Review Article Regulation of p63 Protein Stability via...

Review ArticleRegulation of p63 Protein Stability viaUbiquitin-Proteasome Pathway

Chenghua Li and Zhi-Xiong Xiao

Center of GrowthMetabolism andAging Key Laboratory of Biological Resources and Ecological Environment ofMinistry of EducationCollege of Life Sciences Sichuan University Chengdu 610064 China

Correspondence should be addressed to Zhi-Xiong Xiao bmc605hotmailcom

Received 25 December 2013 Revised 10 March 2014 Accepted 28 March 2014 Published 15 April 2014

Academic Editor Xin-yuan Guan

Copyright copy 2014 C Li and Z-X XiaoThis is an open access article distributed under the Creative Commons Attribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

The p53-related p63 gene encodes multiple protein isoforms which are involved in a variety of biological activities p63 proteinstability is mainly regulated by the ubiquitin-dependent proteasomal degradation pathway Several ubiquitin E3 ligases have beenidentified and some protein kinases as well as other kinds of proteins are involved in regulation of p63 protein stability Theseregulators are responsive to diverse extracellular signaling resulting in changes of the p63 protein levels and impacting differentbiological processes

1 Introduction

The p53 family member p63 gene is located on humanchromosome 3q27ndash29 In contrast to the high frequency ofp53 mutations in cancers p63 gene is rarely mutated [1 2]However up to 60 of squamous cell carcinomas showelevated p63 protein levels [3] In addition mutations in thep63 gene have been linked to several human developmentaldiseases A vast body of evidence demonstrates that p63 arekey transcription factors involved in cell growth prolifer-ation apoptosis and differentiation and play an essentialrole in epithelial stem cell biology and development [4ndash8]Due to their key roles in a variety of essential biologicalprocesses abundances of p63 proteins are tightly controlledUbiquitin-dependent proteasomal degradation is the mostimportant way to eliminate cellular p63 proteins Someimportant regulators including ubiquitin E3 ligases kinasesand proteins in other classes have been reported to controlp63 degradation Multiple extracellular signalings such asgrowth factor signaling and genotoxic stress impact theseregulators which in turn modulate protein stability of p63[9 10] This review is aimed at understanding the molecularmechanisms by which p63 protein stability is regulated andthe players in modulating ubiquitin-dependent proteasomaldegradation of p63 proteins

2 Isoforms of p63 and TheirBiological Functions

The p63 gene consists of 15 exons that can be transcribedfrom two transcriptional start sites The transcript from 51015840promoter of p63 gene proceeding to the first exon encodes TAisotypes of p63 proteins with the full transactivation domain(TAD) homologues to that of p53 on the N-terminus whiletranscript from the cryptic 31015840 intronic promoter gives riseto ΔN isoforms containing a different and weaker domaincapable of transactivation Both TA and ΔN isotypes canundergo alternative splicing to generate different carboxy-termini including 120572 120573 120574 120575 and 120576Thus p63 gene can encodeat least 10 different p63 isoforms TA(120572 sim 120576) and ΔN(120572 sim 120576)(as depicted in Figure 1) Each p63 isoform possesses a DNA-binding domain (DBD) and an oligomerization domain(OLD) both of which are highly homologous to those ofp53 protein Among these p63 isoforms p63120572 contains a full-length C-terminus consisting of a sterile alpha motif (SAM)for protein interaction and a transinhibitory domain (TID)whereas other isoforms have truncated C-termini due toalternative splicing [11ndash13] Evidence from human geneticsand animal models reveals that p63 proteins play crucialroles in stratification of squamous epithelia differentiation ofmature keratinocytes and epidermal morphogenesis during

Hindawi Publishing CorporationBioMed Research InternationalVolume 2014 Article ID 175721 8 pageshttpdxdoiorg1011552014175721

2 BioMed Research International

1 98 182 360 393 432 505 579 680

1 98 182 360 393 432 505 555

1 98 182 360 393 432 487

1 98 182 360 393 432 510

1

1 15 88 266 299 338 411 485 586

1 15 88 266 299 338 411 461

1 15 88 266 299 338 393

1 15 88 266 299 338 416

1 15 88 266 299 338356

98 182 360 393 432 450

ΔNp63120572

ΔNp63120573

ΔNp63120574

ΔNp63120575

ΔNp63120576

TAp63120572

TAp63120573

TAp63120574

TAp63120575

TAp63120576

TAD

TAD

TAD

TAD

TAD

DBD

DBD

DBD

DBD

DBD

DBD

DBD

DBD

DBD

DBD

OLD

OLD

OLD

OLD

OLD

OLD

OLD

OLD

OLD

OLD

SAM

SAM TID

TID

Figure 1 Schematic presentation of p63 isoforms TAD transactivation domain DBDDNA-binding domain OLD oligomerization domainSAM sterile alpha motif TID transinhibitory domain

development [14 15] Multiple p63 isotypes (both TA andΔN isoforms) are expressed in keratinocytes and they aredifferentially modulated during differentiation [16 17]

