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Molecular Pharmacology

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Research ArticleArticle

Norepinephrine-Dependent Protein Phosphorylation in Intact C-6 Glioma Cells

Analysis by Two-Dimensional Gel Electrophoresis

VINCENT E. GROPPI JR. and EDWARD T. BROWNING
Molecular Pharmacology November 1980, 18 (3) 427-437;
VINCENT E. GROPPI JR.
Department of Pharmacology, College of Medicine and Dentistry of New Jersey, Rutgers Medical School, Piscataway, New Jersey 08854
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EDWARD T. BROWNING
Department of Pharmacology, College of Medicine and Dentistry of New Jersey, Rutgers Medical School, Piscataway, New Jersey 08854
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Abstract

Norepinephrine rapidly induced phosphorylation of 7 of the more than 200 neutral and acidic phosphoproteins which were resolved by two-dimensional gel electrophoresis of a whole cell extract of C-6 glioma cells. Cellular ATP pools were nearly optimally labeled by 32Pi, after a 4-hr exposure, and norepinephrine treatment did not modify the specific radioactivity of the γ-phosphate of cellular ATP. That the 32P-labeled spots of two-dimensional gels were indeed phosphopolypeptides was verified by comparing the migration of 35S-methionine-labe1ed and 32Pi-labeled samples and by extensive solvent extractions and chemical treatments. Subcellular fractionation resulted in assignment of a subcellular compartment to five of the seven norepinephrine-dependent phosphopolypeptides. These modified proteins were designated the "phosphoprotein domain" of norepinephrine for the C-6 glioma cell and each was provisionally named in terms of molecular weight, isoelectric point, and, where possible, subcellular compartment. Accordingly the proteins were named: 58K-5.7-nuclear; 50K-6.1, 48K-6.8-cytosolic; 38K-6.4-cytosolic; 20K-6.2, 19K-6.6-mitochondrial; and 16.5K-6.3-cytosolic. We have concluded that protein 58K-5.7-nuclear is the intermediate filament protein of the C-6 glioma cell based on similarities of molecular weight, isoelectric point, abundance, subcellular fractionation, nuclear binding, and elution as well as phosphorylation.

  • Copyright © 1980 by The American Society for Pharmacology and Experimental Therapeutics

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Molecular Pharmacology
Vol. 18, Issue 3
1 Nov 1980
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Research ArticleArticle

Norepinephrine-Dependent Protein Phosphorylation in Intact C-6 Glioma Cells

VINCENT E. GROPPI and EDWARD T. BROWNING
Molecular Pharmacology November 1, 1980, 18 (3) 427-437;

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Research ArticleArticle

Norepinephrine-Dependent Protein Phosphorylation in Intact C-6 Glioma Cells

VINCENT E. GROPPI and EDWARD T. BROWNING
Molecular Pharmacology November 1, 1980, 18 (3) 427-437;
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