Recognition mechanism of p63 by the E3 ligase Itch

Bellomaria, Alessia; Barbato, Gaetano; Melino, Gerry; Paci, Maurizio; Melino, Sonia

doi:10.4161/cc.21918

Cited by 44 publications

(21 citation statements)

References 61 publications

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“…108,109 Thus, altogether these results may be explained with an increase of the trans proline isomers of Ppep63 catalyzed by the Pin1 isomerase during the pre-incubation. The Arg-373 residue of Itch E3-ligase is involved in the interaction with pep63 97 and as previously observed it goes from fast to slow exchange regime when interacts with pep63 and pep63 cyclic form, respectively. This evidence seems to be in agreement with a trans proline conformer of the cyclic form and explain the absence of K d variation between cPpep63 and both the WW domains after pre-incubation with Pin1.…”

Section: Effect Of the Phosphorylation And The Cis/trans Isomerizatiomentioning

confidence: 64%

“…[94][95][96][97] In order to understand the biochemical properties of the interaction between these 2 proteins, the interaction between the Itch-WW2 and the synthetic peptide pep63, including the PPxY recognition motif, has been monitored using spectroscopic techniques; the apparent dissociation constant value of the Itch-WW2-pep63 complex has been reported previously. 96 The presence of the consensus TP motif, close to the PPxY recognition motif, and also the recent studies on the Pin1/p63 interaction 98,99 led us to evaluate the effect of the threonine phosphorylation of the (T/S)PPPxY motif on the binding of the Itch-WW domains to the pep63 peptide.…”

Section: Resultsmentioning

confidence: 99%

“…Phosphorylation of the TPPPxY recognition motif of the p63 protein facilitates the Itch-p63 recognition promoting the ubiquitylation p63. 94,96,97,100,117 This recognition can be modulate by the Pin1 that interacting with the (T/S) p P motif catalyzes the proline isomerization from trans to cis configuration of the recognition motif in p63 and decreases the Itchp63 interaction, resulting in a stabilization of the protein steady state level. Accordingly, the sequence of events is (i) phosphorylation by a kinase, (ii) recognition by Itch, poly-ubiquitylation and proteasomal degradation (right path), (iii) alternatively, Pin1 recognition and trans-cis prolyl isomerization of the proline, reducing interaction with Itch, hence stabilizing the steady-state protein levels of p63 (left path).…”

Section: Discussionmentioning

confidence: 99%

“…5A). 97 The intrinsic fluorescence changes of both WW1 and WW2 domains of the Itch protein were investigated by addition of the phosphorylated form of the cyclic pep63, in order to evaluate the effects of the phosphorylation on the molecular recognition and stabilization of the ItchWW-cPpep63 complex. Figure 5B shows the DFs of the maximal values of the intrinsic fluorescence of both Itch-WW1 and -WW2 at increased concentrations of the cPpep63.…”

Section: Effect Of the Phosphorylation And The Cis/trans Isomerizatiomentioning

confidence: 99%

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p63 threonine phosphorylation signals the interaction with the WW domain of the E3 ligase Itch

et al. 2014

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Both in epithelial development as well as in epithelial cancers, the p53 family member p63 plays a crucial role acting as a master transcriptional regulator. P63 steady state protein levels are regulated by the E3 ubiquitin ligase Itch, via a physical interaction between the PPxY consensus sequence (PY motif) of p63 and one of the 4 WW domains of Itch; this substrate recognition process leads to protein-ubiquitylation and p63 proteasomal degradation. The interaction of the WW domains, a highly compact protein-protein binding module, with the short proline-rich sequences is therefore a crucial regulatory event that may offer innovative potential therapeutic opportunity. Previous molecular studies on the Itch-p63 recognition have been performed in vitro using the Itch-WW2 domain and the peptide interacting fragment of p63 (pep63), which includes the PY motif. Itch-WW2-pep63 interaction is also stabilized in vitro by the conformational constriction of the S-S cyclization in the p63 peptide. The PY motif of p63, as also for other proteins, is characterized by the nearby presence of a (T/S)P motif, which is a potential recognition site of the WW domain of the IV group present in the prolyl-isomerase Pin1. In this study, we demonstrate, by in silico and spectroscopical studies using both the linear pep63 and its cyclic form, that the threonine phosphorylation of the (T/S)PPPxY motif may represent a crucial regulatory event of the Itch-mediated p63 ubiquitylation, increasing the Itch-WW domains-p63 recognition event and stabilizing in vivo the Itch-WW-p63 complex. Moreover, our studies confirm that the subsequently trans/cis proline isomerization of (T/S)P motif by the Pin1 prolyl-isomerase, could modulate the E3-ligase interaction, and that the (T/S) p P trans PPxY motif represent the best conformer for the ItchWW-(T/S)PPPxY motif recognition.

