Deubiquitinating enzyme
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Deubiquitinating enzymes (DUBs), also known as deubiquitinating peptidases, deubiquitinating isopeptidases, deubiquitinases, ubiquitin proteases, ubiquitin hydrolases, ubiquitin isopeptidases, are a large group of
protease A protease (also called a peptidase, proteinase, or proteolytic enzyme) is an enzyme that catalyzes (increases reaction rate or "speeds up") proteolysis, breaking down proteins into smaller polypeptides or single amino acids, and spurring the ...
s that cleave
ubiquitin Ubiquitin is a small (8.6 kDa) regulatory protein found in most tissues of eukaryotic organisms, i.e., it is found ''ubiquitously''. It was discovered in 1975 by Gideon Goldstein and further characterized throughout the late 1970s and 1980s. Fo ...
from proteins. Ubiquitin is attached to proteins in order to regulate the degradation of proteins via the
proteasome Proteasomes are protein complexes which degrade unneeded or damaged proteins by proteolysis, a chemical reaction that breaks peptide bonds. Enzymes that help such reactions are called proteases. Proteasomes are part of a major mechanism by w ...
and
lysosome A lysosome () is a membrane-bound organelle found in many animal cells. They are spherical vesicles that contain hydrolytic enzymes that can break down many kinds of biomolecules. A lysosome has a specific composition, of both its membrane prot ...
; coordinate the cellular localisation of proteins; activate and inactivate proteins; and modulate protein-protein interactions. DUBs can reverse these effects by cleaving the peptide or isopeptide bond between ubiquitin and its substrate protein. In humans there are nearly 100 DUB genes, which can be classified into two main classes:
cysteine protease Cysteine proteases, also known as thiol proteases, are hydrolase enzymes that degrade proteins. These proteases share a common catalytic mechanism that involves a nucleophilic cysteine thiol in a catalytic triad or dyad. Discovered by Gopal Chund ...
s and
metalloprotease A metalloproteinase, or metalloprotease, is any protease enzyme whose catalytic mechanism involves a metal. An example is ADAM12 which plays a significant role in the fusion of muscle cells during embryo development, in a process known as myo ...
s. The cysteine proteases comprise ubiquitin-specific proteases (USPs), ubiquitin C-terminal hydrolases (UCHs), Machado-Josephin domain proteases (MJDs) and ovarian tumour proteases (OTU). The metalloprotease group contains only the Jab1/Mov34/Mpr1 Pad1 N-terminal+ (MPN+) (JAMM) domain proteases.


Classes

In humans there are 102 putative DUB genes, which can be classified into two main classes:
cysteine protease Cysteine proteases, also known as thiol proteases, are hydrolase enzymes that degrade proteins. These proteases share a common catalytic mechanism that involves a nucleophilic cysteine thiol in a catalytic triad or dyad. Discovered by Gopal Chund ...
s and
metalloprotease A metalloproteinase, or metalloprotease, is any protease enzyme whose catalytic mechanism involves a metal. An example is ADAM12 which plays a significant role in the fusion of muscle cells during embryo development, in a process known as myo ...
s, consisting of 58 ubiquitin-specific proteases (USPs), 4 ubiquitin C-terminal hydrolases (UCHs), 5 Machado-Josephin domain proteases (MJDs), 14 ovarian tumour proteases (OTU), and 14 Jab1/Mov34/Mpr1 Pad1 N-terminal+ (MPN+) (JAMM) domain-containing genes. 11 of these proteins are predicted to be non-functional, leaving 79 functional enzymes. In yeast, the USPs are known as ubiquitin-specific-processing proteases (UBPs).


