ERAD
Encyclopedia
Endoplasmic-reticulum-associated protein degradation (ERAD) designates a cellular
pathway which targets misfolded proteins of the endoplasmic reticulum
for ubiquitination and subsequent degradation by a protein-degrading complex, called the proteasome
.
residues and immature glycans
.
In mammalian cells for example, there exists a mechanism called glycan processing. In this mechanism, the lectin
-type chaperones calnexin
/calreticulin
(CNX/CRT) provide immature glycoproteins the opportunity to reach their native conformation. They can do this by way of reglucosylating these glycoproteins by an enzyme called UDP
-glucose
-glycoprotein
glucosyltransferase. Terminally misfolded proteins, however, must be extracted from CNX/CRT. This is carried out by EDEM and ER mannosidase I. This mannosidase removes one mannose
residue from the glycoprotein and the latter is recognized by EDEM. Eventually EDEM will target the misfolded glycoproteins for degradation.
, constitutes the channel necessary for the transport of these misfolded proteins. Further, this translocation requires a driving force that determines the direction of transport. Since polyubiquitination is essential for the export of substrates, it is likely that this driving force is provided by ubiquitin-binding factors. One of these ubiquitin-binding factors is the Cdc48p-Npl4p-Ufd1p complex. It is known that human have the homolog of Cdc48p which is called Valosine-containing protein (VCP)/p97 and VCP/p97 have the same function of its yeast homolog Cdc48p. VCP/p97 transports substrates from endoplasmic reticulum to cytoplasm with its ATPase activity.
E1 hydrolyses ATP
and forms a high-energy thioester
linkage between a cysteine residue in its active site and the C-terminus of ubiquitin. The resulting activated ubiquitin is then passed to E2, which is a ubiquitin-conjugating enzyme
. Another group of enzymes, more specifically ubiquitin protein ligases called E3, bind to the misfolded protein. Next they align the protein and E2, thus facilitating the attachment of ubiquitin to lysine residues of the misfolded protein. Following successive addition of ubiquitin molecules to lysine residues of the previously attached ubiquitin, a polyubiquitin chain is formed. A polyubiquitinated protein is produced and this is recognized by specific subunits
in the 19S capping complexes of the 26S proteasome. Hereafter, the polypeptide chain is fed into the central chamber of the 20S core region that contains the proteolytically active sites. Ubiquitin is cleaved before terminal digestion by deubiquitinating enzymes. This third step is very closely associated with the second one, since ubiquitination takes place during the translocation event. However, the proteasomal degradation takes place in the cytoplasm.
Ubc6 as well as Ubc1 and the Cue1 dependent membrane bound Ubc7 are the ubiquitin conjugating enzymes involved in ERAD.
and transmembrane proteins were recognized by different mechanisms. This led to the identification of 3 different pathways that constitute in fact 3 checkpoints.
The first group is the result of mutations in ERAD components, which subsequently lose their function. By losing their function, these components are not longer able to stabilize aberrant proteins so that the latter accumulate and damage the cell. A very important example of a disease caused by this first group of disorders is Parkinson's disease
. It is caused by a mutation in the parkin gene
. Parkin
is a protein that functions in complex with CHIP as a ubiquitin ligase and overcomes the accumulation and aggregation of misfolded proteins.
[There are numerous debatable theories addressing the causes of Parkinson's Disease, besides the one presented here. Many of these can be found in the section of Wikipedia devoted to Parkinson's Disease]
In contrast to this first group of disorders, the second group is caused by premature degradation of secretory or membrane proteins. In this way, these proteins aren’t able to be deployed to distal compartments, as is the case in cystic fibrosis
.
uses an efficient mechanism to dislocate a single-membrane-spanning host protein, CD4
, from the ER and submits it to ERAD. CD4 is normally a stable protein and is not likely to be a target for ERAD. However, HIV produces the membrane protein Vpu
that binds to CD4. As Vpu is phosphorylated, it mimics substrates for the E3 complex SCFβTrCP. In cells that are infected with HIV, SCFβTrCP interacts with Vpu and ubiquitinates CD4, which is subsequently degraded by the proteasome. Vpu itself escapes from the degradation.