Endogenous TAp63 proteins are barely detectable inembryos and adult (except in oocytes) presumably becauseof their low expression or rapid degradation as well as lackof antibodies able to detect weak expression [18] It is wellsupported that like p53 TAp63 proteins promote cell cyclearrest and apoptotic cell death via activating proapoptotictargets such as Puma Bax and Noxa in somatic cells [119 20] In oocytes TAp63120572 expresses at relatively higherlevels and functions as a quality control factor in the femalegermline upon genotoxic stress [21ndash23] TAp63 knockoutmice are highly tumor prone and develop metastatic diseasereaffirming the antitumor activities of TAp63 [5 6] Loss ofTAp63 also results in premature aging and reduced lifespaninmice [5 24] Recently increased obesity insulin resistanceand glucose intolerance were reported in TAp63-null mice[25]ΔNp63 especially ΔNp63120572 are predominant p63 iso-

forms which are overexpressed in a highly specificmanner inthe embryonic ectoderm and the basal regenerative compart-ment of epithelial tissues such as skin teeth and hair [26]Similar to mice deleted with all p63 isoforms ΔNp63-nullmice have striking developmental defects including truncatedforelimbs the absence of hind limbs and a lack of stratified

epidermis [27] In contrast to the strong transactivationfunction of TAp63 proteins ΔNp63 isoforms were tradition-ally believed to inhibit p53 members including p53 TAp63and TAp73 proteins via forming complexes with them orcompetitive binding to p53-responsive elements This tran-scriptional repressor activity enablesΔNp63120572 to promote cellproliferation and tumorigenesis under certain circumstances[4 11 19 28 29] According to this model the fine balancebetween the TA and ΔN isotypes determines the functionof p63 proteins However mounting evidence reveals thatΔNp63 has an intrinsic transcriptional activity owing to asecond TA domain (TA2) ΔNp63120572 has been shown to regu-late the expression of several adhesion molecules includingintegrins (1205731 1205734 and 1205726) and desmosome protein PERPas well as MAP kinase phosphatase 3 (MKP3) heat shockprotein 70 (HSP70) multidrug resistant gene 1 (MDR-1)and ATM kinase implicating its functions in cell growthinvasion survival drug resistance andDNA repair [8 10 30ndash32]

3 Properties of p63 Protein Stability

Due to their potent proapoptotic activities TAp63 proteinsare generally expressed at very low levels likely owing to thetranscriptional regulation Alternatively TAp63 proteins arehighly labile It has been documented that the half-life of

BioMed Research International 3

TAp63120574 is about 15 hours and anunknown factormay play asa feedback regulator of TAp63 degradation ΔNp63 proteinsare much more stable than TAp63 [33] They are found over-expressed in keratinocyte and squamous carcinoma cells andassociated with proliferation It has been shown that whileΔNp63 undergoes degradation [34] TAp63 accumulates inresponse to some extrinsic stresses such as actinomycin Dbleomycin etoposide and UV irradiation [10] Some intrinsicphysiological processes such as cell differentiation are alsolikely to regulate degradation of p63 proteins [16 35 36]

Although lysosomal degradation may be involved inregulation of p63 abundance the most important way of p63degradation is the proteasome-dependent pathway [10] Thestabilities of p63 proteins are modulated by diverse post-translational modification such as phosphorylation ubiq-uitylation SUMOylation and ISGylation in which variousproteins are involved [26 37ndash39]

4 E3 Ligases Targeting p63 forProteasomal-Mediated Degradation

As the primary pathway of p63 protein degradation ubiq-uitin-dependent proteasomal degradation of p63 proteinswas reported by several laboratories E3 ligase-mediatedubiquitylation is the essential step for proteasomal degrada-tion of a specific protein Up to now several E3 ligases for p63proteins have been identified (as listed in Table 1)

41 Nedd4 Nedd4 is the first identified E3 ligase for p63Using a yeast-two-hybrid screening system Bakkers etal found that two modifying enzymes the E3 ubiquitinligase Nedd4 and the SUMO-conjugating enzyme Ubc9bind to distinct sites in the unique C-terminal region ofΔNp63120572 These physical interactions lead to ubiquitylationand SUMOylation of ΔNp63120572 resulting in vulnerabilityof ΔNp63120572 to proteasomal degradation [36] In zebrafishembryos ΔNp63120572 are expressed at a high level on thedorsal side due to the restricted expression of ubc91and nedd4 in this region [36] However how does theUbc9-mediated SUMOylation of Lys582 destabilize ΔNp63120572remains unknown In addition whether TAp63120572 whichshares the same C-terminus with ΔNp63120572 is modulated byNedd4 and Ubc9 remains to be investigated It was recentlyreported that ΔNp63120572 can transcriptionally repress Nedd4[40] suggesting that downregulation of Nedd4 may functionas a feedforward pathway to increase ΔNp63120572 intracellularconcentration under certain circumstances

42 Itch Itch is Nedd4-like ubiquitin E3 ligase found totarget p63 for ubiquitin-mediated proteasomal degradationWork from Melinorsquos group found that Itch directly binds tothe PPPY motif existing in the SAM domain of p63120572 Thisphysical interaction leads to ubiquitylation of either TA- orΔN-p63120572 isoforms consequently promoting their proteaso-mal degradation A Yrarr F substitution in the PPPY motifcan abolish the binding of Itch and significantly increasesp63120572 protein stability Their data suggest that Itch playsa fundamental role in controlling endogenous p63 protein

levels and regulating p63-mediated physiological functionsparticularly in the epidermis and keratinocytes [26] Thisis similar to the case of p73 which is also ubiquitylatedand targeted to degradation by Itch through PPPY-Itchinteraction [41] And this is also consistent with cases of p63degradation mediated by Nedd4 and WWP1 which both areanalogues of Itch and bind to PPPY motif of p63 [36 42]

Nevertheless Calabro group found that Itch bound to adifferent region of p63 the region encompassing aminoacids109 to 120 of TAp63 (corresponding to amino acids 15 to 26of ΔNp63120572) This association between Itch and N-termini ofp63 promotes degradation of all p63 isoforms including bothTA- and ΔN-isotypes [37]