Cysteine proteases

There are six main superfamilies of cysteine protease DUBs: * the ubiquitin-specific protease (USP/UBP) superfamily; ( USP1, USP2, USP3, USP4,
USP5 Ubiquitin specific peptidase 5 is an enzyme that in humans is encoded by the ''USP5'' gene. Interactions USP5 has been shown to interact with TADA3L Transcriptional adapter 3-like is a protein that in humans is encoded by the ''TADA3'' gene. ...
,
USP6 Ubiquitin carboxyl-terminal hydrolase 6 (USB6), also termed TRE17 and Tre-2, is a deubiquitinating enzyme that in humans is encoded by the hominid (i.e. found only in primates) ''USP6'' gene located at band 13.2 on the short (i.e. "p") arm of chro ...
,
USP7 Ubiquitin-specific-processing protease 7 (USP7), also known as ubiquitin carboxyl-terminal hydrolase 7 or herpesvirus-associated ubiquitin-specific protease (HAUSP), is an enzyme that in humans is encoded by the ''USP7'' gene. Function Regula ...
,
USP8 Ubiquitin carboxyl-terminal hydrolase 8 is an enzyme that in humans is encoded by the ''USP8'' gene. Interactions USP8 has been shown to interact with RNF41 and STAM2 Signal transducing adapter molecule 2 is a protein that in humans is encod ...
, USP9X, USP9Y,
USP10 Ubiquitin specific peptidase 10, also known as USP10, is an enzyme which in humans is encoded by the ''USP10'' gene. Function Ubiquitin is a highly conserved protein that is covalently linked to other proteins to regulate their function and de ...
, USP11, USP12, USP13,
USP14 Ubiquitin-specific protease 14 is an enzyme that in humans is encoded by the ''USP14'' gene. This gene encodes a member of the ubiquitin-specific processing (UBP) family of proteases that is a deubiquitinating enzyme (DUB) with His and Cys domains ...
, USP15, USP16, USP17, USP17L2, USP17L3, USP17L4, USP17L5, USP17L7, USP17L8,
USP18 Ubiquitin specific peptidase 18 (USP18), also known as UBP43, is a type I interferon receptor repressor and an isopeptidase. In humans, it is encoded by the ''USP18'' gene. USP18 is induced by the immune response to type I and III interferons, ...
, USP19,
USP20 Ubiquitin carboxyl-terminal hydrolase 20 is an enzyme that in humans is encoded by the ''USP20'' gene. Ubiquitin-specific protease 20 (USP20), also known as ubiquitin-binding protein 20 and VHL protein-interacting deubiquitinating enzyme 2 (VDU2), ...
, USP21, USP22, USP23, USP24, USP25,
USP26 USP26 is a peptidase enzyme. The USP26 gene is an X-linked gene exclusively expressed in the testis and it codes for the ubiquitin-specific protease 26. The USP26 gene is found at Xq26.2 on the X-chromosome as a single exon. The enzyme that this g ...
, USP27X,
USP28 USP may refer to: Government institutions * ''Unité Spéciale de la Police'', Luxembourg * United States Penitentiary, a prison * Utah State Prison, Draper, US Math and science * Ultra-short period planets, orbiting planets with periods shor ...
, USP29, USP30, USP31, USP32,
USP33 Ubiquitin carboxyl-terminal hydrolase 33 is an enzyme that in humans is encoded by the ''USP33'' gene. Interactions USP33 has been shown to interact with DIO2, SELENBP1 and Von Hippel–Lindau tumor suppressor ) hockey league, Supreme Hocke ...
, USP34, USP35, USP36, USP37, USP38,
USP39 U4/U6.U5 tri-snRNP-associated protein 2 is a protein that in humans is encoded by the ''USP39'' gene In biology, the word gene (from , ; "...Wilhelm Johannsen coined the word gene to describe the Mendelian units of heredity..." meaning '' ...
, USP40, USP41, USP42, USP43, USP44, USP45, USP46) * the ovarian tumour (OTU) superfamily ( OTUB1, OTUB2); * and the Machado-Josephin domain (MJD) superfamily. (
ATXN3 Ataxin-3 is a protein that in humans is encoded by the ''ATXN3'' gene. Clinical significance Machado–Joseph disease, also known as spinocerebellar ataxia-3, is an autosomal dominant neurologic disorder. The protein encoded by the ''ATXN3'' ...
, ATXN3L) * the ubiquitin C-terminal hydrolase (UCH) superfamily; (
BAP1 BRCA1 associated protein-1 (ubiquitin carboxy-terminal hydrolase) is a deubiquitinating enzyme that in humans is encoded by the ''BAP1'' gene. ''BAP1'' encodes an 80.4 kDa nuclear-localizing protein with a ubiquitin carboxy-terminal hydrolase (U ...
, UCHL1, UCHL3, UCHL5) *the MINDY family of K48-specific deubiquitinases; ( MINDY1, MINDY2, MINDY3, MINDY4) *the recently discovered ZUFSP family, at present solely represented by ZUP1 There is also a little known putative group of DUBs called the permutated papain fold peptidases of dsDNA viruses and eukaryote (PPPDEs) superfamily, which, if shown to be bona fide DUBs, would be the seventh in the cysteine protease class.