Cell (biology)
The cell is the basic structural and functional unit of all known living organisms. It is the smallest unit of life that is classified as a living thing, and is often called the building block of life. The Alberts text discusses how the "cellular building blocks" move to shape developing embryos....
pathway which targets misfolded proteins of the endoplasmic reticulum
Endoplasmic reticulum
The endoplasmic reticulum is an organelle of cells in eukaryotic organisms that forms an interconnected network of tubules, vesicles, and cisternae...
for ubiquitination and subsequent degradation by a protein-degrading complex, called the proteasome
Proteasome
Proteasomes are very large protein complexes inside all eukaryotes and archaea, and in some bacteria. In eukaryotes, they are located in the nucleus and the cytoplasm. The main function of the proteasome is to degrade unneeded or damaged proteins by proteolysis, a chemical reaction that breaks...
.
Recognition of misfolded or mutated proteins in the endoplasmic reticulum
The recognition of misfolded or mutated proteins depends on the detection of substructures within proteins such as exposed hydrophobic regions, unpaired cysteineCysteine
Cysteine is an α-amino acid with the chemical formula HO2CCHCH2SH. It is a non-essential amino acid, which means that it is biosynthesized in humans. Its codons are UGU and UGC. The side chain on cysteine is thiol, which is polar and thus cysteine is usually classified as a hydrophilic amino acid...
residues and immature glycans
Glycans
The term glycan refers to a polysaccharide or oligosaccharide. Glycans usually consist solely of O-glycosidic linkages of monosaccharides. For example, cellulose is a glycan composed of beta-1,4-linked D-glucose, and chitin is a glycan composed of beta-1,4-linked N-acetyl-D-glucosamine...
.
In mammalian cells for example, there exists a mechanism called glycan processing. In this mechanism, the lectin
Lectin
Lectins are sugar-binding proteins that are highly specific for their sugar moieties. They play a role in biological recognition phenomena involving cells and proteins. For example, some viruses use lectins to attach themselves to the cells of the host organism during infection...
-type chaperones calnexin
Calnexin
Calnexin is a 90kDa integral protein of the endoplasmic reticulum . It consists of a large N-terminal calcium-binding lumenal domain, a single transmembrane helix and a short , acidic cytoplasmic tail....
/calreticulin
Calreticulin
Calreticulin also known as calregulin, CRP55, CaBP3, calsequestrin-like protein, and endoplasmic reticulum resident protein 60 is a protein that in humans is encoded by the CALR gene....
(CNX/CRT) provide immature glycoproteins the opportunity to reach their native conformation. They can do this by way of reglucosylating these glycoproteins by an enzyme called UDP
Uridine diphosphate
Uridine diphosphate, abbreviated UDP, is a nucleoside diphosphate. It is an ester of pyrophosphoric acid with the nucleoside uridine. UDP consists of the pyrophosphate group, the pentose sugar ribose, and the nucleobase uracil.-See also:* Nucleoside...
-glucose
Glucose
Glucose is a simple sugar and an important carbohydrate in biology. Cells use it as the primary source of energy and a metabolic intermediate...
-glycoprotein
Glycoprotein
Glycoproteins are proteins that contain oligosaccharide chains covalently attached to polypeptide side-chains. The carbohydrate is attached to the protein in a cotranslational or posttranslational modification. This process is known as glycosylation. In proteins that have segments extending...
glucosyltransferase. Terminally misfolded proteins, however, must be extracted from CNX/CRT. This is carried out by EDEM and ER mannosidase I. This mannosidase removes one mannose
Mannose
Mannose is a sugar monomer of the aldohexose series of carbohydrates. Mannose is a C-2 epimer of glucose. It is not part of human metabolism, but is a component of microbial cell walls, and is therefore a target of the immune system and also of antibiotics....
residue from the glycoprotein and the latter is recognized by EDEM. Eventually EDEM will target the misfolded glycoproteins for degradation.
Retro-translocation into the cytosol
Because the ubiquitin–proteasome system (UPS) is located in the cytosol, terminally misfolded proteins have to be transported from the endoplasmic reticulum back into cytoplasm. It seems that a protein complex, called Sec61Sec61
Sec61 is an endoplasmic reticulum membrane protein translocator . It is a doughnut shaped pore through the membrane with 3 major subunits . It has a region called the plug that blocks transport into or out of the ER...
, constitutes the channel necessary for the transport of these misfolded proteins. Further, this translocation requires a driving force that determines the direction of transport. Since polyubiquitination is essential for the export of substrates, it is likely that this driving force is provided by ubiquitin-binding factors. One of these ubiquitin-binding factors is the Cdc48p-Npl4p-Ufd1p complex. It is known that human have the homolog of Cdc48p which is called Valosine-containing protein (VCP)/p97 and VCP/p97 have the same function of its yeast homolog Cdc48p. VCP/p97 transports substrates from endoplasmic reticulum to cytoplasm with its ATPase activity.