43 WWP1 Li et al found that WWP1 the homologue ofItch can also bind to the PPPY motif of either TAp63120572 orΔNp63120572 and ubiquitinate them in culturedmammalian cellsconsequently promoting their proteasomal degradationAdditionallyWWP1 can target endogenousΔNp63120572proteinsfor degradation and sensitizes immortalized breast epithelialcells to chemotherapeutic drug doxorubicin-induced apop-tosis Intriguingly WWP1 can be upregulated at both mRNAand protein levels upon chemotherapeutic drug treatment ina p53-dependent manner [42] This suggests that ΔNp63120572which is well documented to confer cells to resistance toDNAdamage agent-mediated apoptosis [43] may be destabilizedby accumulated WWP1 E3 ligase resulting in cell deathunder genotoxic stress

44 Fbw7 Although several groups reported that MDM2which is a nuclear E3 ubiquitin ligase playing key rolesin controlling cellular p53 abundance cannot individuallymediate ubiquitylation of p63 [44ndash46] Galli et al foundthat MDM2 can facilitate ubiquitylation of p63 mediated byanother E3 ligase Fbw7 Upon DNA damage or keratinocytesdifferentiation MDM2 binds to the C-terminal SAM regionof ΔNp63120572 in the nucleus and promotes its translocation tothe cytoplasm then p63 is targeted for degradation by theFbw7 E3 ubiquitin ligase in the cytosome In this processGSK3 kinase activity is required for efficient degradation ofp63 by Fbw7 [35]

45 Pirh2 Pirh2 (p53-induced RING-H2) is another E3ligase found to target p63 for degradation It was recentlyreported that Pirh2 physically interacts with TAp63 andΔNp63 and targets them for polyubiquitylation and subse-quently proteasomal degradation This Pirh2-mediated post-transcriptional regulation of p63maymodulate keratinocytesdifferentiation [47] Arsenic trioxide a frontline agent foracute promyelocytic leukemia stimulates Pirh2 expressionand consequently promotes proteasomal degradation ofΔNp63120572 but not TAp63120572 [48] This is interesting and usefulbecause ΔNp63120572 is generally considered as a potent onco-protein playing key roles in tumor cell proliferationsurvivalwhereas TAp63120572 is a tumor suppressor with proapoptoticactivity It remains obscure why these two isoforms whichcan both be targeted for degradation by Pirh2 have thisdiscrepancy in response to arsenic trioxide treatment


Table 1 Regulators of p63 protein stability + positively regulates p63 protein stability minus negatively regulates p63 protein stability lowast MDM2functions as not an E3 ligase here but a helper to promote translocation of ΔNp63120572 to cytoplasm [35]

Regulators Stability regulation p63 isoform Interaction or modification sites ReferencesE3 ligases

Nedd4 minus ΔNp63120572 PPPY motif in SAM domain [36]

Itchminus ΔNp63120572 PPPY motif in SAM domain [26]

minus All p63 isoforms A region encompassing aminoacids 109 to 120 ofTAp63 (aminoacids 15 to 26 of ΔNp63) [37]

WWP1 minus p63120572 PPPY motif in SAM domain [42]Fbw7 minus ΔNp63120572 Likely a region encompassing S383 of ΔNp63120572 [35]Pirh2 minus TAp63 and ΔNp63 [47 48]

KinaseATM minus ΔNp63120572 S385 of ΔNp63120572 [49]CDK2 minus ΔNp63120572 T397 of ΔNp63120572 [49]p70s6K minus ΔNp63120572 S466 of ΔNp63120572 [49]

IKK120573 + TAp63120574 TAD of TAp63120574 [51]minus ΔNp63120572 Unknown [52]]

HIPK2 minus ΔNp63120572 T397 of TAp63120574 [55]

c-Abl + TAp63 Y149 Y171 and Y289 of TAp63 [22]minus ΔNp63120572 Y55 Y137 and Y308 of ΔNp63120572 [58]

Plk1 minus TAp63 S52 of TAp63 [59]p38 minus ΔNp63120572 Unknown [60]GSK3 minus ΔNp63120572 S383 of ΔNp63120572 [35]Raf1 minus ΔNp63120572 S383 or T397 of ΔNp63120572 [16]

OthersGadd45a minus ΔNp63120572 No direct interaction [60]p53 minus ΔNp63120572 DBD of ΔNp63120572 [29]MDM2lowast minus ΔNp63120572 SAM domain [35]Cabbles1 + TAp63120572 TAD and SAM of TAp63120572 [65]YAP + ΔNp63120572 PPPY motif in SAM domain [58]Dlx3 minus ΔNp63120572 No direct interaction [16]Stratifin minus p63120572 RQTISFP motif in TID of p63120572 [61]RACK1 minus p63120572 C-ter of p63120572 [61]Stxbp4 + ΔNp63120572 PPPY motif in SAM domain [64]Ubc9 minus ΔNp63120572 K582 of ΔNp63120572 [36]PML + p63120572 Unknown [66]Pin1 + p63120572 PPPY motif in SAM domain [67]

5 Kinases Involved in p63Protein Degradation

Phosphorylation mediated by diverse kinases plays key rolesin protein degradation So far several kinases have beenreported to be involved in p63 protein stability (as listed inTable 1)

51 ATM CDK2 and p70s6K Sidransky group found thatkinases including ATM CDK2 and p70s6K can phospho-rylate ΔNp63120572 in HNSCC (head and neck squamous cellcarcinoma) cells upon DNA damage Phosphorylation atS385 T397 and S466 mediated by these kinases respectively

promotes degradation of ΔNp63120572 [49] Given that ΔNp63120572can transcriptionally regulate ATM [50] the latter mayfunction as a feedback regulator via a p53MDM2-like loopto tightly control protein level of ΔNp63120572