Metalloproteases

The Jab1/Mov34/Mpr1 Pad1 N-terminal+ (MPN+) (JAMM) domain superfamily proteins bind zinc and hence are metalloproteases.


Role of deubiquitinating enzymes

DUBs play several roles in the ubiquitin pathway. One of the best characterised functions of DUBs is the removal of monoubiqutin and polyubiquitin chains from proteins. These modifications are a
post translational modification Post-translational modification (PTM) is the covalent and generally enzymatic modification of proteins following protein biosynthesis. This process occurs in the endoplasmic reticulum and the golgi apparatus. Proteins are synthesized by ribosome ...
(addition to a protein after it has been made) where single ubiquitin proteins or chains of ubiquitin are added to lysines of a substrate protein. These ubiquitin modifications are added to proteins by the ubiquitination machinery;
ubiquitin-activating enzyme Ubiquitin-activating enzymes, also known as E1 enzymes, catalyze the first step in the ubiquitination reaction, which (among other things) can target a protein for degradation via a proteasome. This covalent bond of ubiquitin or ubiquitin-like pro ...
s (E1s),
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 ...
s (E2s) and
ubiquitin ligase A ubiquitin ligase (also called an E3 ubiquitin ligase) is a protein that recruits an E2 ubiquitin-conjugating enzyme that has been loaded with ubiquitin, recognizes a protein substrate, and assists or directly catalyzes the transfer of ubiquitin ...
s (E3s). The end result is ubiquitin bound to lysine residues via an isopeptide bond. Proteins are affected by these modifications in a number of ways: they regulate the degradation of proteins via the
proteasome Proteasomes are protein complexes which degrade unneeded or damaged proteins by proteolysis, a chemical reaction that breaks peptide bonds. Enzymes that help such reactions are called proteases. Proteasomes are part of a major mechanism by w ...
and
lysosome A lysosome () is a membrane-bound organelle found in many animal cells. They are spherical vesicles that contain hydrolytic enzymes that can break down many kinds of biomolecules. A lysosome has a specific composition, of both its membrane prot ...
; coordinate the cellular localisation of proteins; activate and inactivate proteins; and modulate protein-protein interactions. DUBs play the antagonistic role in this axis by removing these modifications, therefore reversing the fate of the proteins. In addition, a less understood role of DUBs is the cleavage of
ubiquitin-like protein Ubiquitin-like proteins (UBLs) are a family of small proteins involved in post-translational modification of other proteins in a cell, usually with a regulatory function. The UBL protein family derives its name from the first member of the class ...
s such as
SUMO is a form of competitive full-contact wrestling where a ''rikishi'' (wrestler) attempts to force his opponent out of a circular ring (''dohyō'') or into touching the ground with any body part other than the soles of his feet (usually by thr ...
and
NEDD8 NEDD8 is a protein that in humans is encoded by the ''NEDD8'' gene. (in ''saccharomyces cerevisiae'' this protein is known as Rub1) This ubiquitin-like (UBL) protein becomes covalently conjugated to a limited number of cellular proteins, in a proc ...
. Some DUBs may have the ability to cleave isopeptide bonds between these proteins and substrate proteins. They activate ubiquitin by the
proteolysis Proteolysis is the breakdown of proteins into smaller polypeptides or amino acids. Uncatalysed, the hydrolysis of peptide bonds is extremely slow, taking hundreds of years. Proteolysis is typically catalysed by cellular enzymes called protease ...