Ubiquitin-dependent degradation by the proteasome
The ubiquitination of terminally misfolded proteins is caused by a cascade of enzymatic reactions. The first of these reactions takes place when the ubiquitin-activating enzymeUbiquitin-activating enzyme
Ubiquitin-activating enzymes, also known as E1 enzymes, catalyze the first step in the ubiquitination reaction, which targets a protein for degradation via a proteasome. This covalent attachment of ubiquitin or ubiquitin-like proteins to targeted proteins is a major mechanism for regulating protein...
E1 hydrolyses ATP
Adenosine triphosphate
Adenosine-5'-triphosphate is a multifunctional nucleoside triphosphate used in cells as a coenzyme. It is often called the "molecular unit of currency" of intracellular energy transfer. ATP transports chemical energy within cells for metabolism...
and forms a high-energy thioester
Thioester
Thioesters are compounds with the functional group C-S-CO-C. They are the product of esterification between a carboxylic acid and a thiol. Thioesters are widespread in biochemistry, the best-known derivative being acetyl-CoA.-Synthesis:...
linkage between a cysteine residue in its active site and the C-terminus of ubiquitin. The resulting activated ubiquitin is then passed to E2, which is a ubiquitin-conjugating enzyme
Ubiquitin-conjugating enzyme
Ubiquitin-conjugating enzymes, also known as E2 enzymes and more rarely as ubiquitin-carrier enzymes, perform the second step in the ubiquitination reaction that targets a protein for degradation via the proteasome.The ubiquitination process covalently attaches ubiquitin, a short protein of 76...
. Another group of enzymes, more specifically ubiquitin protein ligases called E3, bind to the misfolded protein. Next they align the protein and E2, thus facilitating the attachment of ubiquitin to lysine residues of the misfolded protein. Following successive addition of ubiquitin molecules to lysine residues of the previously attached ubiquitin, a polyubiquitin chain is formed. A polyubiquitinated protein is produced and this is recognized by specific subunits
Protein subunit
In structural biology, a protein subunit or subunit protein is a single protein molecule that assembles with other protein molecules to form a protein complex: a multimeric or oligomeric protein. Many naturally occurring proteins and enzymes are multimeric...
in the 19S capping complexes of the 26S proteasome. Hereafter, the polypeptide chain is fed into the central chamber of the 20S core region that contains the proteolytically active sites. Ubiquitin is cleaved before terminal digestion by deubiquitinating enzymes. This third step is very closely associated with the second one, since ubiquitination takes place during the translocation event. However, the proteasomal degradation takes place in the cytoplasm.
ERAD ubiquitination machinery
The ER membrane anchored RING finger containing ubiquitin ligases Hrd1 and Doa10 are the major mediators of substrate ubiquitination during ERAD. The tail anchored membrane proteinMembrane protein
A membrane protein is a protein molecule that is attached to, or associated with the membrane of a cell or an organelle. More than half of all proteins interact with membranes.-Function:...
Ubc6 as well as Ubc1 and the Cue1 dependent membrane bound Ubc7 are the ubiquitin conjugating enzymes involved in ERAD.
Checkpoints
As the variation of ERAD-substrates is enormous, several variations of the ERAD mechanism have been proposed. Indeed, it was confirmed that soluble, membraneMembrane protein
A membrane protein is a protein molecule that is attached to, or associated with the membrane of a cell or an organelle. More than half of all proteins interact with membranes.-Function:...
and transmembrane proteins were recognized by different mechanisms. This led to the identification of 3 different pathways that constitute in fact 3 checkpoints.
- The first checkpoint is called ERAD-C and monitors the folding state of the cytosolic domains of membrane proteins. If defaults are detected in the cytosolic domains, this checkpoint will remove the misfolded protein.
- When the cytosolic domains are found to be correctly folded, the membrane protein will pass to a second checkpoint where the luminalLuminalLuminal may refer to:* A trade name for the anti-epileptic drug phenobarbital* Luminal , a 2004 film by Italian director Andrea Vecchiato starring Denis Lavant* In biology, pertaining to the lumen, the interior of a hollow structure...
domains are monitored. This second checkpoint is called the ERAD-L pathway. Not only membrane proteins surviving the first checkpoint are controlled for their luminal domains, also soluble proteins are inspected by this pathway as they are entirely luminal and thus bypass the first checkpoint. If a lesion in the luminal domains is detected, the involved protein is processed for ERAD using a set of factors including the vesicular trafficking machinery that transports misfolded proteins from the endoplasmic reticulum to the Golgi apparatusGolgi apparatusThe Golgi apparatus is an organelle found in most eukaryotic cells. It was identified in 1898 by the Italian physician Camillo Golgi, after whom the Golgi apparatus is named....