52 IKK120573 TAp63120574 exhibits potent proapoptotic activity dueto possessing a full-length transactivation domain and lack-ing a transinhibitory domain Normally TAp63120574 expressesat very low levels in cells It was reported that in responseto 120574 radiation or tumor necrosis factor-120572 (TNF-120572) I120581Bkinase120573 (IKK120573) is activated and phosphorylates TAp63120574 (butnot ΔNp63120574) at its N-terminus this phosphorylation cansignificantly block ubiquitylation and possible degradation


of TAp63120574 in H1299 and HEK293 cells resulting in elevatedprotein levels of cellular TAp63120574 [33 51]

On the contrary Sidransky group found that cytokine- orchemotherapy-induced stimulation of IKK120573 kinase promotesubiquitin-mediated proteasomal degradation of ΔNp63120572 inthe human head and neck cancer cell line JHU-022 con-sequently augmenting transactivation of p53 family-inducedgenes involved in the cellular response to DNA damage [52]It is unclear how IKK120573 kinase exhibits opposite effects onstabilities of these two different isoforms An interpretationis that ΔNp63120572 and TAp63120574 possess different C-terminus orN-terminus

53 HIPK2 Homeodomain-interacting protein kinase 2(HIPK2) is an evolutionarily conserved serinethreoninekinase involved in the regulation of gene transcription duringdevelopment and in cell response to several types of stressHIPK2 is often activated under diverse genotoxic stimuliincluding treatmentwith ultraviolet ionizing irradiation andanticancer drugs such as cisplatin doxorubicin and roscovi-tine [53 54] Lazzari et al found that HIPK2 phosphorylatesΔNp63120572 at T397 residue in response to chemotherapy Thismodification promotes proteasomal degradation of ΔNp63120572which is mediated by neither Itch nor MDM2 [55]

54 c-Abl c-Abl (also known as ABL1) is a tyrosine kinasewhich can be activated upon DNA damage [56 57] Gonfloniet al recently reported that in response to cisplatin treatmentc-Abl phosphorylates TAp63120572 on Y149 Y171 and Y289residues resulting in increased protein stability of TAp63120572This can lead to accumulated TAp63120572 proteins and eventuallycell death in mouse oocytes [22]

Another group found that c-Abl phosphorylates Y55Y137 andY308 residues ofΔNp63120572 and stabilizes it via induc-ing its binding to Yes-associated protein (YAP) consequentlyregulating cell viability in head and neck cancer cells [58]Since Y149 Y171 and Y289 residues of TAp63120572 are differentfrom Y55 Y137 and Y308 of ΔNp63120572 it is interesting whyTAp63120572 and ΔNp63120572 are stabilized by c-Abl in differentmechanism It also remains unclear if other isoforms of p63can be regulated by c-Abl given that these six putative c-Ablphosphorylation sites are included in all of p63 isoforms

55 Other Kinases In addition to the abovementionedkinases Plk1 [59] p38 [60] GSK3 [35] and Raf1 [16] kinasesmay be also involved in proteasome-dependent degradationof p63

6 Other Proteins Regulating p63Protein Stability

Besides E3 ubiquitin ligases and kinases several other pro-teins were found to be related to protein stability of p63(as listed in Table 1) Most of them function as regulatorsor cofactors of the abovementioned E3 ligases or kinases topromote or inhibit proteasomal degradation of p63

61 p53 Ratovitski group found that p53 can bind to DNA-binding domain of p63 in the absence of DNA and promotep63 degradation through a pathway mediated by caspase-1Interestingly this p63 caspase-mediated degradation of p63resulting from p53-p63 physical interaction needs no furtherp63 posttranslational modifications [29]

62 Stratifin RACK1 and Stxbp4 In another work Ratovit-ski et al found that in response to DNA damage stratifinbinds to the putatively phosphorylated ΔNp63120572 protein andregulates its level through nuclear-cytoplasmic traffickingwhile scaffold protein RACK1 (receptor for activated Ckinase 1) subsequently targets ΔNp63120572 into a 26S-dependentproteasomal degradation pathwayThough overexpression ofRACK1 dramatically enhances ubiquitylation of ΔNp63120572 itneeds further confirmation if RACK1 serves as an E3 ligase inthis pathway [61]

Since it has been documented that RACK1 can regulateFAK activity [62] it is also possible that RACK1 promotesubiquitylation via affecting phosphorylation of p63 Notablyit has been reported that stratifin is transcriptionally regu-lated by p63 and p53 [28 63] So this modulation of p63mediated by stratifinRACK1may also forma regulation loop

Another scaffold protein Stxbp4 was found to counteractthis RACK1-dependent degradation pathway to maintainhigh basal levels of ΔNp63 in stratified epithelial cells undernormal growth conditions but in response to DNA damageStxbp4 is downregulated correlating with ΔNp63120572 destabi-lization mediated in part by RACK1 [64]

63 Other Proteins Hildesheim et al reported that undergenotoxic stress Gadd45a mediates activation of p38 MAPkinase and consequently downregulates ΔNp63120572 protein[60] Wang et al found that Cables1 (CDK5 and Abl enzymesubstrate 1) associates with both TAD and SAM regions ofTAp63120572 to protect TAp63120572 from proteasomal degradation[65] Bernassola et al found that the promyelocytic leukaemiaprotein (PML) physically interacts with TAp63120572 andΔNp63120572and increases their protein levels [66] According to resultsof Di Costanzo et al homeodomain protein Dlx3 physicallyinteractswith and activates Raf1 kinase resulting in enhancedphosphorylation and degradation of ΔNp63120572 [16] As men-tioned above YAP and Ubc9 are also involved in regulationof p63 degradation [36 58] Recently we found that peptidyl-prolyl isomerase Pin1 inhibits binding of WWP1 E3 ligase top63120572 consequently preventing its proteasomal degradation[67]