(breaking down) of the inactive expressed forms of ubiquitin. Ubiquitin is encoded in mammals by 4 different genes:
UBA52 60S ribosomal protein L40 (RPL40) is a protein that in humans is encoded by the ''UBA52'' gene. Function Ubiquitin is a highly conserved nuclear and cytoplasmic protein that has a major role in targeting cellular proteins for protein degradat ...
,
RPS27A 40S ribosomal protein S27a is a protein that in humans is encoded by the ''RPS27A'' gene. Ubiquitin, a highly conserved protein that has a major role in targeting cellular proteins for degradation by the 26S proteosome, is synthesized as a precurs ...
, UBB and
UBC The University of British Columbia (UBC) is a public research university with campuses near Vancouver and in Kelowna, British Columbia. Established in 1908, it is British Columbia's oldest university. The university ranks among the top three ...
. A similar set of genes is found in other eukaryotes such as yeast. The UBA52 and RPS27A genes produce ubiquitin that is fused to
ribosomal Ribosomes ( ) are macromolecular machines, found within all cells, that perform biological protein synthesis (mRNA translation). Ribosomes link amino acids together in the order specified by the codons of messenger RNA (mRNA) molecules to for ...
proteins and the UBB and UBC genes produce polyubiquitin (a chain of ubiquitin joined by their C- and
N-termini The N-terminus (also known as the amino-terminus, NH2-terminus, N-terminal end or amine-terminus) is the start of a protein or polypeptide, referring to the free amine group (-NH2) located at the end of a polypeptide. Within a peptide, the amin ...
). DUBs cleave the ubiquitin from these proteins, producing active single units of ubiquitin. DUBs also cleave single ubiquitin proteins that may have had their C-terminal tails accidentally bound to small cellular
nucleophile In chemistry, a nucleophile is a chemical species that forms bonds by donating an electron pair. All molecules and ions with a free pair of electrons or at least one pi bond can act as nucleophiles. Because nucleophiles donate electrons, they are ...
s. These ubiquitin-
amide In organic chemistry, an amide, also known as an organic amide or a carboxamide, is a compound with the general formula , where R, R', and R″ represent organic groups or hydrogen atoms. The amide group is called a peptide bond when it is ...
s and ubiquitin-
thioester In organic chemistry, thioesters are organosulfur compounds with the functional group . They are analogous to carboxylate esters () with the sulfur in the thioester playing the role of the linking oxygen in the carboxylate ester, as implied by t ...
s may be formed during standard ubiquitination reactions by the E1-E2-E3 cascade.
Glutathione Glutathione (GSH, ) is an antioxidant in plants, animals, fungi, and some bacteria and archaea. Glutathione is capable of preventing damage to important cellular components caused by sources such as reactive oxygen species, free radicals, pero ...
and polyamines are two nucleophiles that might attack the thiolester bond between ubiquitin and these enzymes. Ubiquitin C-terminal hydrolase is an example of the DUB that hydrolyses these bonds with broad specificity. Free polyubiquitin chains are cleaved by DUBs to produce monoubiquitin. The chains may be produced by the E1-E2-E3 machinery in the cell free from any substrate protein. Another source of free polyubiquitin is the product of ubiquitin-substrate cleavage. If DUBs cleave the base of the polyubiquitin chain that is attached to a protein, the whole chain will become free and needs to be recycled by DUBs.


