The 3T3-L1 adipocyte glycogen proteome

Abstract

BackgroundGlycogen is a branched polysaccharide of glucose residues, consisting of α-1-4 glycosidic linkages with α-1-6 branches that together form multi-layered particles ranging in size from 30 nm to 300 nm. Glycogen spatial conformation and intracellular organization are highly regulated processes. Glycogen particles interact with their metabolizing enzymes and are associated with a variety of proteins that intervene in its biology, controlling its structure, particle size and sub-cellular distribution. The function of glycogen in adipose tissue is not well understood but appears to have a pivotal role as a regulatory mechanism informing the cells on substrate availability for triacylglycerol synthesis. To provide new molecular insights into the role of adipocyte glycogen we analyzed the glycogen-associated proteome from differentiated 3T3-L1-adipocytes.ResultsGlycogen particles from 3T3-L1-adipocytes were purified using a series of centrifugation steps followed by specific elution of glycogen bound proteins using α-1,4 glucose oligosaccharides, or maltodextrins, and tandem mass spectrometry. We identified regulatory proteins, 14-3-3 proteins, RACK1 and protein phosphatase 1 glycogen targeting subunit 3D. Evidence was also obtained for a regulated subcellular distribution of the glycogen particle: metabolic and mitochondrial proteins were abundant. Unlike the recently analyzed hepatic glycogen proteome, no endoplasmic proteins were detected, along with the recently described starch-binding domain protein 1. Other regulatory proteins which have previously been described as glycogen-associated proteins were not detected, including laforin, the AMPK beta-subunit and protein targeting to glycogen (PTG).ConclusionsThese data provide new molecular insights into the regulation of glycogen-bound proteins that are associated with the maintenance, organization and localization of the adipocyte glycogen particle.

Role of glycogen in development and adult fitness in Drosophila

“…This idea is supported by the fact that glycogen inhibits the kinase activity of AMPK by binding to the glycogen-binding domain of the β-subunit (McBride et al, 2009). Moreover, glycogen granules act as a signaling scaffold via physical interaction with a variety of proteins (Graham, 2009;Philp et al, 2012;Stapleton et al, 2013). Thus, the flux of either glycolysis or fat oxidation may be promoted through a metabolic shift to compensate for the energy deficit in a mutant-specific manner.…”

Algorithmic approaches to protein-protein interaction site prediction

“…Thus, unsurprisingly, training on one interface type to predict on the other tends to decrease scores [ 18 , 33 ], though this is sometimes not the case [ 13 ]. Generally, analysis of transient versus permanent complexes uses predefined sets [ 13 , 60 , 66 ] or programs designed to separate them [ 52 , 67 - 69 ].…”

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