Nutrient modulation of palmitoylated 24-kilodalton protein in rat pancreatic islets.
Protein acylation in glucose stimulation of insulin secretion in the beta-cells has been implicated. Accordingly, we attempted to identify the target(s) of acylation in the pancreatic islets. Rat pancreatic islets were labeled with [3H]palmitic acid for 1 h at 37 C, and the whole cell lysate was analyzed by SDS-PAGE and two-dimensional gel electrophoresis. The labeling of the proteins by [3H]palmitic acid was shown to be palmitoylation by chemical analyses. Palmitoylation of four distinct bands was recognized, and the palmitoylation was significantly reduced in all of them when the labeling was performed with high glucose. Quite interestingly, the degree of attenuation was particularly dominant for a 24-kDa doublet. Palmitoylation of the 24-kDa doublet was preferentially attenuated also by the mitochondrial fuels and an acylation inhibitor, cerulenin. The half-life of the labeling of the doublet was apparently shorter (approximately 45 min) than that of other bands on pulse chasing of the islets, irrespective of the presence or absence of high glucose. High glucose attenuation of the palmitoylation of the 24-kDa doublet was partially blocked by 20 mm mannoheptulose, a glucokinase inhibitor. Two-dimensional gel electrophoresis revealed that the doublet was composed of acidic peptides, and, by immunoprecipitation, it was shown not to be synaptosome-associated protein of 25 kDa. We identified rapidly turning over palmitoylated 24-kDa acidic proteins distinct from synaptosome-associated protein of 25 kDa in the pancreatic islets, which are preferentially modulated by fuel secretagogues. The data suggested a functional role of the palmitoylated 24-kDa doublet in nutrient stimulation of insulin secretion.
Yamada
Satoko
S
Institute for Molecular and Cellular Regulation, Gunma University, Maebashi 371-8512, Japan. ikojima@showa.gunma-u.ac.jp.
Komatsu
Mitsuhisa
M
Sato
Yoshihiko
Y
Yamauchi
Keishi
K
Aizawa
Toru
T
Kojima
Itaru
I
D000215:Acylation / Q000187:drug effects; D000818:Animals; D000906:Antibodies; D000935:Antifungal Agents / Q000494:pharmacology; D002569:Cerulenin / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology; D004435:Eating / Q000502:physiology*; D015180:Electrophoresis, Gel, Two-Dimensional; D005941:Glucokinase / Q000037:antagonists & inhibitors / Q000378:metabolism; D005947:Glucose / Q000494:pharmacology; D066298:In Vitro Techniques; D007515:Islets of Langerhans / Q000378:metabolism*; D008297:Male; D008356:Mannoheptulose / Q000494:pharmacology; D008565:Membrane Proteins / Q000276:immunology; D008928:Mitochondria / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000276:immunology; D019308:Palmitic Acid / Q000493:pharmacokinetics*; D011233:Precipitin Tests; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D050825:Synaptosomal-Associated Protein 25; D014316:Tritium
D000906:Antibodies; D000935:Antifungal Agents; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D011500:Protein Synthesis Inhibitors; C494210:Snap25 protein, rat; D050825:Synaptosomal-Associated Protein 25; D014316:Tritium; D002569:Cerulenin; D019308:Palmitic Acid; D008356:Mannoheptulose; D003513:Cycloheximide; D005941:Glucokinase; D005947:Glucose