Two-dimensional electrophoretic analysis of secretory-granule, granule-membrane, and plasma-membrane proteins of rat parotid cells

1983 ◽  
Vol 234 (1) ◽  
Author(s):  
MargaretA. Cascieri ◽  
EthelW. Somberg
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Secretory Granule ◽  
Electrophoretic Analysis ◽  
Two Dimensional ◽  
Plasma Membrane Proteins ◽  
Parotid Cells ◽  
Granule Membrane ◽  
Rat Parotid

scholarly journals Evidence for G proteins in rat parotid plasma membranes and secretory granule membranes

1992 ◽  
Vol 285 (2) ◽  
pp. 441-449 ◽  
Author(s):  
E L Watson ◽  
D DiJulio ◽  
D Kauffman ◽  
J Iversen ◽  
M R Robinovitch ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
G Proteins ◽  
Secretory Granule ◽  
Membrane Fraction ◽  
Immunoblot Analysis ◽  
Plasma Membrane Fraction ◽  
Adp Ribosylation ◽  
Granule Membrane ◽  
Rat Parotid ◽  
Gs Alpha

G proteins were identified in rat parotid plasma membrane-enriched fractions and in two populations of isolated secretory granule membrane fractions. Both [32P]ADP-ribosylation analysis with bacterial toxins and immunoblot analysis with crude and affinity-purified antisera specific for alpha subunits of G proteins were utilized. Pertussis toxin catalysed the ADP-ribosylation of a 41 kDa substrate in the plasma membrane fraction and both secretory granule membrane fractions. Cholera toxin catalysed the ADP-ribosylation of two substrates with molecular masses of 44 kDa and 48 kDa in the plasma membrane fraction but not in the secretory granule fractions. However, these substrates were detected in the secretory granule fractions when recombinant ADP-ribosylating factor was present in the assay medium. Immunoblot analysis of rat parotid membrane fractions using both affinity-purified and crude antisera revealed strong immunoreactivity of these membranes with anti-Gs alpha, -Gi alpha 1/alpha 2 and -Gi alpha 3 sera. In contrast Gs alpha was the major substrate found in both of the secretory granule fractions. Granule membrane fractions also reacted moderately with anti-Gi alpha 3 antiserum, and weakly with anti-Gi alpha 1/alpha 2 and -G(o) alpha sera. The results demonstrate that the parotid gland membranes express a number of G proteins. The presence of G proteins in secretory granule membranes suggests that they may play a direct role in regulating exocytosis in exocrine glands.

Regulated phosphorylation of secretory granule membrane proteins of the rat parotid gland

1990 ◽  
Vol 259 (1) ◽  
pp. G70-G77 ◽  
Author(s):  
C. R. Marino ◽  
J. D. Castle ◽  
F. S. Gorelick
Keyword(s):  
Membrane Proteins ◽  
Parotid Gland ◽  
Secretory Granule ◽  
Integral Membrane Proteins ◽  
Dependent Manner ◽  
Amylase Release ◽  
Rat Parotid Gland ◽  
Granule Membrane ◽  
Rat Parotid ◽  
Granule Membrane Proteins

An antiserum raised against purified rat parotid secretory granule membrane proteins has been used to identify organelle-specific protein phosphorylation events following stimulation of intact cells from the rat parotid gland. After lobules were prelabeled with [32P]orthophosphate and exposed to secretagogues, phosphoproteins were immunoprecipitated with the granule membrane protein antiserum, separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and visualized by autoradiography. Parallel studies of stimulated amylase release were performed. Isoproterenol treatment of parotid lobules resulted in an increase in the phosphate content of immunoprecipitable 60- and 72-kDa proteins that correlated with amylase release in a time-dependent manner. Forskolin addition mimicked these effects, but only the isoproterenol effects were reversed by propranolol treatment. To confirm the specificity of the antiserum to the secretory granule membrane fraction, subcellular isolation techniques were employed following in situ phosphorylation. The 60- and 72-kDa phosphoproteins were immunoprecipitated from both a particulate fraction and a purified secretory granule fraction. Furthermore, the extraction properties of both species suggest that they are integral membrane proteins. These findings support the possibility that stimulus-regulated secretion may involve phosphorylation of integral membrane proteins of the exocrine secretory granule.

Construction of a directory of tobacco plasma membrane proteins by combined two-dimensional gel electrophoresis and protein sequencing

1997 ◽  
Vol 18 (3-4) ◽  
pp. 654-660 ◽  
Author(s):  
David Rouquié ◽  
Jean-Beno??t Peltier ◽  
Monique Marquis-Mansion ◽  
Colette Tournaire ◽  
Patrick Doumas ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Gel Electrophoresis ◽  
Protein Sequencing ◽  
Two Dimensional ◽  
Plasma Membrane Proteins ◽  
Two Dimensional Gel Electrophoresis
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