The appearance of acetylcholine receptors triggered by fusion of myoblasts in vitro

Shainberg, Asher; Brik, H.

doi:10.1016/0014-5793(78)80204-x

Cited by 33 publications

(9 citation statements)

References 21 publications

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“…The fact that the induction of the acetylcholine receptor precedes the induction of creative kinase by one day suggests that cells that are initiating the synthesis of creative kinase are already committed to the synthesis of at least one set of muscle-specific proteins ; thus, a cell partially induced for creative kinase is almost fully induced for the acetylcholine receptor . Similar differences in the time of appearance of muscle-specific proteins have been noted, for example, with chick myoblasts (19) . There is similarity between these observations and previous reports on the induction of S-100 protein in C-6 cells.…”

supporting

confidence: 72%

“…The control of the synthesis of specific proteins during myogenesis has been the subject of intensive investigation in various tissue culture systems using myoblasts or established cell lines such as L-6 (1,17,20). Most of these systems use fusing cells, where the events related to cell fusion cannot always be readily separated from the cell recognition events necessary for the induction of muscle-specific proteins (such as creatine kinase and the acetylcholine receptor) (2,7,19,21) . Less information is available regarding the induction of muscle-specific proteins in smooth muscle cells (5) .…”

mentioning

confidence: 99%

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Multiple controls for the synthesis of muscle-specific proteins in BC3H1 cells.

Munson¹,

Caldwell²,

Glaser³

1982

The Journal of Cell Biology

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The regulation of the synthesis of muscle-specific proteins has been examined in BCH1 cells, a smooth muscle-like cell line isolated by Schubert et al . (l . Cell Biol., 1974, 61 : 398-413 .) . The synthesis of both creatine kinase and the acetylcholine receptor appear to be under dual control, a positive control due to cell-cell contact which increases the rate of synthesis of this protein, and a negative signal, elicited by serum components, that decreases the rate of synthesis of these proteins . Induction of muscle-specific proteins in BCH1 cells is a reversible process and can be arrested after partial induction has taken place by the addition of serum or high-molecular-weight protein fraction from serum to these cells. The highmolecular-weight protein fraction from serum is not by itself mitogenic for BCH1 cells and cannot be replaced by a variety of known hormones (mitogenic factors) .BC,H1 cells, a nonfusing muscle cell line, was obtained from Dr. D. Schubert at the Salk Institute. The cells were grown at 37°C in a 10% C02-enriched 350 atmosphere in Dulbecco's modified Eagle's medium (DMEM) supplemented, where indicated, with 15% nutrient medium F12 and the specified concentration of dialyzed fetal calf serum or dialyzed horse serum and L-glutamine (0.1 mg/ ml), penicillin (0 U/ml), and streptomycin (100 /+g/ml) .Cells were transferred from log phase cultures by trypsinization with crystalline trypsin, and replated . Fresh cultures were started from frozen stock at 4-to 6-wk intervals. Media were obtained from Gibco Laboratories (Grand Island Biological Co., Grand Island, NY) or the Washington University Basic Cancer Center. Sera were obtained from Gibco Laboratories and K. C. Biologicals.In some experiments cells were grown on collagen-coated dishes. Dishes were coated by Nlls vapor precipitation of acid-soluble calf skin collagen (Sigma Chemical Co ., St. Louis, MO) (14). The dishes were stored dry after sterilization with UV light. Before use, dishes were washed twice with Hepes-buffered Ca''-and Mg"-free Hanks' solution and once with complete medium .Creatine kinase was determined by the following modification of published procedures (11,20). Cultures in 35-mm dishes were washed once with 2 ml of phosphate-buffered saline (PBS) and the cell layer was suspended in 0.5 ml of 1% nonidet P-40 (NP-40) in 50 glycyl-glycine, pH 6.75. The cells were homogenized (10 to 15 strokes) in a 1-ml Dounce homogenizer. For assay, the extract was appropriately diluted in the same buffer containing 1 mg/ml bovine serum albumin (BSA).A two-stage assay for creatine kinase is used. In the first stage, the reaction mixture contains l mM ADP, 10 mM MgC1% 0.2% BSA, 0.1 mM P',P°diaden-osine, 5' pentaphosphate (Boehringer Mannheim Biochemicals, Indianapolis, IN), 2.5 mM dithiothreitol, 25 mM phosphocreatine, 3 MM D-glucose and 20 U/ ml hexokinase, and extract in a total volume of 40 Ial. Controls lack phosphocreatine . The reaction is initiated by the addition of extract. After incubation at 37°C for 30 min, the rea...

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supporting

confidence: 72%

mentioning

confidence: 99%

Multiple controls for the synthesis of muscle-specific proteins in BC3H1 cells.

