Publications (31)384.11 Total impact
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Article: SnapShot: ER-associated protein degradation pathways.
Cell 07/2007; 129(6):1230. · 32.40 Impact Factor -
Article: Have you HRD? Understanding ERAD is DOAble!
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ABSTRACT: Numerous factors are involved in the eradication of misfolded proteins, yet how these factors achieve substrate specificity remains unclear. In this issue of Cell, Denic et al. (2006) and Carvalho et al. (2006) report that two distinct protein complexes at the endoplasmic reticulum membrane are responsible for the recognition and degradation of specific subsets of protein substrates.Cell 08/2006; 126(2):237-9. · 32.40 Impact Factor -
Article: Yos9p detects and targets misfolded glycoproteins for ER-associated degradation.
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ABSTRACT: Endoplasmic reticulum (ER) quality control mechanisms monitor the folding of nascent secretory and membrane polypeptides. Immature molecules are actively retained in the folding compartment whereas proteins that fail to fold are diverted to proteasome-dependent degradation pathways. We report that a key pathway of ER quality control consists of a two-lectin receptor system consisting of Yos9p and Htm1/Mnl1p that recognizes N-linked glycan signals embedded in substrates. This pathway recognizes lumenally oriented determinants of soluble and membrane proteins. Yos9p binds directly to substrates to discriminate misfolded from folded proteins. Substrates displaying cytosolic determinants can be degraded independently of this system. Our studies show that mechanistically divergent systems collaborate to guard against passage and accumulation of misfolded proteins in the secretory pathway.Molecular Cell 10/2005; 19(6):753-64. · 14.18 Impact Factor -
Article: Screening for mutants defective in secretory protein maturation and ER quality control.
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ABSTRACT: A genetic strategy devised to understand the physiology of the unfolded protein response serendipitously generated mutants affecting a broad spectrum of functions needed for secretory protein biogenesis and quality control. These included N- and O-linked glycosylation, glycosylphosphatidylinositol anchor biosynthesis and transfer, protein folding, protein trafficking, lumenal ionic homeostasis, ER quality control, and ER associated protein degradation. As these pathways are incompletely understood, the screen provides a simple method for their genetic dissection. This article describes methods for isolating novel mutants of these pathways and strategies for identifying corresponding genes.Methods 05/2005; 35(4):366-72. · 4.01 Impact Factor -
Article: Single, context-specific glycans can target misfolded glycoproteins for ER-associated degradation.
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ABSTRACT: The endoplasmic reticulum (ER) maintains an environment essential for secretory protein folding. Consequently, the premature transport of polypeptides would be harmful to the cell. To avert this scenario, mechanisms collectively termed "ER quality control" prevent the transport of nascent polypeptides until they properly fold. Irreversibly misfolded molecules are sorted for disposal by the ER-associated degradation (ERAD) pathway. To better understand the relationship between quality control and ERAD, we studied a new misfolded variant of carboxypeptidase Y (CPY). The molecule was recognized and retained by ER quality control but failed to enter the ERAD pathway. Systematic analysis revealed that a single, specific N-linked glycan of CPY was required for sorting into the pathway. The determinant is dependent on the putative lectin-like receptor Htm1/Mnl1p. The discovery of a similar signal in misfolded proteinase A supported the generality of the mechanism. These studies show that specific signals embedded in glycoproteins can direct their degradation if they fail to fold.The Journal of Cell Biology 05/2005; 169(1):73-82. · 10.26 Impact Factor -
Article: Misfolded proteins are sorted by a sequential checkpoint mechanism of ER quality control.
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ABSTRACT: Misfolded proteins retained in the endoplasmic reticulum (ER) are degraded by the ER-associated degradation pathway. The mechanisms used to sort them from correctly folded proteins remain unclear. Analysis of substrates with defined folded and misfolded domains has revealed a system of sequential checkpoints that recognize topologically distinct domains of polypeptides. The first checkpoint examines the cytoplasmic domains of membrane proteins. If a lesion is detected, it is retained statically in the ER and rapidly degraded without regard to the state of its other domains. Proteins passing this test face a second checkpoint that monitors domains localized in the ER lumen. Proteins detected by this pathway are sorted from folded proteins and degraded by a quality control mechanism that requires ER-to-Golgi transport. Although the first checkpoint is obligatorily directed at membrane proteins, the second monitors both soluble and membrane proteins. Our data support a model whereby "properly folded" proteins are defined biologically as survivors that endure a series of distinct checkpoints.The Journal of Cell Biology 05/2004; 165(1):41-52. · 10.26 Impact Factor -
Article: Stress tolerance of misfolded carboxypeptidase Y requires maintenance of protein trafficking and degradative pathways.