Domains

DUBs often contain a
catalytic Catalysis () is the process of increasing the rate of a chemical reaction by adding a substance known as a catalyst (). Catalysts are not consumed in the reaction and remain unchanged after it. If the reaction is rapid and the catalyst recyc ...
domain Domain may refer to: Mathematics *Domain of a function, the set of input values for which the (total) function is defined **Domain of definition of a partial function **Natural domain of a partial function **Domain of holomorphy of a function * Do ...
surrounded by one or more accessory domains, some of which contribute to target recognition. These additional domains include domain present in ubiquitin-specific proteases (DUSP) domain; ubiquitin-like (UBL) domain; meprin and TRAF homology (MATH) domain; zinc-finger ubiquitin-specific protease (ZnF-UBP) domain; zinc-finger myeloid, nervy and DEAF1 (ZnF-MYND) domain; ubiquitin-associated (UBA) domain; CHORD-SGT1 (CS) domain; microtubule-interacting and trafficking (MIT) domain; rhodenase-like domain; TBC/RABGAP domain; and B-box domain.


Catalytic domain

The catalytic domain of DUBs is what classifies them into particular groups; USPs, OTUs, MJDs, UCHs and MPN+/JAMMs. The first 4 groups are
cysteine protease Cysteine proteases, also known as thiol proteases, are hydrolase enzymes that degrade proteins. These proteases share a common catalytic mechanism that involves a nucleophilic cysteine thiol in a catalytic triad or dyad. Discovered by Gopal Chund ...
s, whereas the latter is a zinc
metalloprotease A metalloproteinase, or metalloprotease, is any protease enzyme whose catalytic mechanism involves a metal. An example is ADAM12 which plays a significant role in the fusion of muscle cells during embryo development, in a process known as myo ...
. The cysteine protease DUBs are papain-like and thus have a similar mechanism of action. They use either catalytic dyads or triads (either two or three
amino acids Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although hundreds of amino acids exist in nature, by far the most important are the alpha-amino acids, which comprise proteins. Only 22 alpha am ...
) to catalyse the
hydrolysis Hydrolysis (; ) is any chemical reaction in which a molecule of water breaks one or more chemical bonds. The term is used broadly for substitution reaction, substitution, elimination reaction, elimination, and solvation reactions in which water ...
of the amide bonds between ubiquitin and the substrate. The active site residues that contribute to the catalytic activity of the cysteine protease DUBs are
cysteine Cysteine (symbol Cys or C; ) is a semiessential proteinogenic amino acid with the formula . The thiol side chain in cysteine often participates in enzymatic reactions as a nucleophile. When present as a deprotonated catalytic residue, sometime ...
(dyad/triad),
histidine Histidine (symbol His or H) is an essential amino acid that is used in the biosynthesis of proteins. It contains an α-amino group (which is in the protonated –NH3+ form under biological conditions), a carboxylic acid group (which is in the de ...
(dyad/triad) and
aspartate Aspartic acid (symbol Asp or D; the ionic form is known as aspartate), is an α-amino acid that is used in the biosynthesis of proteins. Like all other amino acids, it contains an amino group and a carboxylic acid. Its α-amino group is in the pro ...
or
asparagine Asparagine (symbol Asn or N) is an α-amino acid that is used in the biosynthesis of proteins. It contains an α-amino group (which is in the protonated −NH form under biological conditions), an α-carboxylic acid group (which is in the depro ...
(triad only). The histidine is polarised by the aspartate or asparagine in catalytic triads or by other ways in dyads. This polarised residue lowers the pKa of the cysteine, allowing it to perform a
nucleophilic attack In chemistry, a nucleophile is a chemical species that forms bonds by donating an electron pair. All molecules and ions with a free pair of electrons or at least one pi bond can act as nucleophiles. Because nucleophiles donate electrons, they are ...
on the isopeptide bond between the ubiquitin
C-terminus The C-terminus (also known as the carboxyl-terminus, carboxy-terminus, C-terminal tail, C-terminal end, or COOH-terminus) is the end of an amino acid chain (protein or polypeptide), terminated by a free carboxyl group (-COOH). When the protein is ...
and the substrate
lysine Lysine (symbol Lys or K) is an α-amino acid that is a precursor to many proteins. It contains an α-amino group (which is in the protonated form under biological conditions), an α-carboxylic acid group (which is in the deprotonated −C ...
. Metalloproteases coordinate
zinc Zinc is a chemical element with the symbol Zn and atomic number 30. Zinc is a slightly brittle metal at room temperature and has a shiny-greyish appearance when oxidation is removed. It is the first element in group 12 (IIB) of the periodi ...
ions with histidine, aspartate and
serine Serine (symbol Ser or S) is an α-amino acid that is used in the biosynthesis of proteins. It contains an α-amino group (which is in the protonated − form under biological conditions), a carboxyl group (which is in the deprotonated − form un ...
residues, which activate water molecules and allows them to attack the isopeptide bond.


