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In JoVE (1)
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Articles by Yana Izenshtein in JoVE
Pulso-chase Análise de Cadeias N-linked Açúcar de Glicoproteínas em células de mamíferos
Edward Avezov, Efrat Ron, Yana Izenshtein, Yosef Adan, Gerardo Z. Lederkremer
Department of Cell Research and Immunology, George Wise Faculty of Life Sciences, Tel Aviv University
Nós descrevemos um método para análise da alteração da N-glicanos ligados através do início da vida de glicoproteínas após a sua biossíntese em células de mamíferos. Isto é conseguido através de pulso-chase análise de glicanos metabolicamente marcados, a liberação enzimática das glicoproteínas e análise por HPLC.
Other articles by Yana Izenshtein on PubMed
Molecular Biology of the Cell. Nov, 2011 | Pubmed ID: 21917589
Trimming of mannose residues from the N-linked oligosaccharide precursor is a stringent requirement for glycoprotein endoplasmic reticulum (ER)-associated degradation (ERAD). In this paper, we show that, surprisingly, overexpression of ER degradation-enhancing α-mannosidase-like protein 1 (EDEM1) or its up-regulation by IRE1, as occurs in the unfolded protein response, overrides this requirement and renders unnecessary the expression of ER mannosidase I. An EDEM1 deletion mutant lacking most of the carbohydrate-recognition domain also accelerated ERAD, delivering the substrate to XTP3-B and OS9. EDEM1 overexpression also accelerated the degradation of a mutant nonglycosylated substrate. Upon proteasomal inhibition, EDEM1 concentrated together with the ERAD substrate in the pericentriolar ER-derived quality control compartment (ERQC), where ER mannosidase I and ERAD machinery components are localized, including, as we show here, OS9. We suggest that a nascent glycoprotein can normally dissociate from EDEM1 and be rescued from ERAD by reentering calnexin-refolding cycles, a condition terminated by mannose trimming. At high EDEM1 levels, glycoprotein release is prevented and glycan interactions are no longer required, canceling the otherwise mandatory ERAD timing by mannose trimming and accelerating the targeting to degradation.