Tankyrase Requires SAM Domain-Dependent Polymerization to Support Wnt-β-Catenin Signaling

Mariotti, Laura; Templeton, Catherine M.; Ranes, Michael; Paracuellos, Patricia; Cronin, Nora; Beuron, Fabienne; Morris, Edward P.; Guettler, Sebastian

doi:10.1016/j.molcel.2016.06.019

Cited by 80 publications

(167 citation statements)

References 67 publications

(104 reference statements)

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“…(A)]. Immediately upstream of this catalytic domain lies a SAM domain responsible for TNKSs polymerization . The extreme N‐terminal end harbors a histidine, serine, proline‐rich segment (a region lacking in tankyrase‐2), followed by a large ankyrin repeat domain composed of five conserved subdomains or ankyrin repeat clusters (ARCs) connected by conserved linkers.…”

Section: Introductionmentioning

confidence: 99%

Structural basis for tankyrase‐RNF146 interaction reveals noncanonical tankyrase‐binding motifs

DaRosa¹,

Klevit²,

Xu³

2018

Protein Science

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Poly(ADP-ribosyl)ation (PARylation) catalyzed by the tankyrase enzymes (Tankyrase-1 and -2; a.k.a. PARP-5a and -5b) is involved in mitosis, telomere length regulation, GLUT-4 vesicle transport, and cell growth and differentiation. Together with the E3 ubiquitin ligase RNF146 (a.k.a. Iduna), tankyrases regulate the cellular levels of several important proteins including Axin, 3BP2, and angiomotins, which are key regulators of Wnt, Src and Hippo signaling, respectively. These tankyrase substrates are first PARylated and then ubiquitylated by RNF146, which is allosterically activated by binding to PAR polymer. Each tankyrase substrate is recognized by a tankyrase-binding motif (TBM). Here we show that RNF146 binds directly to tankyrases via motifs in its C-terminal region. Four of these RNF146 motifs represent novel, extended TBMs, that have one or two additional amino acids between the most conserved Arg and Gly residues. The individual RNF146 motifs display weak binding, but together mediate a strong multivalent interaction with the substrate-binding region of TNKS, forming a robust one-to-one complex. A crystal structure of the first RNF146 noncanonical TBM in complex with the second ankyrin repeat domain of TNKS shows how an extended motif can be accommodated in a peptide-binding groove on tankyrases. Overall, our work demonstrates the existence of a new class of extended TBMs that exist in previously uncharacterized tankyrase-binding proteins including those of IF4A1 and NELFE.

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Section: Introductionmentioning

confidence: 99%

Structural basis for tankyrase‐RNF146 interaction reveals noncanonical tankyrase‐binding motifs

DaRosa¹,

Klevit²,

Xu³

2018

Protein Science

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“…Since the stabilization of Axin can lead to a concomitant reduction in β-catenin protein levels, which blocks its transcriptional activity in vitro and in vivo, inhibition of tankyrases has been proposed as an exciting new cancer therapeutic approach to block Wnt signaling (Huang et al 2009;Waaler et al 2011Waaler et al , 2012. However, catalytic inhibition of tankyrases might not be sufficient, as polymers of tankyrases were discovered to act as scaffolds to promote Wnt signaling independently of their PARP activity (Mariotti et al 2016). Thus, inhibiting the catalytic activity of tankyrases per se may not be adequate for suppressing Wnt activity in cells with high levels of tankyrases.…”

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confidence: 99%

USP25 regulates Wnt signaling by controlling the stability of tankyrases

Liu

et al. 2017

Genes Dev.

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Aberrant activation of the Wnt signaling pathway plays an important role in human cancer development. Wnt signaling is negatively regulated by Axin, a scaffolding protein that controls a rate-limiting step in the destruction of β-catenin, the central activator of the Wnt pathway. In Wnt-stimulated cells, Axin is rapidly modified by tankyrasemediated poly(ADP-ribosyl)ation, which promotes the proteolysis of Axin and consequent stabilization of β-catenin. Thus, regulation of the levels and activity of tankyrases is mechanistically important in controlling Wnt signaling. Here, we identify ubiquitin-specific protease 25 (USP25) as a positive regulator of Wnt/β-catenin signaling. We found that USP25 directly interacted with tankyrases to promote their deubiquitination and stabilization. We demonstrated that USP25 deficiency could promote the degradation of tankyrases and consequent stabilization of Axin to antagonize Wnt signaling. We further characterized the interaction between TNKS1 and USP25 by X-ray crystal structure determination. Our results provide important new insights into the molecular mechanism that regulates the turnover of tankyrases and the possibility of targeting the stability of tankyrases by antagonizing their interaction with USP25 to modulate the Wnt/β-catenin pathway.

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“…Standard site-directed mutagenesis is performed to mutate the Gly at position 6 of the FP-validated TBMs Validation of candidate TBM Fuorescence polarization (FP) to Arg. The FLAG-taggedFLAG tag candidate proteins and corresponding TBM mutant derivatives are next used as “baits” in co-immunoprecipitation experiments, to assess their binding to and their PARylation by MYC 2 -TNKS2, using catalytically inactive TNKS2 (G1032W) as control [31]. …”

Section: Methodsmentioning

confidence: 99%

“…4Assessing Tankyrase binding and PARylation by tankyrase in the full-length protein context. ( a ) FLAG 3 -TRF1, either in its wild-typeTankyrase Binders form or as a G18R TBM mutant (“mut.”), was co-expressed with the indicated MYC 2 -TNKS2 constructs, either in wild-type form or as a G1032W PARP-inactive mutant (“GW”) [31]. FLAG-TRF1 was immunoprecipitated and input and immunoprecipitate (IP) samples analyzed by SDS-PAGE and Western blotting as indicated.…”

Section: Methodsmentioning

confidence: 99%

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Identifying and Validating Tankyrase Binders and Substrates: A Candidate Approach

Pollock

Ranes

Collins

et al. 2017

Methods in Molecular Biology

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The poly(ADP-ribose)polymerase (PARP) enzyme tankyrase (TNKS/ARTD5, TNKS2/ARTD6) uses its ankyrin repeat clusters (ARCs) to recognize degenerate peptide motifs in a wide range of proteins, thereby recruiting such proteins and their complexes for scaffolding and/or poly(ADP-ribosyl)ation. Here, we provide guidance for predicting putative tankyrase-binding motifs, based on the previously delineated peptide sequence rules and existing structural information. We present a general method for the expression and purification of tankyrase ARCs from Escherichia coli and outline a fluorescence polarization assay to quantitatively assess direct ARC–TBM peptide interactions. We provide a basic protocol for evaluating binding and poly(ADP-ribosyl)ation of full-length candidate interacting proteins by full-length tankyrase in mammalian cells.

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Tankyrase Requires SAM Domain-Dependent Polymerization to Support Wnt-β-Catenin Signaling

Cited by 80 publications

References 67 publications

Structural basis for tankyrase‐RNF146 interaction reveals noncanonical tankyrase‐binding motifs

Structural basis for tankyrase‐RNF146 interaction reveals noncanonical tankyrase‐binding motifs

USP25 regulates Wnt signaling by controlling the stability of tankyrases

Identifying and Validating Tankyrase Binders and Substrates: A Candidate Approach

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