Cytoskeletal filaments of heart conducting system localized by antibody against a 55,000 dalton protein.

Cow heart conducting cells characteristically contain cytoplasmic intermediate-sized filaments. We report here the preparation of a specific antibody to a 55,000 dalton protein of isolated cow Purkinje fibres. Confirmation has been obtained that these filaments consist of the 55,000 dalton protein, using the indirect immunofluorescence technique. Cross-reaction is seen with vascular endothelium and smooth muscle cells of various origin, suggesting close identity of different types of intermediate-sized filaments.     

Isolation of filaments of the chick lens

A preparation of intermediate filaments isolated from the chicken lens is enriched with a 50,000 dalton polypeptide.     

Isolation of filaments of the chick lens

Summary A preparation of intermediate filaments isolated from the chicken lens is enriched with a 50,000 dalton polypeptide.This study was supported by research grant EY 01417 of the National Institutes of Health, Bethesda, Maryland, USA.     

Purification from a total cellular lysate of a protein modifying the sensitivity of Na+/K+ ATPase to ouabain]

A protein (32,000 dalton) has been purified from M2FS cells derived from the murine plasmocytoma MOPC 173. Like tropomyosin, this protein when added with Ca++ to EDTA-treated plasma membranes prepared from the same cell, induced a drastic increase in the Na+/K+ atpase resistance to ouabain.     

Ultrastructural aspects of the cell fusion induced by Visna virus on sheep choroid plexus cell in culture]

Sheep choroid plexus cells infected with low multiplicities of infection of Visna Virus were stellate and had long and thin processes containing filaments and forming cytoplasmic bridges between adjacent cells. Enlargement of the bridges resulted in the formation of multinucleated cells. Some glycoproteins were clustered on filaments outside the cell. The cytoplasmic changes showed : an intensive protein synthesis; numerous mitochondria closely associated with filaments and some lysosomes and numerous vesicules near the plasma membrane.     

Importance of protein-protein interactions for the structural integrity of membrane framents from Torpedo marmorata electric organ]

Electron spin resonance and electron microscopy were used to show that alkaline extraction of a 43,000 dalton protein from torpedo membrane induces structural destabilization of the membrane and rotational diffusion of the cholinergic receptor protein.     

Spiralig angeordnete Untereinheiten in den A-Filamenten der Kamptozoenmuskulatur

Summary Thick A-Filaments (myosin filaments) of entoproctan muscle cells each consist of 9–11 fibrillar subunits, ca. 30 å in diameter, embedded in a protein matrix of lower electron density (tropomyosin ?). Unlike hitherto described paramyosin filaments, these subunits are regularly arranged in a single circle near the outer edge of each filament. They seem to run in spiral windings around the filaments axis. The protein matrix shows a faint banding along the filament, resembling to the tropomyosin-A pattern but with a much shorter periodicity (ca. 60 å).

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Mit UnterstÜtzung der Deutschen Forschungsgemeinschaft.     

VIP receptors and control of short circuit current in the human intestinal clonal cell line Cl.19A

At the maximally effective concentration of 10 nM, VIP induced a marked (12.5-fold stimulation above basal), and sustained increase in short circuit current in the human intestinal epithelial cell line Cl.19A grown on permeable filters and placed in Ussing chambers. Half-maximal increase of Isc was observed for 0.1 nM VIP. This was well correlated with the VIP-stimulated adenylate cyclase activity (ED50:0.07 nM). Binding studies using 125I-VIP indicated that Cl.19A cells express a peptide-specific VIP receptor with a dissociation constant of 0.07 nM. Covalent labeling of receptors followed by SDS-PAGE analysis of membrane proteins resulted in the identification of a 63,000 dalton binding protein in Cl.19A cells.     

