<i>PNG1</i>, a Yeast Gene Encoding a Highly Conserved Peptide:<i>N</i>-Glycanase

Suzuki, Tadashi; Park, Hangil; Hollingsworth, Nancy M.; Sternglanz, Rolf; Lennarz, William J.

doi:10.1083/jcb.149.5.1039

Cited by 212 publications

(229 citation statements)

References 63 publications

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“…Next, we examined the involvement of the proteasome system using MG-132, a specific inhibitor of the proteasome (Suzuki et al, 2000). Gas1*p was dramatically stabilized by the addition of MG-132 ( Figure 4E).…”

Section: Construction and Characterization Of Misfolded Gas1mentioning

confidence: 99%

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Inositol Deacylation by Bst1p Is Required for the Quality Control of Glycosylphosphatidylinositol-anchored Proteins

Fujita

Yoko-o

Jigami

2006

MBoC

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Misfolded proteins are recognized in the endoplasmic reticulum (ER), transported back to the cytosol, and degraded by the proteasome. A number of proteins are processed and modified by a glycosylphosphatidylinositol (GPI) anchor in the ER, but the quality control mechanisms of GPI-anchored proteins remain unclear. Here, we report on the quality control mechanism of misfolded GPI-anchored proteins. We have constructed a mutant form of the ␤-1,3-glucanosyltransferase Gas1p (Gas1*p) as a model misfolded GPI-anchored protein. Gas1*p was modified with a GPI anchor but retained in the ER and was degraded rapidly via the proteasome. Disruption of BST1, which encodes GPI inositol deacylase, caused a delay in the degradation of Gas1*p. This delay was because of an effect on the deacylation activity of Bst1p. Disruption of genes involved in GPI-anchored protein concentration and N-glycan processing caused different effects on the degradation of Gas1*p and a soluble misfolded version of carboxypeptidase Y. Furthermore, Gas1*p associated with both Bst1p and BiP/Kar2p, a molecular chaperone, in vivo. Our data suggest that GPI inositol deacylation plays important roles in the quality control and ER-associated degradation of GPI-anchored proteins. INTRODUCTIONCells possess several quality control mechanisms for the maintenance of proper protein folding and function. On synthesis, membrane and secretory proteins are inserted into the lumen of the endoplasmic reticulum (ER), where they are folded and undergo oligomerization. There are a number of chaperones and enzymes in the ER required for proper protein folding (Ellgaard et al., 1999). Before exiting from the ER, proteins are monitored by a quality control system that ensures correct folding. Misfolded proteins that fail to pass the quality control checkpoint are transported back to the cytosol and degraded by an ER-associated degradation (ERAD) mechanism that involves the ubiquitinproteasome pathway (Kopito, 1997;Kostova and Wolf, 2003). N-linked oligosaccharide, one of the major posttranslational modifications in the ER, is trimmed and processed by glucosidase I, glucosidase II, and mannosidase I (Jakob et al., 1998;Helenius and Aebi, 2001). The processing of Nlinked oligosaccharides plays important roles in the quality control of glycoprotein folding in the ER Aebi, 2001, 2004).A number of cell surface proteins are posttranslationally modified in the ER with glycosylphosphatidylinositol (GPI). Mechanisms for quality control and degradation of GPIanchored proteins are important in folding diseases, including prion diseases and transmissible spongiform encephalopathies, which are caused by a conformational modification of prion, a GPI-anchored protein (Prusiner, 1998). There have been several biochemical studies on both the degradation of mutated prions and the quality control of proteins with mutated GPI attachment signals (Field et al., 1994;Oda et al., 1996;Wainwright and Field, 1997;Jin et al., 2000;Ito et al., 2002;Ishida et al., 2003). In contrast, the molecular mech...

