Myosin Messenger RNA: Studies on Its Purification, Properties and Translation during Myogenesis in Culture

Morris, Glenn E.; Buzash, Elizabeth A.; Rourke, A. W.; Tepperman, Katherine; Thompson, William C.; Heywood, Stuart M.

doi:10.1101/sqb.1973.037.01.064

Cited by 52 publications

(24 citation statements)

References 0 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The apparent molecular weight of the estrogen-induced RNA obtained by gel electrophoresis under non-denaturating conditions (2.0 x lo6) is considerably higher than that determined by sedimentation on dodecylsulphate-sucrose gradients (1.5 x lo6). This feature was already observed for other poly(A)-containing RNA's such as myosin-mRNA [23] and ovalbumin mRNA [24]. Since all these separations are very dependent on the secondary structure of the RNA, the molecular weights determined by reference to rRNA are misleading.…”

Section: Discussionmentioning

confidence: 94%

Size, Complexity and Abundance of a Specific Poly(A)‐Containing RNA of Liver from Male Xenopus Induced to Vitellogenin Synthesis by Estrogen

Wahli

Wyler

Weber

et al. 1976

European Journal of Biochemistry

View full text Add to dashboard Cite

Estrogen treatment of Xenopus males leads to the appearance of a new species of poly(A)-containing RNA in the liver, at a stage when large amounts of the estrogen-induced yolk precursor protein, vitellogenin, is produced. This estrogen-induced RNA sediments at 28 S and migrates on gels in aqueous solution with an apparent molecular weight of 2.0 x lo6. Contour length measurements under denaturing conditions in the electron microscope reveal a molecular weight of 2.34 x lo6 compared to the mouse 28-S rRNA.Labeling experiments show that the estrogen-induced RNA has a higher stability than the average liver poly(A)-containing RNA and represents 10 -20 of the poly(A)-containing RNA in the cytoplasm after 24 h of labeling.Hybridization of complementary DNA, synthesized on the isolated estrogen-induced RNA, with its template suggests a complexity corresponding to a single species of poly(A)-containing RNA of such a high molecular weight. Hybridization of the complementary DNA with cytoplasmic poly(A)-containing RNA from estrogen-treated Xenopus males and control toads show that the estrogen-induced RNA constitutes 12 -15 % of all cytoplasmic poly(A)-containing RNA, and is at least 2000-fold less abundant in untreated males.Size, complexity and abundance of the estrogen-induced RNA are characteristics expected for a mRNA coding for vitellogenin.

show abstract

Section: Discussionmentioning

confidence: 94%

Size, Complexity and Abundance of a Specific Poly(A)‐Containing RNA of Liver from Male Xenopus Induced to Vitellogenin Synthesis by Estrogen

Wahli

Wyler

Weber

et al. 1976

European Journal of Biochemistry

View full text Add to dashboard Cite

show abstract

“…Myosin mRNA has been assigned a molecular weight of 2,000,000 based on both aqueous and formamide gel electrophoresis, but sediments at 26 5 in sucrose gradients (24,25); sedimentation at 26 S implies a molecular weight of about 1,400,000, which is obviously too small to code for myosin. It seems likely that the anomolously low sedimentation constant is due to myosin 'mRNA having a more extended mean configuration than that of ribosomal RNAs.…”