α-Actinin structure and regulation

Alpha-actinin is a cytoskeletal actin-binding protein and a member of the spectrin superfamily, which comprises spectrin, dystrophin and their homologues and isoforms. It forms an anti-parallel rod-shaped dimer with one actin-binding domain at each end of the rod and bundles actin filaments in multiple cell-type and cytoskeleton frameworks. In non-muscle cells, alpha-actinin is found along the actin filaments and in adhesion sites. In striated, cardiac and smooth muscle cells, it is localized at the Z-disk and analogous dense bodies, where it forms a lattice-like structure and stabilizes the muscle contractile apparatus. Besides binding to actin filaments alpha-actinin associates with a number of cytoskeletal and signaling molecules, cytoplasmic domains of transmembrane receptors and ion channels, rendering it important structural and regulatory roles in cytoskeleton organization and muscle contraction. This review reports on the current knowledge on structure and regulation of alpha-actinin.     

Swiprosin-1 modulates actin dynamics by regulating the F-actin accessibility to cofilin

Membrane protrusions, like lamellipodia, and cell movement are dependent on actin dynamics, which are regulated by a variety of actin-binding proteins acting cooperatively to reorganize actin filaments. Here, we provide evidence that Swiprosin-1, a newly identified actin-binding protein, modulates lamellipodial dynamics by regulating the accessibility of F-actin to cofilin. Overexpression of Swiprosin-1 increased lamellipodia formation in B16F10 melanoma cells, whereas knockdown of Swiprosin-1 inhibited EGF-induced lamellipodia formation, and led to a loss of actin stress fibers at the leading edges of cells but not in the cell cortex. Swiprosin-1 strongly facilitated the formation of entangled or clustered F-actin, which remodeled the structural organization of actin filaments making them inaccessible to cofilin. EGF-induced phosphorylation of Swiprosin-1 at Ser183, a phosphorylation site newly identified using mass spectrometry, effectively inhibited clustering of actin filaments and permitted cofilin access to F-actin, resulting in actin depolymerization. Cells overexpressing a Swiprosin-1 phosphorylation-mimicking mutant or a phosphorylation-deficient mutant exhibited irregular membrane dynamics during the protrusion and retraction cycles of lamellipodia. Taken together, these findings suggest that dynamic exchange of Swiprosin-1 phosphorylation and dephosphorylation is a novel mechanism that regulates actin dynamics by modulating the pattern of cofilin activity at the leading edges of cells.     

Solubilization and characterization of a ouabain-sensitive protein from transverse tubule membrane-junctional sarcoplasmic reticulum complexes (TTM-JSR) in cat cardiac muscle

A new glycoprotein of 31,500 dalton, which has a high affinity for ouabain, and is independent of (Na+-K+)-ATPase, was solubilized from transverse tubule membrane and junctional sarcoplasmic reticulum complexes (TTM-JSR) of cat cardiac muscle. This protein could be extracted only in small amounts from sarcolemma-plasma membrane (SLM-PL) fragments, suggesting that it indeed originates from the TTM-JSR.     

Polypeptides of the lens fibre cell intracellular matrix

Summary The cytoskeletal proteins of the vertebrate lens fibre cell comprise polypeptides ranging in mol.wt from 43,000 to 250,000 daltons. The main intermediate filament polypeptide of the pickerel, Northern frog, chick, bovine, and human lens has a mol.wt of 54,000 daltons. Peptide analysis revealed that the chick 54,000 dalton protein was more similar to the bovine protein than to the pickerel protein.Supported by grants EY 01417 (H. Maisel) and HD-06390 (J.C. Brown, University of Virginia, Charlottesville) from the National Institutes of Health, U.S.P.H.S.     

Protein aggregation as primary and characteristic cell reaction to various stresses

Ehrlich carcinoma and EL-4 thymoma ascites cells were subjected in vitro to heat shock, ATP depletion, oxidative stress, Ca²⁺ overlading and iodoacetamide treatment. After the transient stresses, Triton (X-100)-insoluble TIS) fractions were isolated from the cells and analysed by electrophoresis and immunoblotting. All stresses used caused rapid aggregation of cell proteins. This was manifested in a signficant rise in protein content in the TIS fractions. The protein increase was mostly due to and increase in the insolubility of actin, 57 kDa protein of intermediate filaments, 70 kDa heat shock protein (HSP 70), and some specific proteins whose insolubilization was a characteristic sign for each type of cell injury. Different survival rates in the cell lines after either stress corrlated well with differences in their TIS protein accretion. Possible mechanisms for stress-induced protein aggregation and its relationship with cell viability are suggested.     