UBL

Ubiquitin-like (UBL) domains have a similar structure (fold) to ubiquitin, except they lack the terminal glycine residues. 18 USPs are proposed to have UBL domains. Only 2 other DUBs have UBLs outside the USP group: OTU1 and VCPIP1. USP4, USP7, USP11, USP15, USP32, USP40 and USP47 have multiple UBL domains. Sometimes the UBL domains are in tandem, such as in USP7 where 5 tandem
C-terminal The C-terminus (also known as the carboxyl-terminus, carboxy-terminus, C-terminal tail, C-terminal end, or COOH-terminus) is the end of an amino acid chain (protein or polypeptide), terminated by a free carboxyl group (-COOH). When the protein is ...
UBL domains are present. USP4, USP6, USP11, USP15, USP19, USP31, USP32 and USP43 have UBL domains inserted into the catalytic domain. The functions of UBL domains are different between USPs, but commonly they regulate USP catalytic activity. They can coordinate localisation at the proteasome (USP14); negatively regulate USPs by competing for the catalytic site of the USP (USP4), and induce conformational changes to increase catalytic activity (USP7). Like other UBL domains, the structure of USP UBL domains show a β-grasp fold.


DUSP

Single or multiple tandem DUSP domains of approximately 120 residues are found in six USPs. The function of the DUSP domain is currently unknown but it may play a role in protein-protein interaction, in particular to DUBs substrate recognition. This is predicted because of the hydrophobic cleft present in the DUSP domain of USP15 and that some protein interactions with DUSP containing USPs do not occur without these domains. The DUSP domain displays a novel tripod-like fold comprising three
helices A helix () is a shape like a corkscrew or spiral staircase. It is a type of smooth space curve with tangent lines at a constant angle to a fixed axis. Helices are important in biology, as the DNA molecule is formed as two intertwined helices, ...
and an anti-parallel
beta-sheet The beta sheet, (β-sheet) (also β-pleated sheet) is a common motif of the regular protein secondary structure. Beta sheets consist of beta strands (β-strands) connected laterally by at least two or three backbone hydrogen bonds, forming a g ...
made of three strands. This fold resembles the legs (helices) and seat (beta-sheet) of the tripod. Within most DUSP domains in USPs there is a conserved sequence of amino acids known as the PGPI motif. This is a sequence of four amino acids;
proline Proline (symbol Pro or P) is an organic acid classed as a proteinogenic amino acid (used in the biosynthesis of proteins), although it does not contain the amino group but is rather a secondary amine. The secondary amine nitrogen is in the prot ...
,
glycine Glycine (symbol Gly or G; ) is an amino acid that has a single hydrogen atom as its side chain. It is the simplest stable amino acid (carbamic acid is unstable), with the chemical formula NH2‐ CH2‐ COOH. Glycine is one of the proteinogeni ...
, proline and
isoleucine Isoleucine (symbol Ile or I) is an α-amino acid that is used in the biosynthesis of proteins. It contains an α-amino group (which is in the protonated −NH form under biological conditions), an α-carboxylic acid group (which is in the deprot ...
, which packs against the three-helix bundle and is highly ordered.