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Section: Construction and Characterization Of Misfolded Gas1mentioning

confidence: 99%

“…The preparation of samples and immunoblotting were performed as described above. The effect of the proteasome inhibitor was analyzed as described previously (Suzuki et al, 2000). Ten minutes before CHX was added, MG-132 (Merck, Darmstadt, Germany) was added to a final concentration of 50 M from a freshly made 5 mM solution in dimethyl sulfoxide (DMSO).…”

mentioning

confidence: 99%

Inositol Deacylation by Bst1p Is Required for the Quality Control of Glycosylphosphatidylinositol-anchored Proteins

Fujita

Yoko-o

Jigami

2006

MBoC

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“…Des études in vivo montrent que le taux de dégradation de la majorité des substrats ERAD étudiés n'est pas profondément altéré par un grand nombre de conditions qui induisent une inhibition de la PNGase, comme la délétion du gène correspondant chez la levure [6], ou l'utilisation d'ARNi (interférents) spécifiques [20], ou de l'inhibiteur Z-VAD-fmk (un inhibiteur des caspases) [19]. In vitro, la présence de la chaîne N-glycane n'empêche pas la dégradation de glycoprotéines dénaturées ou de substrats ERAD par les structures 20 S et 26 S du protéasome, mais la ralentit [21].…”

Section: La Pngase Modifie Peu Le Taux De Dégradation Des Substrats Eradunclassified

“…Il avait été observé que la délétion du gène PNG1 n'était pas associée à un phénotype particulier, en dehors de l'absence d'activité de déglycosylation, chez les organismes unicellulaires comme la levure S. cerevisiae [6]. Des articles apparus récemment dans la littérature décrivent cependant que la délétion du gène PNG1 dans des organismes pluricellulaires entraîne l'apparition de défauts morphogéniques.…”

Section: Png1 Joue Un Rôle Dans La Mise En Place De La Polarité Celluunclassified

Nouvelles fonctions de la peptideN-glycanase indépendantes de sa capacité de déglycosylation

2014

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> La première fonction qui a été attribuée à la peptide N-glycanase (PNGase) est de déglycosyler les N-glycosylprotéines mal repliées avant leur dégradation par le protéasome. Il s'avère cependant que l'inhibition de cette enzyme modifie peu le taux de dégradation de ces N-glycosylprotéines. Des données récentes montrent que cette enzyme a également la capacité de moduler la morphogenèse de façon indépendante de sa fonction de déglycosylation. La caractérisation du premier déficit en PNGase devrait apporter une meilleure compréhension du rôle de cette enzyme multifonctionnelle dans la physiologie humaine. < stockage lysosomal la plus fréquente, et elle se manifeste par une accumulation de petits glycopeptides dans les lysosomes. Les symptômes cliniques sont un retard psychomoteur progressif, ainsi que des anomalies du squelette et du visage. De très nombreuses mutations du gène AGA codant pour cette hydrolase lysosomale ont été caractéri-sées [2]. La première description d'un patient porteur d'un déficit en PNGase ne date, elle, que de 2012 [3]. Les symptômes de ce patient peuvent être la conséquence d'un dysfonctionnement, soit de l'activité d'hydrolyse de la chaîne N-glycane, soit de fonctions nouvellement décrites qui sont indépendantes de l'activité de déglycosylation de l'enzyme. Dans cette revue, seront donc successivement exposées les données conséquentes concernant le rôle de la PNGase dans la dégradation des N-glycosylprotéines mal repliées, puis les données plus restreintes concernant les autres fonctions potentielles de cette enzyme. Principales caractéristiques de la PNGaseUne activité PNGase s'exerçant sur une N-glycosylprotéine produit de façon concomitante un oligosaccharide libre (fOS pour free oligosaccharide) et la conversion du résidu asparagine porteur de la chaîne N-glycane en résidu aspartate (Asp) ( Figure 1A). Bien que des activités PNGase aient été trouvées depuis de nombreuses années chez les plantes et les bactéries, la découverte d'activités PNGase cytosoliques chez les mammifères [4], puis chez la levure Saccharomyces Inserm U773, centre de recherche Bichat Beaujon CRB3,

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“…After exit from the ER lumen, HCs encounter the deglycosylating peptide:N-glycanase (PNGase; Suzuki et al, 2000) and consequently are deglycosylated at the ER membrane or in the cytosol (Misaghi et al, 2004). Although deglycosylation is not an absolute requirement, it is thought to facilitate proteasomal degradation by providing increased accessibility to the proteasome pore by removal of the bulky glycan chains from misfolded glycoproteins (Hirsch et al, 2003).…”

