediate-sized filaments of the prekeratin type in myoepithelial cells.

1980 ◽  
Vol 84 (3) ◽  
pp. 633-654 ◽  
Author(s):  
W W Franke ◽  
E Schmid ◽  
C Freudenstein ◽  
B Appelhans ◽  
M Osborn ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Immunofluorescence Microscopy ◽  
Myogenic Differentiation ◽  
Muscle Cells ◽  
Myoepithelial Cells ◽  
Sweat Glands ◽  
Dense Bodies ◽  
Cell Specialization

Myoepithelial cells from mammary glands, the modified sweat glands of bovine muzzle, and salivary glands have been studied by electron microscopy and by immunofluorescence microscopy in frozen sections in an attempt to further characterize the type of intermediate-sized filaments present in these cells. Electron microscopy has shown that all myoepithelial cells contain extensive meshworks of intermediate-sized (7--11-nm) filaments, many of which are anchored at typical desmosomes or hemidesmosomes. The intermediate-sized filaments are also intimately associated with masses of contractile elements, identified as bundles of typical 5--6-nm microfilaments and with characteristically spaced dense bodies. This organization resembles that described for various smooth muscle cells. In immunofluorescence microscopy, using antibodies specific for the various classes of intermediate-sized filaments, the myoepithelial cells are strongly decorated by antibodies to prekeratin. They are not specifically stained by antibodies to vimentin, which stain mesenchymal cells, nor by antibodies to chick gizzard desmin, which decorate fibrils in smooth muscle Z bands and intercalated disks in skeletal and cardiac muscle of mammals. Myoepithelial cells are also strongly stained by antibodies to actin. The observations show (a) that the epithelial character, as indicated by the presence of intermediate-sized filaments of the prekeratin type, is maintained in the differentiated contractile myoepithelial cell, and (b) that desmin and desmin-containing filaments are not generally associated with musclelike cell specialization for contraction but are specific to myogenic differentiation. The data also suggest that in myoepithelial cells prekeratin filaments are arranged--and might function--in a manner similar to the desmin filaments in smooth muscle cells.

Differentiation of smooth muscle cells in chicken gizzard

2002 ◽  
Vol 307 (2) ◽  
pp. 211-223
Author(s):  
Takefumi Kofuji ◽  
Akio Inoue
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Chicken Gizzard

A 36-kDa Protein of the Dense Bodies of Smooth Muscle Cells

1994 ◽  
Vol 116 (6) ◽  
pp. 1354-1359 ◽  
Author(s):  
Kazuyo Ohashi ◽  
Mari Nishimura ◽  
Asako Goi Terasaki ◽  
Hiroyuki Nakagawa
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Dense Bodies

scholarly journals Smitin, a novel smooth muscle titin–like protein, interacts with myosin filaments in vivo and in vitro

2002 ◽  
Vol 156 (1) ◽  
pp. 101-112 ◽  
Author(s):  
Kyoungtae Kim ◽  
Thomas C.S. Keller
Keyword(s):  
Smooth Muscle ◽  
Ionic Strength ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Smooth Muscle Myosin ◽  
Globular Domain ◽  
Contractile Apparatus ◽  
Myosin Filaments ◽  
Muscle Myosin

Smooth muscle cells use an actin–myosin II-based contractile apparatus to produce force for a variety of physiological functions, including blood pressure regulation and gut peristalsis. The organization of the smooth muscle contractile apparatus resembles that of striated skeletal and cardiac muscle, but remains much more poorly understood. We have found that avian vascular and visceral smooth muscles contain a novel, megadalton protein, smitin, that is similar to striated muscle titin in molecular morphology, localization in a contractile apparatus, and ability to interact with myosin filaments. Smitin, like titin, is a long fibrous molecule with a globular domain on one end. Specific reactivities of an anti-smitin polyclonal antibody and an anti-titin monoclonal antibody suggest that smitin and titin are distinct proteins rather than differentially spliced isoforms encoded by the same gene. Smitin immunofluorescently colocalizes with myosin in chicken gizzard smooth muscle, and interacts with two configurations of smooth muscle myosin filaments in vitro. In physiological ionic strength conditions, smitin and smooth muscle myosin coassemble into irregular aggregates containing large sidepolar myosin filaments. In low ionic strength conditions, smitin and smooth muscle myosin form highly ordered structures containing linear and polygonal end-to-end and side-by-side arrays of small bipolar myosin filaments. We have used immunogold localization and sucrose density gradient cosedimentation analyses to confirm association of smitin with both the sidepolar and bipolar smooth muscle myosin filaments. These findings suggest that the titin-like protein smitin may play a central role in organizing myosin filaments in the contractile apparatus and perhaps in other structures in smooth muscle cells.