.
- Also a third checkpoint has been described that relies on the inspection of transmembrane domains of proteins. It is called the ERAD-M pathway but it is not very clear in which order it has to be placed with regard to the two previously described pathways.
Diseases associated with ERAD-malfunctioning
As ERAD is a central element of the secretory pathway, disorders in its activity can cause a range of human diseases. These disorders can be classified into two groups.The first group is the result of mutations in ERAD components, which subsequently lose their function. By losing their function, these components are not longer able to stabilize aberrant proteins so that the latter accumulate and damage the cell. A very important example of a disease caused by this first group of disorders is Parkinson's disease
Parkinson's disease
Parkinson's disease is a degenerative disorder of the central nervous system...
. It is caused by a mutation in the parkin gene
Gene
A gene is a molecular unit of heredity of a living organism. It is a name given to some stretches of DNA and RNA that code for a type of protein or for an RNA chain that has a function in the organism. Living beings depend on genes, as they specify all proteins and functional RNA chains...
. Parkin
Parkin
Parkin may refer to:* Parkin , an enzyme* Parkin , a type of cake* Parkin, Arkansas, a US city* Parkin , people with the surname Parkin* Parkin Archeological State Park also known as Parkin Site...
is a protein that functions in complex with CHIP as a ubiquitin ligase and overcomes the accumulation and aggregation of misfolded proteins.
[There are numerous debatable theories addressing the causes of Parkinson's Disease, besides the one presented here. Many of these can be found in the section of Wikipedia devoted to Parkinson's Disease]
In contrast to this first group of disorders, the second group is caused by premature degradation of secretory or membrane proteins. In this way, these proteins aren’t able to be deployed to distal compartments, as is the case in cystic fibrosis
Cystic fibrosis
Cystic fibrosis is a recessive genetic disease affecting most critically the lungs, and also the pancreas, liver, and intestine...
.
ERAD and HIV
As described before, the addition of polyubiquitin chains to ERAD substrates is crucial for their export. HIVHIV
Human immunodeficiency virus is a lentivirus that causes acquired immunodeficiency syndrome , a condition in humans in which progressive failure of the immune system allows life-threatening opportunistic infections and cancers to thrive...
uses an efficient mechanism to dislocate a single-membrane-spanning host protein, CD4
CD4
CD4 is a glycoprotein expressed on the surface of T helper cells, monocytes, macrophages, and dendritic cells. It was discovered in the late 1970s and was originally known as leu-3 and T4 before being named CD4 in 1984...
, from the ER and submits it to ERAD. CD4 is normally a stable protein and is not likely to be a target for ERAD. However, HIV produces the membrane protein Vpu
Vpu
Vpu is a HIV gene.Vpu stands for "Viral Protein U". Vpu is involved in viral budding, enhancing virion release from the cell by counteracting BST2 . In the absence of vpu, tetherin binds to the viral envelope and ties it to the cell membrane and other viral particles, impeding release of the viral...
that binds to CD4. As Vpu is phosphorylated, it mimics substrates for the E3 complex SCFβTrCP. In cells that are infected with HIV, SCFβTrCP interacts with Vpu and ubiquitinates CD4, which is subsequently degraded by the proteasome. Vpu itself escapes from the degradation.
Questions
The big questions for ERAD are:- How are misfolded proteins more specifically recognized?
- What is the channel for the retrotranslocation of luminal ER proteins?
- Which E3 ligase finally tags the proteins for the proteasomal degradation?
See also
- endoplasmic reticulumEndoplasmic reticulumThe endoplasmic reticulum is an organelle of cells in eukaryotic organisms that forms an interconnected network of tubules, vesicles, and cisternae...
- ubiquitination
- proteasomeProteasomeProteasomes are very large protein complexes inside all eukaryotes and archaea, and in some bacteria. In eukaryotes, they are located in the nucleus and the cytoplasm. The main function of the proteasome is to degrade unneeded or damaged proteins by proteolysis, a chemical reaction that breaks...
- protein foldingProtein foldingProtein folding is the process by which a protein structure assumes its functional shape or conformation. It is the physical process by which a polypeptide folds into its characteristic and functional three-dimensional structure from random coil....
- oxidative foldingOxidative foldingOxidative protein folding is a process that is responsible for the formation of disulfide bonds between cysteine residues in proteins. The driving force behind this process is a redox reaction, in which electrons are passed between several proteins and finally to a terminal electron acceptor.- In...