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E3 Ubiquitin Ligases


Overview of E3 Ubiquitin Ligases

An 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 from the E2 to the protein substrate. The ubiquitin protein is linked to lysine from target protein by ISO peptide bond. E3 ligase interacts with target protein and E2 enzyme, thus giving E2 substrate specificity.

Usually, E3s polyubiquitinate their substrate with Lys48-linked chains of ubiquitin, targeting substrates to be destroyed by proteasomes. However, many other types of connections are possible and change the activity, interaction, or localization of proteins. The ubiquitination of E3 ligase regulates many regions, such as cell transport, DNA repair, and signal transduction, and is important in cell biology. E3 ligase is also a key participant in cell cycle control, mediating the degradation of cyclin and cyclin-dependent kinase inhibitor protein. The human genome encodes more than 600 putative E3 ligase, which makes the substrate have enormous diversity.

Mono- and poly- ubiquitylation

Proteins can be labeled with a single ubiquitin molecule (monoubiquitylation), or a variety of different ubiquitin chains (polyubiquitination). E3 ubiquitin ligase catalyzes polyubiquitination events in the same way as a single ubiquitination mechanism, using lysine residues from ubiquitin molecules currently attached to substrate proteins to attack the C-terminal of new ubiquitin molecules.

Monoubiquitination is related to the membrane protein endocytosis pathway. For example, the phosphorylation of the tyrosine at position 1045 in the epidermal growth factor receptor (EGFR) can recruit RING-type E3 ligase C-Cbl through the SH2 domain. C-Cbl monoubiquitylates EGFR and signal transduction is internalized and transported to lysosomes.

Monoubiquitination can also regulate the localization of cytosolic proteins. For example, the E3 ligase MDM2 can regulate degradation or nuclear output by the ubiquitination of P53. These events occur in a concentration-dependent manner, suggesting that regulating the concentration of E3 ligase is a cellular regulatory strategy for controlling proteins homeostasis and localization.

Ubiquitin ligase families

Humans have estimated 500-1000 E3 ligases, which impart substrate specificity onto the E1 and E2. E3 ligase is divided into four series, HECT, RING-finger, U-box and PHD-finger. The RING-finger E3 ligases are the largest family and contain the anaphase-promoting complex (APC) and the SCF complex (Skp1-Cullin-F-box protein complex).

  • UBE3A

Ubiquitin-protein ligase E3A (UBE3A), also known as E6AP Ubiquitin-protein ligase, is an enzyme encoded by the UBE3A gene in humans. The enzyme participates in targeting protein degradation in cells. Ubiquitin protein ligase E3A attaches a small marker protein called ubiquitin to a protein that should be degraded. The cell structure known as proteasome recognizes and digests ubiquitin-labeled proteins. The UBE3A gene is located in 15q11-13 of human chromosome region. Other abnormalities in chromosome 15 can also lead to Angelman syndrome. Mutations in the UBE3A gene are responsible for some cases of Angelman syndrome and Prader-Willi syndrome. Most of these mutations lead to abnormal short of ubiquitin protein ligase E3A, and the enzyme loses its function.

  • MDM2

Mouse double minute 2 homolog (MDM2), also known as E3 ubiquitin-protein ligase Mdm2, is a protein encoded by the MDM2 gene in mice. Mdm2 is an important negative regulator of P53 tumor suppressor. MDM2 is a key regulator of the activity and stability of P53 tumor suppressor protein. It can bind and inhibit the transactivation domain of P53. In addition, MDM2 controls P53 stability by acting as an E3 ligase in ubiquitination and shuttling P53 from the nucleus to the cytoplasm for subsequent degradation.

  • NEDD4

E3 ubiquitin-protein ligase NEDD4, also known as neural precursor cells, expresses a developmentally down-regulated protein 4 (NEDD4), an enzyme encoded by the NEDD4 gene in humans. NEDD4 regulates a large number of membrane proteins, such as ion channels and membrane receptors, through ubiquitination and endocytosis. This means that NEDD4 can bind to many proteins and regulate protein stability. NEDD4 is involved in many processes, including insulin-like growth factor signaling, neuronal structure and virus germination. NEDD4 is an essential protein for animal development and survival.

  • NEDD4L

NEDD4L or NEDD4-2 are enzymes of the NEDD4 family (ubiquitin ligase). In humans, these proteins are encoded by the NEDD4L gene. In mice, this protein are commonly referred to as NEDD4-2. Although NEDD4 homologues exist in all eukaryotes, NEDD4L protein is restricted to vertebrates. NEDD4L protein is known to regulate many membrane proteins through ubiquitination and endocytosis. Ubiquitin-induced NEDD4-2 down-regulates the epithelial sodium channel (ENaC) of the renal collecting duct, thereby antagonizing aldosterone and increasing salt excretion.

  • RNF4

RING finger protein 4 is a protein encoded by the RNF4 gene in humans. The protein encoded by this gene contains RING finger domain and acts as a transcription factor. The protein has been shown to interact with and inhibit the activity of TRPS1, a GATA-mediated transcription inhibitor. It was found that ZNF278/PATZ1 interacted with this protein, thereby reducing the increase of androgen receptor-dependent transcription mediated by this protein. Studies on the mouse and rat counterparts showed the role of the protein in spermatogenesis.

  • SMURF2

E3 ubiquitin-protein ligase SMURF2 is an enzyme that in humans is encoded by the SMURF2 gene. SMURF2 (SMAD ubiquitin regulatory factor 2) is a cytoplasmic E3 ubiquitin ligase of about 86 kDa, 748 amino acids that negatively regulates TGF-beta signaling by facilitating proteasome degradation of some SMAD signaling molecules. It also affects cell cycle progression, reaching a peak during mitosis and binding to MAD2L1 and NEDD9 / CASL.

  • PIAS1

E3 SUMO-protein ligase PIAS1 is an enzyme that in humans is encoded by the PIAS1 gene. The gene encodes a member of the mammalian PIAS family. The member contains putative zinc binding motif and high acid region. It inhibits STAT1-mediated gene activation and DNA binding activity, binds to Gu protein/RNA helicase II/DEAD cassette peptide 21, and interacts with androgen receptor (AR). It acts as a nuclear receptor transcription coregulator in the testis and may play a role in AR initiation and maintenance of spermatogenesis.

Role in disease

E3 ubiquitin ligase regulates homeostasis, cell cycle and DNA repair pathways in vivo, and so many of these proteins, including the well-known MDM2, BRCA1 and Von Hippel-Lindau tumor suppressors, also play an important role in various cancers. For example, MDM2 mutations have been found in gastric, renal and hepatocellular carcinomas (and others), and the MDM2 concentrations will be enhanced by increasing the affinity of their promoters to Sp1 transcription factors, resulting in an increase in the mRNA level of MDM2.

References:

  • Hershko A, Ciechanover A . The ubiquitin system. Annual Review of Biochemistry. 1998, 67: 425–79.
  • Metzger MB, Hristova VA, Weissman AM. HECT and RING finger families of E3 ubiquitin ligases at a glance. Journal of Cell Science. 2012, 125 (Pt 3): 531–7.
  • Nakayama KI, Nakayama K. Ubiquitin ligases: cell-cycle control and cancer. Nature Reviews. Cancer. May 2006, 6 (5): 369–81.
  • Hou YC, Deng JY. Role of E3 ubiquitin ligases in gastric cancer. World Journal of Gastroenterology. Jan 2015, 21 (3): 786–93.

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