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ABSTRACT: The accumulation of aberrantly folded proteins can lead to cell dysfunction and death. Currently, the mechanisms of toxicity and cellular defenses against their effects remain incompletely understood. In the endoplasmic reticulum (ER), stress caused by misfolded proteins activates the unfolded protein response (UPR). The UPR is an ER-to-nucleus signal transduction pathway that regulates a wide variety of target genes to maintain cellular homeostasis. We studied the effects of ER stress in budding yeast through expression of the well-characterized misfolded protein, CPY*. By challenging cells within their physiological limits to resist stress, we show that the UPR is required to maintain essential functions including protein translocation, glycosylation, degradation, and transport. Under stress, the ER-associated degradation (ERAD) pathway for misfolded proteins is saturable. To maintain homeostasis, an "overflow" pathway dependent on the UPR transports excess substrate to the vacuole for turnover. The importance of this pathway was revealed through mutant strains compromised in the vesicular trafficking of excess CPY*. Expression of CPY* at levels tolerated by wild-type cells was toxic to these strains despite retaining the ability to activate the UPR.Molecular Biology of the Cell 08/2003; 14(7):2756-67. · 4.94 Impact Factor -
Article: Two distinctly localized p-type ATPases collaborate to maintain organelle homeostasis required for glycoprotein processing and quality control.
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ABSTRACT: Membrane transporter proteins are essential for the maintenance of cellular ion homeostasis. In the secretory pathway, the P-type ATPase family of transporters is found in every compartment and the plasma membrane. Here, we report the identification of COD1/SPF1 (control of HMG-CoA reductase degradation/SPF1) through genetic strategies intended to uncover genes involved in protein maturation and endoplasmic reticulum (ER)-associated degradation (ERAD), a quality control pathway that rids misfolded proteins. Cod1p is a putative ER P-type ATPase whose expression is regulated by the unfolded protein response, a stress-inducible pathway used to monitor and maintain ER homeostasis. COD1 mutants activate the unfolded protein response and are defective in a variety of functions apart from ERAD, which further support a homeostatic role. COD1 mutants display phenotypes similar to strains lacking Pmr1p, a Ca(2+)/Mn(2+) pump that resides in the medial-Golgi. Because of its localization, the previously reported role of PMR1 in ERAD was somewhat enigmatic. A clue to their respective roles came from observations that the two genes are not generally required for ERAD. We show that the specificity is rooted in a requirement for both genes in protein-linked oligosaccharide trimming, a requisite ER modification in the degradation of some misfolded glycoproteins. Furthermore, Cod1p, like Pmr1p, is also needed for the outer chain modification of carbohydrates in the Golgi apparatus despite its ER localization. In strains deleted of both genes, these activities are nearly abolished. The presence of either protein alone, however, can support partial function for both compartments. Taken together, our results reveal an interdependent relationship between two P-type ATPases to maintain homeostasis of the organelles where they reside.Molecular Biology of the Cell 12/2002; 13(11):3955-66. · 4.94 Impact Factor -
Article: Translocation of lipid-linked oligosaccharides across the ER membrane requires Rft1 protein.
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ABSTRACT: N-linked glycosylation of proteins in eukaryotic cells follows a highly conserved pathway. The tetradecasaccharide substrate (Glc3Man9GlcNAc2) is first assembled at the membrane of the endoplasmic reticulum (ER) as a dolichylpyrophosphate (Dol-PP)-linked intermediate, and then transferred to nascent polypeptide chains in the lumen of the ER. The assembly of the oligosaccharide starts on the cytoplasmic side of the ER membrane with the synthesis of a Man5GlcNAc2-PP-Dol intermediate. This lipid-linked intermediate is then translocated across the membrane so that the oligosaccharides face the lumen of the ER, where the biosynthesis of Glc3Man9GlcNAc2-PP-Dol continues to completion. The fully assembled oligosaccharide is transferred to selected asparagine residues of target proteins. The transmembrane movement of lipid-linked Man5GlcNAc2 oligosaccharide is of fundamental importance in this biosynthetic pathway, and similar processes involving phospholipids and glycolipids are essential in all types of cells. The process is predicted to be catalysed by proteins, termed flippases, which to date have remained elusive. Here we provide evidence that yeast RFT1 encodes an evolutionarily conserved protein required for the translocation of Man5GlcNAc2-PP-Dol from the cytoplasmic to the lumenal leaflet of the ER membrane.Nature 02/2002; 415(6870):447-50. · 36.28 Impact Factor -
Article: The EDEM and Yos9p families of lectin-like ERAD factors
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ABSTRACT: Protein quality control pathways monitor the folding of newly synthesized proteins throughout the cell. Irreversibly misfolded proteins are sorted and degraded to neutralize their potential toxicity. In the secretory pathway, multiple strategies have evolved to test the wide diversity of molecules that traffic through the endoplasmic reticulum. The organelle has adapted the use of N-linked glycans to signal protein folding states. The signals are read by the EDEM and Yos9 protein families that take substrates out of folding cycles for degradation.Seminars in Cell & Developmental Biology. -
Article: Search and destroy: ER quality control and ER-associated protein degradation.