Role in disease

The full extent of the role of DUBs in diseases remains to be elucidated. Their involvement in disease is predicted due to known roles in physiological processes that are involved in disease states; including cancer and neurological disorders. The enzyme USP28 is over-expressed in different types of
cancer Cancer is a group of diseases involving abnormal cell growth with the potential to invade or spread to other parts of the body. These contrast with benign tumors, which do not spread. Possible signs and symptoms include a lump, abnormal b ...
such as colon or lung. In addition,
USP28 USP may refer to: Government institutions * ''Unité Spéciale de la Police'', Luxembourg * United States Penitentiary, a prison * Utah State Prison, Draper, US Math and science * Ultra-short period planets, orbiting planets with periods shor ...
deubiquitinates and stabilizes important oncogenes such as
c-Myc ''Myc'' is a family of regulator genes and proto-oncogenes that code for transcription factors. The ''Myc'' family consists of three related human genes: ''c-myc'' (MYC), ''l-myc'' (MYCL), and ''n-myc'' (MYCN). ''c-myc'' (also sometimes referre ...
,
Notch1 Neurogenic locus notch homolog protein 1 (Notch 1) is a protein encoded in humans by the ''NOTCH1'' gene. Notch 1 is a single-pass transmembrane receptor. Function This gene encodes a member of the Notch family. Members of this Type 1 transm ...
,
c-jun Transcription factor Jun is a protein that in humans is encoded by the ''JUN'' gene. c-Jun, in combination with protein c-Fos, forms the AP-1 early response transcription factor. It was first identified as the Fos-binding protein p39 and only lat ...
or ΔNp63. In squamous tumors, USP28 regulates the resistance to chemotherapy regulating DNA repair via ΔNp63-Fanconia anemia pathway axis. The deubiquitinating enzymes UCH-L3 and YUH1 are able to hydrolyse mutant ubiquitin UBB+1 despite of the fact that the glycine at position 76 is mutated. UCH-L1 levels are high in various types of malignancies (
cancer Cancer is a group of diseases involving abnormal cell growth with the potential to invade or spread to other parts of the body. These contrast with benign tumors, which do not spread. Possible signs and symptoms include a lump, abnormal b ...
).


Role in the Cell Cycle

DUBs play an active role in modulating the cell cycle. Ubiquitin-specific-processing protease (USP) is a family of deubiquitinating enzymes that play a crucial role in cell cycle regulation. Two such enzymes include USP17 and USP44. USP17 regulates pathways responsible for progressing cells through the cell cycle. Its targets include regulators of Ras, CDK2, and Cyclin A. USP44 plays an important role in anaphase initiation. New research into the mitotic checkpoint has revealed a novel role for USP44 in regulating cell cycle progression.


USP Regulation of Ras

The ERK Pathway allows for the transduction of external mitogenic signals into intracellular signals promoting cellular proliferation. One of the key regulators of this pathways is Ras, a GTPase that, upon activation, binds GTP to “turn on” the subsequent signaling cascade. Ras converting enzyme 1 (RCE1) post-translationally cleaves the 3 residues on the C-terminus of Ras, allowing Ras to properly localize to the plasma membrane. USP17 acts to deubiquitinate K63-ubiquitin domains on RCE1. Such stabilization of RCE1 allows for proper localization of Ras, thus promoting proliferation upon activation of early receptors in the ERK Pathway. Ras hyperactivity can result in cell cycle dysregulation. Thus, regulation of Ras through USP17 acts as another point in Ras regulation.