Section: Introductionmentioning

confidence: 99%

The Retrotranslocation Protein Derlin-1 Binds Peptide:N-Glycanase to the Endoplasmic Reticulum

Katiyar

Joshi

Lennarz

2005

MBoC

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The deglycosylating enzyme, peptide:N-glycanase, acts on misfolded N-linked glycoproteins dislocated from the endoplasmic reticulum (ER) to the cytosol. Deglycosylation has been demonstrated to occur at the ER membrane and in the cytosol. However, the mechanism of PNGase association with the ER membrane was unclear, because PNGase lacked the necessary signal to facilitate its incorporation in the ER membrane, nor was it known to bind to an integral ER protein.Using HeLa cells, we have identified a membrane protein that associates with PNGase, thereby bringing it in close proximity to the ER and providing accessibility to dislocating glycoproteins. This protein, Derlin-1, has recently been shown to mediate retrotranslocation of misfolded glycoproteins. In this study we demonstrate that Derlin-1 interacts with the N-terminal domain of PNGase via its cytosolic C-terminus. Moreover, we find PNGase distributed in two populations; ER-associated and free in the cytosol, which suggests the deglycosylation process can proceed at either site depending on the glycoprotein substrate. INTRODUCTIONIn eukaryotes proteins are synthesized on membrane-bound ribosomes and undergo a folding process in the lumen of the endoplasmic reticulum (ER; Helenius, 2001, 2003). Misfolded or incompletely assembled multisubunit glycoproteins are recognized by the endoplasmic reticulumassociated degradation (ERAD) pathway regulated largely by their N-linked polymannose oligosaccharides. In this quality control system, lectin-like molecular chaperones, calnexin and calreticulin, recognize the Glc 1 Man 9 GlcNAc 2 oligosaccharide and assist in folding of newly synthesized glycoproteins. Lectin interaction with Glc 1 Man 9 GlcNAc 2 glycans after trimming with ER alpha-glucosidases, and alpha-mannosidases sorts out persistently unfolded glycoproteins for N-deglycosylation and degradation by the proteasome (Cresswell and Hughes, 1997;Kopito, 1997;Brodsky and McCracken, 1999;Romisch, 1999;Spiro, 2004).The mannose trimming of N-linked glycans plays an important role in the ERAD of glycoproteins (Spiro, 2004). In both yeast and human cells, it is reported that the misfolded glycoproteins in the ER are degraded through ERAD only after the glycan is trimmed to the Man8B form, while misfolded proteins stay within the ER when they retain the Man9 form of oligosaccharides. Accordingly, it was assumed that there existed a Man8-binding lectin, later identified as EDEM, that recognizes misfolded glycoproteins in the Man8B form in the ER and directs them to the ERAD pathway (Hosokawa et al., 2001). The misfolded glycoproteins exit the ER via a multiprotein complex referred to as a retrotranslocon or dislocon (Plemper et al., 1997;Bebok et al., 1998;de Virgilio et al., 1998;Tsai et al., 2002). In alternative models both PNGase and proteasomes may be either free in the cytosol or ER membrane imbedded or attached (Spiro, 2004).A well-studied model for the ERAD pathway utilizes the human cytomegalovirus, which affects the MHC class I antigen presentation. The vira...

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PNG1, a Yeast Gene Encoding a Highly Conserved Peptide:N-Glycanase

Cited by 212 publications

References 63 publications

Inositol Deacylation by Bst1p Is Required for the Quality Control of Glycosylphosphatidylinositol-anchored Proteins

Inositol Deacylation by Bst1p Is Required for the Quality Control of Glycosylphosphatidylinositol-anchored Proteins

Nouvelles fonctions de la peptideN-glycanase indépendantes de sa capacité de déglycosylation

The Retrotranslocation Protein Derlin-1 Binds Peptide:N-Glycanase to the Endoplasmic Reticulum

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