7 Summary and Prospects

Taken together current studies reveal that p63 protein stabil-ity is tightly controlled and closely correlated with cell prolif-eration cell death and cell differentiation Various signalingsand stresses can affect p63 protein stability via regulatingE3 ligases or activities of diverse kinases Notably the sameresidues or same regions are targeted by different E3 ligasesor different signaling For instance T397 of ΔNp63120572 can bephosphorylated by several kinases including CDK2 HIPK2


and Raf1 consequently promoting degradation of ΔNp63120572three E3 ligase homologues NEDD4 Itch andWWP1 targetp63 for proteasomal degradation via binding to PPPY motifof p63120572 and p63120573 Furthermore the regulation of p63 proteinstability is complex since the effects of E3 ligases on p63degradation are modulated by kinases and other regulatorsSome of the factors can be transcriptionally regulated byp63 forming some regulation loops Further investigationsare required to clarify the relationships between differentregulators in p63 protein stability upon exogenous signalingMoreover it is necessary to elucidate how different p63 iso-types are targeted to degradation during cell differentiationsince a switch in expression of different p63 isoforms is pivotalin this process [68]

On the other hand it would be important to investigatekey regulators for p63 degradation that are responsiblefor p63-related human developmental diseases Like theparadigm of promising anticancer drug Nutlin-3 which canstabilize p53 via disrupting its association with MDM2 [69]it would be interesting to develop potent inhibitors targetingE3 ligases in order to stabilize TAp63 tumor suppressors or todestabilize ΔNp63 oncoproteins

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported in part by the National NaturalScience Foundation of China (no 31171362 to Z Xiao and no31100982 to C Li) and Research Foundation for the DoctoralProgram Ministry of Education of China (no 2011018120082to C Li)

References

[1] MOsadaMOhba C Kawahara et al ldquoCloning and functionalanalysis of human p51 which structurally and functionallyresembles p53rdquoNatureMedicine vol 4 no 7 pp 839ndash843 1998

[2] B Trink K Okami L Wu V Sriuranpong J Jen and DSidransky ldquoA new human p53 homologuerdquo Nature Medicinevol 4 no 7 pp 747ndash748 1998

[3] K Hibi B Trink M Patturajan et al ldquoAIS is an oncogeneamplified in squamous cell carcinomardquo Proceedings of theNational Academy of Sciences of the United States of Americavol 97 no 10 pp 5462ndash5467 2000

[4] M Senoo F Pinto C P Crum and F McKeon ldquop63 is essentialfor the proliferative potential of stem cells in stratified epitheliardquoCell vol 129 no 3 pp 523ndash536 2007

[5] X Guo W M Keyes C Papazoglu et al ldquoTAp63 inducessenescence and suppresses tumorigenesis in vivordquo Nature cellbiology vol 11 no 12 pp 1451ndash1457 2009

[6] X Su D Chakravarti M S Cho et al ldquoTAp63 suppressesmetastasis through coordinate regulation of Dicer and miR-NAsrdquo Nature vol 467 no 7318 pp 986ndash990 2010

[7] W M Keyes M Pecoraro V Aranda et al ldquoΔnp63120572 is anoncogene that targets chromatin remodeler Lsh to drive skin

stem cell proliferation and tumorigenesisrdquo Cell Stem Cell vol 8no 2 pp 164ndash176 2011

[8] D K Carroll J S Carroll C-O Leong et al ldquop63 regulates anadhesion programme and cell survival in epithelial cellsrdquoNatureCell Biology vol 8 no 6 pp 551ndash561 2006

[9] CMaisse P Guerrieri andGMelino ldquop73 and p63 protein sta-bility theway to regulate functionrdquoBiochemical Pharmacologyvol 66 no 8 pp 1555ndash1561 2003

[10] L E Finlan and T R Hupp ldquop63 the phantom of the tumorsuppressorrdquo Cell Cycle vol 6 no 9 pp 1062ndash1071 2007

[11] A Yang M Kaghad Y Wang et al ldquop63 a p53 homolog at3q27-29 encodesmultiple products with transactivating death-inducing and dominant-negative activitiesrdquoMolecular Cell vol2 no 3 pp 305ndash316 1998

[12] F Murray-Zmijewski D P Lane and J-C Bourdon ldquop53p63p73 isoforms an orchestra of isoforms to harmonise celldifferentiation and response to stressrdquo Cell Death and Differen-tiation vol 13 no 6 pp 962ndash972 2006

[13] MMangiulli A Valletti M F Caratozzolo et al ldquoIdentificationand functional characterization of two new transcriptionalvariants of the human p63 generdquoNucleic Acids Research vol 37no 18 pp 6092ndash6104 2009

[14] H van Bokhoven B C J Hamel M Bamshad et al ldquop63 genemutations in EEC syndrome limb-mammary syndrome andisolated split hand-split foot malformation suggest a genotype-phenotype correlationrdquo American Journal of Human Geneticsvol 69 no 3 pp 481ndash492 2001

[15] A A Mills B Zheng X-J Wang H Vogel D R Roop andA Bradley ldquop63 is a p53 homologue required for limb andepidermal morphogenesisrdquo Nature vol 398 no 6729 pp 708ndash713 1999

[16] A Di Costanzo L Festa O Duverger et al ldquoHomeodomainproteinDlx3 induces phosphorylation-dependent p63 degrada-tionrdquo Cell Cycle vol 8 no 8 pp 1185ndash1195 2009

[17] K E King R M Ponnamperuma M J Gerdes et al ldquoUniquedomain functions of p63 isotypes that differentially regulatedistinct aspects of epidermal homeostasisrdquo Carcinogenesis vol27 no 1 pp 53ndash63 2006