USP Regulation of G1-S Transition

Cyclin-dependent kinases (CDKs) are a family of enzymes that phosphorylate serine and threonine residues to drive the cell through the cell cycle. Activation of CDK2 is critical for the G1-S transition. For CDK2 to be activated, cyclin A must bind to the cyclin-dependent kinase complex (CDKC). Cell division cycle 25A (CDC25A) is a phosphatase that removes an inhibitory phosphate group from CDK2. While ubiquitination would mark CDC25A for degradation, thus blocking progression to S phase, USP17 deubiquitinates CDC25A. An increase in CDC25A stability promotes CDKC activity, thus driving the cell through the G1-S transition. USP17 also regulates cell cycle progression by acting on SETD8 to downregulate transcription of cyclin-dependent kinase inhibitor 1 (CDKN1A), also known as p21. CDKN1A binds to and inhibits CDK2 using its N-terminal binding domain, thus blocking progression through the G1-S transition. SETD8, a methyltransferase, uses S-Adenosyl methionine to methylate the Lys20 residue of histone 4, resulting in the condensation of chromosomes. This compaction of the DNA downregulates CDKN1A transcription. USP17 deubiquitinates SETD8, thus reducing its propensity for degradation and increasing its intracellular stability. The resulting downregulation in CDKN1A transcription promotes CDK2 activity, allowing the cell to progress through the G1-S transition. See schematic of the role of DUBs in the cell cycle regulation.


USP44 in Anaphase Initiation

The spindle checkpoint (also referred to as the mitotic checkpoint) ensures proper separation of chromosomes. Broadly, the mitotic checkpoint promotes fidelity in chromosomal segregation, increasing the likelihood that each daughter cell receives only one duplicated chromosome. Such a mechanism is crucial, as errors in chromosomal separation have been implicated in cancer, birth defects, and antibiotic resistance in pathogens. One of the core regulator proteins is the anaphase-promoting complex (APC/C). APC/C ubiquitinates securin. The resulting destruction of securing release separase, which hydrolyzes cohesion – the protein that binds sister chromatids together. New research from Stegmeier and colleagues published in the journal Nature demonstrates a crucial role for USP44 in regulating the spindle checkpoint. Using an shRNA screen, USP44 was identified to stabilize the inhibition of APC/C The binding of CDC20 to APC/C is required for the ubiquitination of securin. A protein called hMAD2 can form an inactive trimer with APC and CDC20, forming the hMAD2-CDC-APC complex. Upon the ubiquitination of CDC20 by UbcH10, hMAD2 dissociates, and APC/C becomes active. It is important to note that ubiquitination of CDC20 does not serve to mark it for degradation, but rather promote dissociation of hMAD2 from the hMAD2-CDC-APC complex. USP44, a ubiquitin-specific-processing protease, can stabilize the inactive hMAD2-CDC-APC complex by counteracting UbcH10 ubiquitination. This blocks hMAD2 dissociation and allows for proper regulation of APC/C, keeping it inactive until proper attachment of the mitotic spindle. Upon proper attachment, switch-like behavior allows for the activation of APC/C. This results in the cleavage of cohesion, allowing for the separation of sister chromatids.


Role in p53-mediated DNA Damage Repair

DNA damage can prove catastrophic for an organism. Mechanisms for DNA mutation include oxidative stress, DNA replication errors, exogenous carcinogens, radiation, and spontaneous base mutation. Upon DNA damage, cell cycle progression is halted to prevent propagation of the mutation. The TP53 gene (also known as p53) is crucial in ensuring the conservation of the genome. Deubiquitinating enzymes play an integral role in maintaining p53’s function. In healthy cells, p53 activates the E3 ubiquitin ligase MDM2 which in turn ubiquitinates p53. This creates a negative feedback loop, whereby the degradation of p53 allows for cells to flow through the cell cycle. Upon DNA damage, Ubiquitin-specific-processing protease 7 (USP7) stabilizes p53 by cleaving ubiquitin. For USP7 to deubiquitinate p53, it must localize to the nucleus. However, no nuclear localization sequence (NLS) has been found. Despite no known NLS, one study showed that, upon deletion of USP7’s N-terminus, no nuclear localization occurred. It is possible that other proteins facilitate nuclear entry of USP7. Once stabilized, p53 can exert its tumor suppression function. Downstream pathways of p53 act to either halt cell cycle progression in G1 or G2 phases of the cell cycle or promote cell-death, depending on the severity of the DNA damage. See schematic of the role of USP7 in the p53-dependent pathway. or promote cell-death, depending on the severity of the DNA damage. See schematic of the role of USP7 in the p53-dependent pathway.


References

{{InterPro content, IPR006615 Protein domains Enzymes