Previous studies have shown that the organism's nutritional status changes the protein levels of insulin receptor substrate 1 (IRS-1) in a tissue-specific manner. Although the mechanisms underlying the regulation of IRS-1 in the nutrient-rich conditions associated with diabetes and insulin resistance have been well studied, those under nutrient-poor conditions remain unknown. The aim of this study was to investigate how IRS-1 protein levels change depending on the nutritional status of 3T3-L1 preadipocytes. 3T3-L1 preadipocytes were treated with glucose-, amino acid- and serum-free medium for starvation. IRS-1 protein levels were detected by western blot. Autophagy activity was observed by western blot and fluorescence microscopy. The effect of autophagy and p62, an adaptor for selective autophagy, on IRS-1 protein levels under starvation conditions was examined by western blot and immunocytochemistry. We showed that that the levels of IRS-1, but not those of insulin receptor and Akt, decreased when starvation activated autophagy. The inhibition of autophagy by chloroquine or autophagy-related 7 (Atg7) RNA interference counteracted the starvation-induced decrease of IRS-1. Additionally, Atg7 knockdown increased insulin-stimulated phosphorylation of Akt under starvation conditions. Furthermore, p62 co-localized with IRS-1 under starvation conditions, and p62 knockdown counteracted the starvation-induced degradation of IRS-1. Autophagy through p62 plays an important role in regulating IRS-1 protein levels in response to nutritional deficiency. Our findings suggest that autophagy may function as energy depletion-sensing machinery that finely tunes insulin signal transduction. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
WORKING FREE SYNAPSE X CRACKED LEVEL 7
The characteristics of a GTPase reaction between poly(U)-programmed ribosomes, EFTu . GTP, and the near-cognate aminoacyl (aa)-tRNA, Leu-tRNA Leu 2, have been studied to assess the role of this reaction in proofreading of the codon-anticodon interaction. The reaction resembles the GTPase reaction with cognate aa-tRNAs and EFTu . GTP in its substrate requirements, in its involving EFTu . GTP . aa-tRNA ternary complexes, and in its requiring a free ribosomal A-site. The noncognate reaction differs from the cognate one in that aa-tRNA becomes stably bound to the ribosomes only 5% of the time; it therefore seems best characterized as an abortive enzymatic binding reaction. The rate of reaction is a significant fraction (4%) of that of the cognate aa-tRNA, indicating that recognition of ternary complexes by ribosomes involves a level of error greater than that of translation as a whole. The rejection of the noncognate aa-tRNA following GTP hydrolysis is therefore a vital step in the translation process and fulfills the criteria set for a proofreading reaction. Leu-tRNA Leu 2 which escapes rejection through proofreading, forms a stable complex with the ribosomal A-site, so it appears that the Leu-tRNA2 which was rejected never reached the A-site and that proofreading precedes full A-site binding. 2ff7e9595c
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