[18] M Paris M Rouleau M Puceat and D Aberdam ldquoRegulationof skin aging and heart development by TAp63rdquo Cell Death andDifferentiation vol 19 no 2 pp 186ndash193 2012

[19] G Wu S Nomoto M O Hoque et al ldquoΔNp63120572 and TAp63120572regulate transcription of geneswith distinct biological functionsin cancer and developmentrdquoCancer Research vol 63 no 10 pp2351ndash2357 2003

[20] O Gressner T Schilling K Lorenz et al ldquoTAp63120572 inducesapoptosis by activating signaling via death receptors and mito-chondriardquo The EMBO Journal vol 24 no 13 pp 2458ndash24712005

[21] G B Deutsch E M Zielonka D Coutandin et al ldquoDNAdamage in oocytes induces a switch of the quality control factorTAp63120572 from dimer to tetramerrdquo Cell vol 144 no 4 pp 566ndash576 2011

[22] S Gonfloni L Di Tella S Caldarola et al ldquoInhibition of thec-Abl-TAp63 pathway protects mouse oocytes from chemo-therapy-induced deathrdquo Nature Medicine vol 15 no 10 pp1179ndash1185 2009

[23] G B Deutsch E M Zielonka D Coutandin and V DotschldquoQuality control in oocytes domain-domain interactions regu-late the activity of p63rdquo Cell Cycle vol 10 no 12 pp 1884ndash18852011


[24] X Su M Paris Y J Gi et al ldquoTAp63 prevents premature agingby promoting adult stem cell maintenancerdquo Cell Stem Cell vol5 no 1 pp 64ndash75 2009

[25] X Su Y J Gi D Chakravarti et al ldquoTAp63 is a mastertranscriptional regulator of lipid and glucose metabolismrdquo CellMetabolism vol 16 pp 511ndash525 2012

[26] M Rossi R I Aqeilan M Neale et al ldquoThe E3 ubiquitin ligaseItch controls the protein stability of p63rdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 103 no 34 pp 12753ndash12758 2006

[27] R-A Romano K Smalley C Magraw et al ldquoΔNp63 knockoutmice reveal its indispensable role as a master regulator ofepithelial development and differentiationrdquo Development vol139 no 4 pp 772ndash782 2012

[28] MDWestfall D JMays J C Sniezek and J A Pietenpol ldquoTheΔNp63120572 phosphoprotein binds the p21 and 14-3-3120590 promotersin vivo and has transcriptional repressor activity that is reducedby Hay-Wells syndrome-derived mutationsrdquo Molecular andCellular Biology vol 23 no 7 pp 2264ndash2276 2003

[29] E A Ratovitski M Patturajan K Hibi B Trink K Yamaguchiand D Sidransky ldquop53 associates with and targets ΔNp63 intoa protein degradation pathwayrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 98 no4 pp 1817ndash1822 2001

[30] J Bergholz Y Zhang J Wu et al ldquoΔNp63120572 regulates Erksignaling viaMKP3 to inhibit cancermetastasisrdquoOncogene vol33 no 2 pp 212ndash224 2012

[31] R A Ihrie M R Marques B T Nguyen et al ldquoPerp is a p63-regulated gene essential for epithelial integrityrdquoCell vol 120 no6 pp 843ndash856 2005

[32] G Wu M Osada Z Guo et al ldquoΔNp63120572 up-regulates theHsp70 gene in human cancerrdquo Cancer Research vol 65 no 3pp 758ndash766 2005

[33] H Ying D L F Chang H Zheng F McKeon and Z-X JXiao ldquoDNA-binding and transactivation activities are essentialfor TAp63 protein degradationrdquoMolecular and Cellular Biologyvol 25 no 14 pp 6154ndash6164 2005

[34] Y Okada M Osada S-I Kurata et al ldquop53 gene fam-ily p51(p63)-encoded secondary transactivator p51b(TAp63120572)occurs without forming an immunoprecipitable complex withMDM2 but responds to genotoxic stress by accumulationrdquoExperimental Cell Research vol 276 no 2 pp 194ndash200 2002

[35] F Galli M Rossi Y DrsquoAlessandra et al ldquoMDM2 and Fbw7cooperate to induce p63 protein degradation following DNAdamage and cell differentiationrdquo Journal of Cell Science vol 123no 14 pp 2423ndash2433 2010

[36] J Bakkers M Camacho-Carvajal M Nowak C Kramer BDanger and M Hammerschmidt ldquoDestabilization of ΔNp63120572byNedd4-mediated ubiquitination andUbc9-mediated sumoy-lation and its implications on dorsoventral patterning of thezebrafish embryordquo Cell Cycle vol 4 no 6 pp 790ndash800 2005

[37] M Rossi M De Simone A Pollice et al ldquoItchAIP4 associateswith and promotes p63 protein degradationrdquo Cell Cycle vol 5no 16 pp 1816ndash1822 2006

[38] Y J Jeon M G Jo H M Yoo et al ldquoChemosensitivityis controlled by p63 modification with ubiquitin-like proteinISG15rdquo The Journal of Clinical Investigation vol 122 pp 2622ndash2636 2012

[39] P Ghioni Y DrsquoAlessandra G Mansueto et al ldquoThe proteinstability and transcriptional activity of p63120572 are regulated bySUMO-1 conjugationrdquoCell Cycle vol 4 no 1 pp 183ndash190 2005

[40] M K Leonard N T Hill E D Grant and M P KadakialdquoDeltaNp63alpha represses nuclear translocation of PTEN byinhibition of NEDD4-1 in keratinocytesrdquo Archives of Dermato-logical Research vol 305 pp 733ndash739 2013

[41] M Rossi V De Laurenzi E Munarriz et al ldquoThe ubiquitin-protein ligase Itch regulates p73 stabilityrdquo The EMBO Journalvol 24 no 4 pp 836ndash848 2005