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ABSTRACT: Proteins synthesized in the endoplasmic reticulum (ER) encounter quality control checkpoints that verify their fitness to proceed in the secretory pathway. Molecules undergoing folding and assembly are kept out of the exocytic pathway until maturation is complete. Misfolded side products that inevitably form are removed from the mixture of conformers and returned to the cytosol for degradation. How unfolded proteins are recognized and how irreversibly misfolded proteins are sorted to ER-associated degradation pathways was poorly understood. Recent developments from a combination of genetic and biochemical analyses has revealed new insights into these mechanisms. The emerging view shows distinct pathways working in collaboration to filter the diverse range of unfolded proteins from the transport flow and to divert misfolded molecules for destruction.Critical Reviews in Biochemistry and Molecular Biology 40(2):75-91. · 7.66 Impact Factor -
Article: How cells decide that a protein is misfolded is a mystery. The endoplasmic reticulum integrates n -linked glycosylation into the decision as to whether a protein is misfolded. The basic strategy of glycan-based recognition, previously identified in yeast, is conserved in mammals but is expanded, possibly to accommodate a more complex client portfolio
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ABSTRACT: The endoplasmic reticulum (ER) is a cel- lular factory that produces a wide range of proteins and lipids. These products are then transported to where they are needed, in the cell and beyond. Making accurate proteins is a challenge because folding and assembly take place at various sites within the lumen or membrane of the ER and in the cytosol. Furthermore, the specific and limited ER localization of many chaperones and modi- fying enzymes means that it is essential that molecules are not delivered to later compart- ments prematurely. For this reason, ER quality control (ERQC) mechanisms have evolved in all eukaryotes to monitor the folding and oli- gomerization states of nascent polypeptides. These systems retain proteins that are still in the process of folding in a particular compart- ment until folding is complete, and they target misfolded proteins for destruction by ER- associated degradation (ERAD) pathways. ERAD mechanisms vary between organisms. The common link among all ERAD pathways is the final step: substrate degradation by the 26S proteasome. Mechanistically, all ERAD systems must have factors that recognize substrates, translocate them across ER membranes and target them to the proteasome, usually through polyubiquitination. The systems operating in yeast are the most basic, with two defined path- ways organized around the two E3 ubiquitin ligases Hrd1p and Doa10p (ref. 1). The impor- tance of these pathways is underscored by their conservation in all eukaryotes. Metazoans have evolved diverse ERAD pathways, with the com- plete list still to be revealed 2 . Although the rea- sons for ERAD complexity continue to emerge, early indications are that different complexes are used to monitor different classes of substrate. The simplest version is found in yeast where the Hrd1p complex detects substrates with luminal or transmembrane lesions and the Doa10p com- plex monitors the folding of cytosolic domains 3 . This arrangement might seem sufficient to cover all types of substrate. Therefore, understanding why mammals expanded this basic framework several-fold should provide a new insight into the evolution of complex biosynthetic pathways. Characterization of distinct Hrd1 complexes in mammals highlights one mechanism that has evolved to use different components to detect misfolded proteins. N-linked glycans participate in the folding pathways of many glycoproteins 4
Top co-authors
Top Journals
- Molecular biology of the cell (4)
- The Journal of Cell Biology (2)
- Nature (2)
- Cell (2)
- Molecular Biology of the Cell (2)
Institutions
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2010–2012
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Temasek Life Sciences Laboratory
Singapore, Singapore
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2006–2012
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National University of Singapore
- Department of Biological Sciences
Singapore, Singapore
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2002–2008
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Pennsylvania State University
- Department of Biochemistry and Molecular Biology
University Park, MD, USA
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