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Section: Effect Of the Phosphorylation And The Cis/trans Isomerizatiomentioning

confidence: 64%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Effect Of the Phosphorylation And The Cis/trans Isomerizatiomentioning

confidence: 99%

See 2 more Smart Citations

p63 threonine phosphorylation signals the interaction with the WW domain of the E3 ligase Itch

et al. 2014

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“…[121][122][123][124][125][126] While the DBD domain is highly conserved among vertebrates and invertebrates, the C terminus varies, resulting in a change from dimeric structure to a tetramer in the vertebrates. [127][128][129] The more ancient members of the family include p73, involved in cancer, 130 neurodevelopment, 131,132 and aging, 133 and p63, involved in epidermal development, 119,[134][135][136] cancer, [137][138][139][140][141] reproduction, 142 and heart development. 143 Understanding the structural restrain of its structure is pivotal to understand the function of p53 [144][145][146] as well as its potential therapeutic exploitation.…”

Section: Discussionmentioning

confidence: 99%

Molecular dynamics of the full-length p53 monomer

Chillemi¹,

Davidovich

D’Abramo³

et al. 2013

Cell Cycle

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The p53 protein is frequently mutated in a very large proportion of human tumors, where it seems to acquire gain-of-function activity that facilitates tumor onset and progression. A possible mechanism is the ability of mutant p53 proteins to physically interact with other proteins, including members of the same family, namely p63 and p73, inactivating their function. Assuming that this interaction might occurs at the level of the monomer, to investigate the molecular basis for this interaction, here, we sample the structural flexibility of the wild-type p53 monomeric protein. The results show a strong stability up to 850 ns in the DNA binding domain, with major flexibility in the N-terminal transactivations domains (TAD1 and TAD2) as well as in the C-terminal region (tetramerization domain). Several stable hydrogen bonds have been detected between N-terminal or C-terminal and DNA binding domain, and also between N-terminal and C-terminal. Essential dynamics analysis highlights strongly correlated movements involving TAD1 and the proline-rich region in the N-terminal domain, the tetramerization region in the C-terminal domain; Lys120 in the DNA binding region. The herein presented model is a starting point for further investigation of the whole protein tetramer as well as of its mutants

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Cullin3/KCTD5 induces monoubiquitination of ΔNp63α and impairs its activity

Peng

Fan

et al. 2018

FEBS Letters

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Potassium channel tetramerization domain containing 5 (KCTD5) was previously documented as a component of the Cullin3-RING ligase (CRL3). It has been reported that KCTD5 can induce enrichment of polyubiquitinated proteins, and KCTD5-based CRL3 destabilizes several proteins. In our present study, we report that KCTD5 may physically interact with ΔNp63α, which is a member of the p53 family. Our further investigation revealed that Cullin3/KCTD5 can induce monoubiquitination of ΔNp63α. Cullin3/KCTD5 downregulates the DNA-binding affinity of ΔNp63α, impairing either its transactivity or its transinhibitory activity. Functionally, Cullin3/KCTD5 abates the proproliferation activity of ΔNp63α. These findings suggest that KCTD5-based CRL3 may mediate monoubiquitination and is a novel regulator of ΔNp63α.

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Recognition mechanism of p63 by the E3 ligase Itch

Cited by 44 publications

References 61 publications

p63 threonine phosphorylation signals the interaction with the WW domain of the E3 ligase Itch

p63 threonine phosphorylation signals the interaction with the WW domain of the E3 ligase Itch

Molecular dynamics of the full-length p53 monomer

Cullin3/KCTD5 induces monoubiquitination of ΔNp63α and impairs its activity

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