Unique geometry of actin-membrane anchorage sites in avian gizzard smooth muscle cells

1989 ◽  
Vol 94 (4) ◽  
pp. 703-711
Author(s):  
A. Draeger ◽  
E.H. Stelzer ◽  
M. Herzog ◽  
J.V. Small
Keyword(s):  
Smooth Muscle ◽  
Cell Surface ◽  
Smooth Muscle Cells ◽  
Laser Scanning ◽  
Smooth Muscles ◽  
Muscle Cells ◽  
Laser Scanning Microscopy ◽  
Surface Pattern ◽  
Confocal Laser ◽  
Dense Bodies

Adherens junctions in isolated avian gizzard smooth muscle cells appear as short longitudinal streaks or chevrons that are arranged in periodic, mainly transverse bands along the cell surface. This barrel-like geometry, revealed by antibodies to either vinculin or talin, was seen also in teased gizzard strips by confocal laser-scanning microscopy and contrasted with the rib-like surface pattern observed here and previously in other avian and mammalian smooth muscles. There were on average 67 transverse bands per gizzard cell and an estimated total of around 800 vinculin/talin sites. The longitudinal spacing between the transverse bands of vinculin streaks in the gizzard cells changed from 4–5 microns in extended cells to around 1 micron in shortened cells and the bands remained essentially transverse at all cell lengths, inconsistent with a screw-like mode of cell shortening as has been invoked for smooth muscle cells by others. The absence of rotation on shortening was confirmed by observations on isolated and bead-decorated skinned cells that were induced to contract with ATP. Counterlabelling of cells with alpha-actinin antibodies produced more or less exclusive staining of the cytoplasmic dense bodies, and little surface label: the total number of dense bodies per cell, estimated from confocal microscope through focal series was in the range of 3000. The data are consistent with a periodic anchorage of actin filaments to the cell surface and, in turn, with the existence of regularly spaced contractile assemblies.

Intercellular calcium signaling and nitric oxide feedback during constriction of rabbit renal afferent arterioles

2007 ◽  
Vol 292 (4) ◽  
pp. F1124-F1131 ◽  
Author(s):  
T. R. Uhrenholt ◽  
J. Schjerning ◽  
P. M. Vanhoutte ◽  
B. L. Jensen ◽  
O. Skøtt
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Smooth Muscle ◽  
Endothelial Cell ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Calcium Sensitivity ◽  
No Production ◽  
Contractile Apparatus ◽  
Afferent Arterioles

Vasoconstriction and increase in the intracellular calcium concentration ([Ca2+]i) of vascular smooth muscle cells may cause an increase of endothelial cell [Ca2+]i, which, in turn, augments nitric oxide (NO) production and inhibits smooth muscle cell contraction. This hypothesis was tested in microperfused rabbit renal afferent arterioles, using fluorescence imaging microscopy with the calcium-sensitive dye fura-2 and the NO-sensitive dye 4-amino-5-methylamino-2′,7′-difluorescein. Both dyes were loaded into smooth muscle and endothelium. Depolarization with 100 mmol/l KCl led to a transient vasoconstriction which was converted into a sustained response by N-nitro-l-arginine methyl ester (l-NAME). Depolarization increased smooth muscle cell [Ca2+]ifrom 162 ± 15 nmol/l to a peak of 555 ± 70 nmol/l ( n = 7), and this response was inhibited by 80% by the l-type calcium channel blocker calciseptine. After a delay of 10 s, [Ca2+]iincreased in endothelial cells immediately adjacent to reactive smooth muscle cells, and this calcium wave spread in a nonregenerative fashion laterally into the endothelial cell layer with a velocity of 1.2 μm/s. Depolarization with 100 mmol/l KCl led to a significant increase in NO production ([NO]i) which was inhibited by l-NAME ( n = 5). Acetylcholine caused a rapid increase in endothelial [Ca2+]i, which did not transfer to the smooth muscle cells. l-NAME treatment did not affect changes in smooth muscle [Ca2+]iafter depolarization, but it did increase the calcium sensitivity of the contractile apparatus. We conclude that depolarization increases smooth muscle [Ca2+]iwhich is transferred to the endothelial cells and stimulates NO production which curtails vasoconstriction by reducing the calcium sensitivity of the contractile apparatus.

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