Munson¹,

Caldwell²,

Glaser³

1982

The Journal of Cell Biology

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“…The level of acetylcholine receptors (AChR) was measured by incubating intact cultures with 6 x lo-' M '251-a-bungarotoxin (50-150 Ci/mmole) for 1 h in DMEM, as previously described [48]. The cells were then washed six times with 1 ml medium (each wash remaining on the culture dishes for 10 min) and radioactivity determined by laying the dishes on a flat crystal 2-inch-diameter y counter (Elscint, Haifa, Israel).…”

Section: Determination Of Muscle-specific Proteinsmentioning

confidence: 99%

Activation of the interferon system during myogenesis in vitro

et al. 1990

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“…Analysis with specific cDNA clones has shown that during this period, there is a dramatic accumulation of mRNA coding for ankyrin and a-and/~-spectrin (39). Together, these results suggest that transcriptional (or posttranscriptional) control of gene expression plays an important role in regulating the expression of these components of the muscle membrane-skeleton in early myogenesis, as has been shown also for other muscle-specific proteins such as myosin heavy chain (1, 13,15,17,53,57,63), a-actin (15,59,63), tropomyosin (41), and the acetylcholine receptor (16,35,55,62).…”

mentioning

confidence: 71%

Posttranslational control of membrane-skeleton (ankyrin and alpha beta-spectrin) assembly in early myogenesis.

Nelson¹,

Lazarides²

1985

The Journal of Cell Biology

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Adult chicken skeletal muscle cells express polypeptides that are antigenically related to ce-spectrin (Mr 240,000) and/3-spectrin (Mr 220,000-225,000), the major components of the erythrocyte membrane-skeleton, and to ankyrin (Mr 237,000; also termed goblin in chicken erythrocytes), which binds spectrin to the transmembrane anion transporter in erythrocytes. Comparative immunoblotting of SDS-solubilized extracts of presumptive myoblasts and fully differentiated myotubes cultured in vitro demonstrated that there is a dramatic accumulation of ankyrin and c~-and/3-spectrin during myogenesis and a concomitant switch in the subunit composition of spectrin from c~, to c~3. Analysis of early time points in myogenesis (12-96 h) revealed that these changes occur shortly after the main burst of cell fusion. To determine the temporal relationship between cell fusion and the accumulation of ankyrin and ~x-and B-spectrin, we treated presumptive myoblasts with 2 mM EGTA, which resulted in the complete inhibition of cell fusion. The incorporation of [35S]methionine into total protein and, specifically, into ~x-, 3,-, and/3-spectrin remained the same in EGTA-treated and control cells. Analysis by immunoblotting of the amounts of ankyrin and ce-and f3-spectrin in fusion-blocked cells revealed that there was no effect on accumulation for the first 19 h. However, there was then a dramatic cessation in their accumulation, and thereafter, the amount of each protein at steady state remained constant. Upon release from the EGTA block, the cells fused rapidly (< 11 h), and the accumulation of ankyrin and c~-and g3-spectrin was reinitiated after a lag period of 3-5 h at a rate similar to that in control cells. The inhibition in the accumulation of newly synthesized ankyrin, c~-spectrin, and B-spectrin in EGTA-treated myoblasts was not characteristic of all structural proteins, since the accumulation of the muscle-specific intermediate filament protein desmin was the same in control and fusionblocked cells. These results show that in myogenesis, the synthesis of ankyrin and c~-and Bspectrin and their accumulation as a complex, although concurrent, are not coupled events. We hypothesize that the extent of assembly of these components of the membrane-skeleton in muscle cells is determined by a control mechanism(s) operative at the posttranslational level that is triggered near the time of cell fusion and the onset of terminal differentiation.Adult chicken skeletal muscle cells express a class of proteins that are antigenically and structurally related to the major components of the erythrocyte membrane-skeleton. These proteins are a-spectrin (Mr 240,000; references 43 and 56), ~-spectrin (Mr 220,000-225,000; reference 43), and ankyrin (Mr 235,000; reference 46). In erythrocytes, a-and/3-spectrin form an (aj3)2 tetramer that binds actin oligomers (reviewed in references 2 and 10). Ankyrin (termed goblin in chicken erythrocytes; see references 38, 46, and 56) binds spectrin (4, 5; specifically the/3-spectrin subunit [33,40,65]) to ...

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The appearance of acetylcholine receptors triggered by fusion of myoblasts in vitro

Cited by 33 publications

References 21 publications

Multiple controls for the synthesis of muscle-specific proteins in BC3H1 cells.

Multiple controls for the synthesis of muscle-specific proteins in BC3H1 cells.

Activation of the interferon system during myogenesis in vitro

Posttranslational control of membrane-skeleton (ankyrin and alpha beta-spectrin) assembly in early myogenesis.

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