Mouse double minute 2 homolog (MDM2) also known asE3 ubiquitin-protein ligase Mdm2 is aprotein that in humans is encoded by theMDM2gene.[5][6] Mdm2 is an important negative regulator of thep53 tumor suppressor. Mdm2 protein functions both as anE3 ubiquitin ligase that recognizes theN-terminal trans-activation domain (TAD) of the p53 tumor suppressor and as an inhibitor of p53 transcriptional activation.
The murinedouble minute (mdm2)oncogene, which codes for the Mdm2 protein, was originally cloned, along with two other genes (mdm1 and mdm3) from the transformed mouse cell line 3T3-DM. Mdm2 overexpression, in cooperation with oncogenicRas, promotes transformation of primary rodent fibroblasts, andmdm2 expression led to tumor formation innude mice. The human homologue of this protein was later identified and is sometimes called Hdm2. Further supporting the role of mdm2 as anoncogene, several humantumor types have been shown to have increased levels of Mdm2, including soft tissue sarcomas and osteosarcomas as well as breast tumors.
An additional Mdm2 family member, Mdm4 (also called MdmX), has been discovered and is also an important negative regulator ofp53.
The key target of Mdm2 is thep53 tumor suppressor. Mdm2 has been identified as a p53 interacting protein that represses p53 transcriptional activity. Mdm2 achieves this repression by binding to and blocking theN-terminal trans-activation domain of p53. Mdm2 is a p53 responsive gene—that is, its transcription can be activated by p53. Thus when p53 is stabilized, the transcription of Mdm2 is also induced, resulting in higher Mdm2 protein levels.
The E3 ubiquitin ligase MDM2 is a negative regulator of the p53 tumor suppressor protein. MDM2 binds and ubiquitinates p53, facilitating it for degradation. p53 can induce transcription of MDM2, generating anegative feedback loop.[7] Mdm2 also acts as anE3 ubiquitin ligase, targeting both itself and p53 for degradation by theproteasome (see alsoubiquitin). Severallysine residues in p53C-terminus have been identified as the sites of ubiquitination, and it has been shown that p53 protein levels are downregulated by Mdm2 in a proteasome-dependent manner. Mdm2 is capable of auto-polyubiquitination, and in complex with p300, a cooperating E3 ubiquitin ligase, is capable of polyubiquitinating p53. In this manner, Mdm2 and p53 are the members of a negative feedback control loop that keeps the level of p53 low in the absence of p53-stabilizing signals. This loop can be interfered with bykinases and genes likep14arf when p53 activation signals, includingDNA damage, are high.
The full-length transcript of the mdm2 gene encodes a protein of 491amino acids with a predicted molecular weight of 56kDa. This protein contains several conservedstructural domains including an N-terminal p53 interaction domain, the structure of which has been solved usingx-ray crystallography. The Mdm2 protein also contains a central acidic domain (residues 230–300). Thephosphorylation of residues within this domain appears to be important for regulation of Mdm2 function. In addition, this region contains nuclear export and import signals that are essential for proper nuclear-cytoplasmic trafficking of Mdm2. Another conserved domain within the Mdm2 protein is azinc finger domain, the function of which is poorly understood.
Mdm2 also contains aC-terminal RING domain (amino acid residues 430–480), which contains a Cis3-His2-Cis3 consensus that coordinates two ions ofzinc. These residues are required for zinc binding, which is essential for proper folding of the RING domain. The RING domain of Mdm2 confersE3 ubiquitin ligase activity and is sufficient for E3 ligase activity in Mdm2 RING autoubiquitination. The RING domain of Mdm2 is unique in that it incorporates a conservedWalker A or P-loop motif characteristic ofnucleotide binding proteins, as well as a nucleolar localization sequence. The RING domain also binds specifically toRNA, although the function of this is poorly understood.
There are several known mechanisms for regulation of Mdm2. One of these mechanisms isphosphorylation of the Mdm2 protein. Mdm2 is phosphorylated at multiple sites in cells. FollowingDNA damage, phosphorylation of Mdm2 leads to changes in protein function and stabilization ofp53. Additionally, phosphorylation at certain residues within the central acidic domain of Mdm2 may stimulate its ability to target p53 for degradation.HIPK2 is a protein that regulates Mdm2 in this way. The induction of thep14arf protein, the alternate reading frame product of thep16INK4a locus, is also a mechanism of negatively regulating the p53-Mdm2 interaction. p14arf directly interacts with Mdm2 and leads to up-regulation of p53 transcriptional response. ARF sequesters Mdm2 in thenucleolus, resulting in inhibition of nuclear export and activation of p53, since nuclear export is essential for proper p53 degradation.
Inhibitors of the MDM2-p53 interaction include the cis-imidazoline analognutlin.[8]
Levels and stability of Mdm2 are also modulated by ubiquitylation. Mdm2 auto ubiquitylates itself, which allows for its degradation by theproteasome. Mdm2 also interacts with a ubiquitin specific protease,USP7, which can reverse Mdm2-ubiquitylation and prevent it from being degraded by the proteasome. USP7 also protects from degradation the p53 protein, which is a major target of Mdm2. Thus Mdm2 and USP7 form an intricate circuit to finely regulate the stability and activity of p53, whose levels are critical for its function.
Mdm2 overexpression was shown to inhibit DNA double-strand break repair mediated through a novel, direct interaction between Mdm2 and Nbs1 and independent of p53. Regardless of p53 status, increased levels of Mdm2, but not Mdm2 lacking its Nbs1-binding domain, caused delays in DNA break repair, chromosomal abnormalities, and genome instability. These data demonstrated Mdm2-induced genome instability can be mediated through Mdm2:Nbs1 interactions and independent from its association with p53.[56]
^Vassilev LT, Vu BT, Graves B, Carvajal D, Podlaski F, Filipovic Z, Kong N, Kammlott U, Lukacs C, Klein C, Fotouhi N, Liu EA (February 2004). "In vivo activation of the p53 pathway by small-molecule antagonists of MDM2".Science.303 (5659):844–8.Bibcode:2004Sci...303..844V.doi:10.1126/science.1092472.PMID14704432.S2CID16132757.
^Zhao L, Samuels T, Winckler S, Korgaonkar C, Tompkins V, Horne MC, Quelle DE (January 2003). "Cyclin G1 has growth inhibitory activity linked to the ARF-Mdm2-p53 and pRb tumor suppressor pathways".Molecular Cancer Research.1 (3):195–206.PMID12556559.
^Yogosawa S, Miyauchi Y, Honda R, Tanaka H, Yasuda H (March 2003). "Mammalian Numb is a target protein of Mdm2, ubiquitin ligase".Biochemical and Biophysical Research Communications.302 (4):869–72.doi:10.1016/S0006-291X(03)00282-1.PMID12646252.
Cahilly-Snyder L, Yang-Feng T, Francke U, George DL (May 1987). "Molecular analysis and chromosomal mapping of amplified genes isolated from a transformed mouse 3T3 cell line".Somatic Cell and Molecular Genetics.13 (3):235–44.doi:10.1007/BF01535205.PMID3474784.S2CID27300300.
Momand J, Zambetti GP, Olson DC, George D, Levine AJ (June 1992). "The mdm-2 oncogene product forms a complex with the p53 protein and inhibits p53-mediated transactivation".Cell.69 (7):1237–45.doi:10.1016/0092-8674(92)90644-R.PMID1535557.S2CID22594319.