Solubilization and characterization of a ouabain-sensitive protein from transverse tubule membranejunctional sarcoplasmic reticulum complexes (TTM-JSR) in cat cardiac muscle

Summary A new glycoprotein of 31,500 dalton, which has a high affinity for ouabain, and is independent of (Na⁺–K⁺)-ATPase, was solubilized from transverse tubule membrane and junctional sarcoplasmic reticulum complexes (TTM-JSR) of cat cardiac muscle. This protein could be extracted only in small amounts from sarcolemmaplasma membrane (SLM-PL) fragments, suggesting that it indeed originates from the TTM-JSR.     

Contribution à l'étude de la morphogenèse du thalle duStigeoclonium farctum berth (Chaetophorale)

Summary All inhibitors or any modifications of the cultural conditions lead to partial or total isolation of the cells of filaments ofStigeoclonium farctum. The suppression of the intracellular correlation leads to the ramification of the filaments.     

Fragmentation of myosin A-1 light chain of fast muscle by trypsin]

Limited proteolysis of myosin by such proteolytic enzymes as trypsin, chymotrypsin or papain produces typical fragmentation of its heavy chain. Presently evidence is given that trypsin treatment cleaves the alkali light chain A-1 (20,700 dalton) to a shorter (ca 20,000 dalton) chain. The two "essential" thiols (SH-1 and 2) of moysin were alkylated with 17-C-N-ethylmaleimide and a non-negligible amount of radioactivity was also found in the two alkali light chains. Using the specific radioactivity of alkali light chain A-1 it was possible to identify it among heavy chain fragmentation products. The molecular weight of the newly formed A-1 indicates that limited tryptic cleavage of this A-1 confers on it a closer similarity with alkali light chain A-2.     

Gonadostatin and gonadocrinin, polypeptides of ovarian origin with hypophysiotropic activity]

Crude acetic acid extract of Rat ovaries pretreated with pregnant mare serum (PMSG) contains native peptides with two types of separable biological activities: one, molecular weight greater than 10,000 dalton inhibits the secretion of both LH and FSH as stimulated by luteinizing hormone releasing factor (LRF) in a pituitary monolayer culture system and is referred to as gonadostatin; the other, less than 3,500 dalton, stimulates the secretion of gonadotropins and is designated as gonadocrinin. The biological activities of ovarian gonadocrinin can be competitively inhibited by an LRF-analog-antagonist, D-Phe2, D-Trp6-LRF. These ovarian peptides may participate in physiological control of pituitary LH/FSH secretion.     

Distribution of cytoskeletal elements in cultured skin fibroblasts of patients with Duchenne's Muscular Dystrophy

Summary Cultured fibroblasts from patients suffering from Duchenne's Muscular Dystrophy were examined by indirect immunofluorescent techniques using antibodies against actin, myosin, tubulin, and intermediate-sized filaments. The cells display normal patterns of microfilamentous bundles (stress fibres), microtubules, and intermediate-sized filaments suggesting a normal organization of these cytoskeletal structures.Acknowledgments. This work was supported by the Swiss National Science Foundation, grant Nos 3.445-0.79 and 3.419.78.     

Genetic analysis of myosin II assembly and organization in model organisms

Myosins are a large family of actin-based motor proteins that are involved in a variety of cellular processes. Class II, or conventional, myosins are organized into a number of multi-component structures such as muscle thick filaments, non-muscle filaments and the actomyosin ring during cell division. A number of conditions must be met for the proper assembly and organization of myosin II-containing structures, including the correct stoichiometry of myosin and its associated proteins, and the conformation and regulation of the myosin molecule itself by molecular chaperones and protein kinases. In this review we discuss the use of model organisms in the genetic analysis of the assembly and organization of myosin-containing structures.     

Elektronenmikroskopische Beobachtungen an den Hülsenarteriolen in der Milz des Hundes

Summary The endothelial cells of sheathed arterioles of dog spleen were found to contain an abundance of intracytoplasmic filaments about 80 Å in diameter. The endothelium is surrounded by a fenestrated clastin skeleton. The sheath cells show signs of phagocytosis. A possible vasomotor function of the endothelial cells is discussed.