[42] Y Li Z Zhou and C Chen ldquoWW domain-containing E3ubiquitin protein ligase 1 targets p63 transcription factorfor ubiquitin-mediated proteasomal degradation and regulatesapoptosisrdquo Cell Death and Differentiation vol 15 no 12 pp1941ndash1951 2008

[43] X Li J Chen Y Yi C Li and Y Zhang ldquoDNA damage down-regulates ΔNp63120572 and induces apoptosis independent of wildtype p53rdquo Biochemical and Biophysical Research Communica-tions vol 423 no 2 pp 338ndash343 2012

[44] M Dobbelstein S Wienzek C Konig and J Roth ldquoInacti-vation of the p53-homologue p73 by the mdm2-oncoproteinrdquoOncogene vol 18 no 12 pp 2101ndash2106 1999

[45] X Wang T Arooz W Y Siu et al ldquoMDM2 and MDMX caninteract differently with ARF and members of the p53 familyrdquoFEBS Letters vol 490 no 3 pp 202ndash208 2001

[46] V Calabro G Mansueto T Parisi M Vivo R A Calogero andG L Mantia ldquoThe human MDM2 oncoprotein increases thetranscriptional activity and the protein level of the p53 homologp63rdquo Journal of Biological Chemistry vol 277 no 4 pp 2674ndash2681 2002

[47] Y S Jung Y Qian W Yan and X Chen ldquoPirh2 E3 ubiquitinligase modulates keratinocyte differentiation through p63rdquoJournal of InvestigativeDermatology vol 133 pp 1178ndash1187 2013

[48] W Yan X Chen Y Zhang J Zhang and Y S Jung ldquoArsenicsuppresses cell survival via Pirh2-mediated proteasomal degra-dation of DeltaNp63 proteinrdquo The Journal of Biological Chem-istry vol 288 pp 2907ndash2913 2013

[49] Y Huang T Sen J Nagpal et al ldquoATMkinase is amaster switchfor the ΔNp63120572 phosphorylationdegradation in human headand neck squamous cell carcinoma cells upon DNA damagerdquoCell Cycle vol 7 no 18 pp 2846ndash2855 2008

[50] A L Craig J Holcakova L E Finlan et al ldquoΔNp63 transcrip-tionally regulates ATM to control p53 Serine-15 phosphoryla-tionrdquoMolecular Cancer vol 9 article 195 2010

[51] M MacPartlin S X Zeng and H Lu ldquoPhosphorylation andstabilization of TAp63120574 by I120581B kinase-120573rdquo Journal of BiologicalChemistry vol 283 no 23 pp 15754ndash15761 2008

[52] A Chatterjee X Chang T Sen R Ravi A Bedi and DSidransky ldquoRegulation of p53 family member isoform ΔNp63120572by the nuclear factor-120581B targeting kinase I120581B kinase 120573rdquo CancerResearch vol 70 no 4 pp 1419ndash1429 2010

[53] G DrsquoOrazi B Cecchinelli T Bruno et al ldquoHomeodomain-interacting protein kinase-2 phosphorylates p53 at Ser 46 andmediates apoptosisrdquo Nature Cell Biology vol 4 no 1 pp 11ndash202002

[54] T G Hofmann A Moller H Sirma et al ldquoRegulation ofp53 activity by its interaction with homeodomain-interactingprotein kinase-2rdquoNatureCell Biology vol 4 no 1 pp 1ndash10 2002

[55] C Lazzari A Prodosmo F Siepi et al ldquoHIPK2 phosphorylatesΔnp63120572 and promotes its degradation in response to DNAdamagerdquo Oncogene vol 30 no 48 pp 4802ndash4813 2011

[56] M Simonatto F Marullo F Chiacchiera et al ldquoDNA damage-activated ABL-MyoD signaling contributes to DNA repair inskeletal myoblastsrdquo Cell Death amp Disease vol 20 no 12 pp1664ndash1674 2013


[57] N Reuven J Adler V Meltser and Y Shaul ldquoThe Hippopathway kinase Lats2 prevents DNAdamage-induced apoptosisthrough inhibition of the tyrosine kinase c-Ablrdquo Cell Death ampDisease vol 20 pp 1330ndash1340 2013

[58] MYuan P Luong CHudson KGudmundsdottir and S BasuldquoC-Abl phosphorylation of Δnp63120572 is critical for cell viabilityrdquoCell Death and Disease vol 1 no 1 article e16 2010

[59] S Komatsu H Takenobu T Ozaki et al ldquoPlk1 regulates livertumor cell death by phosphorylation of TAp63rdquo Oncogene vol28 no 41 pp 3631ndash3641 2009

[60] J Hildesheim G I Belova S D Tyner X Zhou L Vardanianand A J Fornace Jr ldquoGadd45a regulates matrix metallopro-teinases by suppressing ΔNp63120572 and 120573-catenin via p38 MAPkinase and APC complex activationrdquo Oncogene vol 23 no 10pp 1829ndash1837 2004

[61] A Fomenkov R Zangen Y-P Huang et al ldquoRACK1 andstratifin target ΔNp63120572 for a proteasome degradation in headand neck squamous cell carcinoma cells upon DNA damagerdquoCell Cycle vol 3 no 10 pp 1285ndash1295 2004

[62] B Serrels E Sandilands A Serrels et al ldquoA complex betweenFAK RACK1 and PDE4D5 controls spreading initiation andcancer cell polarityrdquo Current Biology vol 20 no 12 pp 1086ndash1092 2010

[63] M Dohn S Zhang and X Chen ldquop63120572 and ΔNp63120572 caninduce cell cycle arrest and apoptosis and differentially regulatep53 target genesrdquoOncogene vol 20 no 25 pp 3193ndash3205 2001