Section: Methodsmentioning

confidence: 99%

The Identification of Collagen Messenger RNA

Boedtker

Crkvenjakov

Doty

1974

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

RNA isolated from calvaria of 16-to 18-day-old chick embryos, assayed in rabbit reticulocyte lysates, programs the synthesis of a collagenase-sensitive protein with the molecular weight of collagen pro-achains. When RNA labeled with [3Hjuridine for 2 hr and chased for 1 or 2 hr was electrophoresed on aqueous polyacrylamide gels, most of the radioactivity not in 28S or 1S rRNA migrated with an apparent-molecular weight of about 1,800,000. After oligo(dT)-cellulose chromotography and analysis in 99% forinamide gels, this nonrihosomal, rapidly labeled calvaria RNA species migrates at 28S-30S and thus has a molecular weight of at least 1,600,000.Both the ability to-program the synthesis of collagenasesensitive protein in reticulocyte lysates and the presence of a single prominent rapidly labeled 30S peak in acrylamide gels strongly support the deduction that there is only one major mRNA species in calvaria and that this species is collagen messenger RNA.Understanding the control of collagen biosynthesis is essential to understanding normal growth and development in vertebrates. Not only is collagen their major structural protein, but the onset of collagen synthesis is coincident with the onset of major differentiational' events. An important step in the study of collagen biosynthesis must be the purification and characterization of the messenger RNAs which code for the five or more different collagen pro-a-chains.Collagen messenger RNA (mRNA) has already been detected in polysomes prepared from the wings and legs of 8-'or 9-day-old chick embryos by assaying for collagen synthesis in homologous cell-free extracts (1, 2) and in heterologous cellfree extracts prepared from Krebs II ascites cells (3). Collagen synthesizing polysomes have also been isolated from L-929 fibroblasts (4). However, collagen mRNA has not been succ'essfully isolated from any of these polysomes.More recently, Benveniste, Wilczek, and Stern (5) reported that total RNA'isolated from the calvaria of 15-day-old chick embryos contained active collagen mRNA. They found that more than half of the [3H]proline-labeled protein made in ascites tumor extracts when calvaria RNA was added could be solubilized by collagenase. Also, some of this protein cochromatographed with carrier collagen on carboxymethyl cellulose. We have thus isolated RNA from calvaria. From a combination of pulse-chase studies of this RNA, under both aqueous and denaturing conditions, and assays of its ability to code for the synthesis of intact procollagen in rabbit reticulocyte lysates, we have been able to show that calvaria contain predominantly one class of mRNA, having a molecular weight of about 1,600,000-1,800,000. Several arguments indicate that this is collagen mRNA. MATERIALS AND METHODSPreparation and Fractionation of RNA from Calvaria. Calvaria RNA was prepared from 16-to 18-day-old chick embryos according to the procedure described by Benveniste et al. (5). The RNA was further purified by centrifugation through 5.7 M CsCl as described by Glisin et al. (6).To prepar...

show abstract

“…TOTAL POLYSOME ABSORBANCE AT 260nni/10^ NUCLEI to fusion of mononucleates myoblasts into multinucleated myotubes (Coleman and Coleman, 1968;Morris et al, 1972;Paterson and Strohman, 1972;Stockdale and O'Neill, 1972;Yaffe and Dym, 1972). Polysomes were isolated from 24-hr and 72-hr myogenic cell cultures as described in Ma terials and Methods and were displayed on a linear 15-^0^ sucrose gradient in isolation buffer after centrifugation at 105,000 for 2.0 hrs.…”

Section: Results MD Discussionmentioning

confidence: 99%

“…The myosin mRKA was then eluted in a single peak of greater than 95^ purity by washing the column with 0.001 M EDTA and 0.001 M Tris, pH 7.0. Purified RNA obtained by the two procedures described above was shown to be myosin mRNA because it directed cell-free synthesis of myosin heavy chains (Morris et al, 1972;Sarkar et al, 197^;Mondai et al, 1974) and because the peptides produced by cyanogen bromide cleavage of iji vitro labeled myosin matched those produced by cyanogen bromide cleavage of in vivo labeled myosin .…”

Section: ^2mentioning

confidence: 99%

“…Fractions collected from these columns were assayed for their ability to promote specific binding of ^^P-labeled myosin mENA to ribosomes in the nitrocellulose filter assay (Heywood, 1969). These chromatographic experiments showed that the factors responsible for mRNA were excluded from these data is that highly differentiated cells such as reticulocytes and muscle contain multiple, messenger-specific IF3's (Rourke and Heywood, 1972).…”

Section: ^2mentioning

confidence: 99%

See 1 more Smart Citation

Effect of myoblast fusion during muscle cell differentiation on myosin polysome accumulation and myosin synthesis

Young

View full text Add to dashboard Cite

Myosin Messenger RNA: Studies on Its Purification, Properties and Translation during Myogenesis in Culture

Cited by 52 publications

References 0 publications

Size, Complexity and Abundance of a Specific Poly(A)‐Containing RNA of Liver from Male Xenopus Induced to Vitellogenin Synthesis by Estrogen

Size, Complexity and Abundance of a Specific Poly(A)‐Containing RNA of Liver from Male Xenopus Induced to Vitellogenin Synthesis by Estrogen

The Identification of Collagen Messenger RNA

Effect of myoblast fusion during muscle cell differentiation on myosin polysome accumulation and myosin synthesis

Contact Info

Product

Resources

About