[64] Y Li M J Peart and C Prives ldquoStxbp4 regulates ΔNp63stability by suppression of RACK1-dependent degradationrdquoMolecular and Cellular Biology vol 29 no 14 pp 3953ndash39632009

[65] N Wang L Guo B R Rueda and J L Tilly ldquoCables1 protectsp63 from proteasomal degradation to ensure deletion of cellsafter genotoxic stressrdquoEMBOReports vol 11 no 8 pp 633ndash6392010

[66] F Bernassola A Oberst G Melino and P P Pandolfi ldquoThepromyelocytic leukaemia protein tumour suppressor functionsas a transcriptional regulator of p63rdquo Oncogene vol 24 no 46pp 6982ndash6986 2005

[67] C Li D L Chang Z Yang et al ldquoPin1 modulates p63alphaprotein stability in regulation of cell survival proliferation andtumor formationrdquo Cell Death amp Disease vol 4 p e943 2013

[68] K Nylander B Vojtesek R Nenutil et al ldquoDifferential expres-sion of p63 isoforms in normal tissues and neoplastic cellsrdquoJournal of Pathology vol 198 no 4 pp 417ndash427 2002

[69] MMichaelis F Rothweiler S Barth et al ldquoAdaptation of cancercells from different entities to the MDM2 inhibitor nutlin-3results in the emergence of p53-mutated multi-drug-resistantcancer cellsrdquo Cell Death and Disease vol 2 no 12 article e2432011

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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International Journal of

Volume 2014

Zoology


Molecular Biology International

GenomicsInternational Journal of


The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014


BioinformaticsAdvances in

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Signal TransductionJournal of


BioMed Research International

Evolutionary BiologyInternational Journal of



Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014


Genetics Research International


Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational



Enzyme Research



Microbiology


1 98 182 360 393 432 505 579 680

1 98 182 360 393 432 505 555

1 98 182 360 393 432 487

1 98 182 360 393 432 510

1

1 15 88 266 299 338 411 485 586

1 15 88 266 299 338 411 461

1 15 88 266 299 338 393

1 15 88 266 299 338 416

1 15 88 266 299 338356

98 182 360 393 432 450

ΔNp63120572

ΔNp63120573

ΔNp63120574

ΔNp63120575

ΔNp63120576

TAp63120572

TAp63120573

TAp63120574

TAp63120575

TAp63120576

TAD

TAD

TAD

TAD

TAD

DBD

DBD

DBD

DBD

DBD

DBD

DBD

DBD

DBD

DBD

OLD

OLD

OLD

OLD

OLD

OLD

OLD

OLD

OLD

OLD

SAM

SAM TID

TID

Figure 1 Schematic presentation of p63 isoforms TAD transactivation domain DBDDNA-binding domain OLD oligomerization domainSAM sterile alpha motif TID transinhibitory domain

development [14 15] Multiple p63 isotypes (both TA andΔN isoforms) are expressed in keratinocytes and they aredifferentially modulated during differentiation [16 17]

Endogenous TAp63 proteins are barely detectable inembryos and adult (except in oocytes) presumably becauseof their low expression or rapid degradation as well as lackof antibodies able to detect weak expression [18] It is wellsupported that like p53 TAp63 proteins promote cell cyclearrest and apoptotic cell death via activating proapoptotictargets such as Puma Bax and Noxa in somatic cells [119 20] In oocytes TAp63120572 expresses at relatively higherlevels and functions as a quality control factor in the femalegermline upon genotoxic stress [21ndash23] TAp63 knockoutmice are highly tumor prone and develop metastatic diseasereaffirming the antitumor activities of TAp63 [5 6] Loss ofTAp63 also results in premature aging and reduced lifespaninmice [5 24] Recently increased obesity insulin resistanceand glucose intolerance were reported in TAp63-null mice[25]ΔNp63 especially ΔNp63120572 are predominant p63 iso-

forms which are overexpressed in a highly specificmanner inthe embryonic ectoderm and the basal regenerative compart-ment of epithelial tissues such as skin teeth and hair [26]Similar to mice deleted with all p63 isoforms ΔNp63-nullmice have striking developmental defects including truncatedforelimbs the absence of hind limbs and a lack of stratified

epidermis [27] In contrast to the strong transactivationfunction of TAp63 proteins ΔNp63 isoforms were tradition-ally believed to inhibit p53 members including p53 TAp63and TAp73 proteins via forming complexes with them orcompetitive binding to p53-responsive elements This tran-scriptional repressor activity enablesΔNp63120572 to promote cellproliferation and tumorigenesis under certain circumstances[4 11 19 28 29] According to this model the fine balancebetween the TA and ΔN isotypes determines the functionof p63 proteins However mounting evidence reveals thatΔNp63 has an intrinsic transcriptional activity owing to asecond TA domain (TA2) ΔNp63120572 has been shown to regu-late the expression of several adhesion molecules includingintegrins (1205731 1205734 and 1205726) and desmosome protein PERPas well as MAP kinase phosphatase 3 (MKP3) heat shockprotein 70 (HSP70) multidrug resistant gene 1 (MDR-1)and ATM kinase implicating its functions in cell growthinvasion survival drug resistance andDNA repair [8 10 30ndash32]

3 Properties of p63 Protein Stability

Due to their potent proapoptotic activities TAp63 proteinsare generally expressed at very low levels likely owing to thetranscriptional regulation Alternatively TAp63 proteins arehighly labile It has been documented that the half-life of




















































Acknowledgments


References
















































































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology




















































Acknowledgments


References
















































































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology
























































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology

Review Article Regulation of p63 Protein Stability via...

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