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1.
Cell Death Dis ; 15(3): 211, 2024 Mar 13.
Article En | MEDLINE | ID: mdl-38480683

Activation of the Hippo pathway by angiomotins to limit colorectal cancer progression is prevalent, whereas the regulation of angiomotins remains elusive. In this study, we uncover the involvement of an upregulated E3 ubiquitin ligase called RNF166, which destabilizes angiomotins, activates YAP, and is associated with a poor prognosis in colorectal cancer patients. Mechanistically, RNF166 specifically recognizes PARsylated angiomotin, a modification mediated by tankyrase at specific amino acid residues (D506, E513, E516, and E528). The tankyrase inhibitor XAV939, effectively prevents RNF166-dependent destabilization of angiomotins and subsequent activation of YAP. Additionally, YAP-5SA, a constitutively active form of YAP, rescues colorectal cancer progression following knockdown of RNF166. Importantly, the C-terminus of RNF66, particularly the Di19-ZF domain, is the crucial region responsible for recognizing ADP-ribosylated angiomotins. Together, this work not only sheds light on the regulation of the Hippo pathway in colorectal cancer but also uncovers a novel poly(ADP-ribose)-binding domain, which may serve as a potential therapeutic target for intervention.


Colorectal Neoplasms , Tankyrases , Humans , Angiomotins , Adaptor Proteins, Signal Transducing/metabolism , Signal Transduction , Tankyrases/metabolism , Colorectal Neoplasms/genetics , Ubiquitin-Protein Ligases/metabolism
2.
Expert Opin Ther Targets ; 28(3): 159-177, 2024 Mar.
Article En | MEDLINE | ID: mdl-38497299

INTRODUCTION: The pathways like Wingless-related integration (Wnt/ß-catenin) and PI3K play an important role in colorectal cancer (CRC) development; however, their roles are distinct in the process of oncogenesis. Despite their differences, these pathways interact through feedback mechanisms and regulate the common effectors both in the upstream and the downstream processes in normal and pathological conditions. Their ability to reciprocally control each other is a primary resistance mechanism for the selective inhibitors in CRC. AREA COVERED: This review highlights the Wnt/ß-catenin and PI3K pathways that are interrelated in CRC, recent advances and some key perspectives in developing inhibitors that could target the tankyrase enzyme and PI3K, apart from a brief description of the potential of dual inhibitors of PI3K and Tankyrases (TNKS). EXPERT OPINION: Recent research has focused on overcoming the challenges particularly relating to the resistance and efficacy of dual inhibitors targeting PI3K and tankyrase proteins. Despite these challenges, PI3K as well as tankyrases remain promising therapeutic targets for the treatment of solid tumors. The design of potent inhibitors is crucial to effectively block these protein signaling pathways. Moreover, it is essential to explore the potential of dual-target inhibition of other signaling pathways in conjunction with PI3K and tankyrase.


Antineoplastic Agents , Colorectal Neoplasms , Molecular Targeted Therapy , Phosphoinositide-3 Kinase Inhibitors , Tankyrases , Wnt Signaling Pathway , Humans , Tankyrases/antagonists & inhibitors , Tankyrases/metabolism , Colorectal Neoplasms/drug therapy , Colorectal Neoplasms/pathology , Animals , Phosphoinositide-3 Kinase Inhibitors/pharmacology , Antineoplastic Agents/pharmacology , Wnt Signaling Pathway/drug effects , Drug Development , Drug Resistance, Neoplasm , Phosphatidylinositol 3-Kinases/metabolism
3.
Nucleic Acids Res ; 52(7): 4002-4020, 2024 Apr 24.
Article En | MEDLINE | ID: mdl-38321934

Poly(ADP-ribosylation) (PARylation) is a post-translational modification mediated by a subset of ADP-ribosyl transferases (ARTs). Although PARylation-inhibition based therapies are considered as an avenue to combat debilitating diseases such as cancer and myopathies, the role of this modification in physiological processes such as cell differentiation remains unclear. Here, we show that Tankyrase1 (TNKS1), a PARylating ART, plays a major role in myogenesis, a vital process known to drive muscle fiber formation and regeneration. Although all bona fide PARPs are expressed in muscle cells, experiments using siRNA-mediated knockdown or pharmacological inhibition show that TNKS1 is the enzyme responsible of catalyzing PARylation during myogenesis. Via this activity, TNKS1 controls the turnover of mRNAs encoding myogenic regulatory factors such as nucleophosmin (NPM) and myogenin. TNKS1 mediates these effects by targeting RNA-binding proteins such as Human Antigen R (HuR). HuR harbors a conserved TNKS-binding motif (TBM), the mutation of which not only prevents the association of HuR with TNKS1 and its PARylation, but also precludes HuR from regulating the turnover of NPM and myogenin mRNAs as well as from promoting myogenesis. Therefore, our data uncover a new role for TNKS1 as a key modulator of RBP-mediated post-transcriptional events required for vital processes such as myogenesis.


Muscle Development , Muscle Fibers, Skeletal , Myogenin , RNA, Messenger , Tankyrases , Tankyrases/metabolism , Tankyrases/genetics , Humans , RNA, Messenger/metabolism , RNA, Messenger/genetics , Muscle Development/genetics , Animals , Muscle Fibers, Skeletal/metabolism , Mice , Myogenin/genetics , Myogenin/metabolism , Nucleophosmin , ELAV-Like Protein 1/metabolism , ELAV-Like Protein 1/genetics , RNA Stability/genetics , Poly ADP Ribosylation/genetics , Cell Line , Nuclear Proteins/metabolism , Nuclear Proteins/genetics , Cell Differentiation/genetics , RNA-Binding Proteins/metabolism , RNA-Binding Proteins/genetics , HEK293 Cells
4.
Br J Cancer ; 130(1): 151-162, 2024 01.
Article En | MEDLINE | ID: mdl-37968472

BACKGROUND: Aberrant WNT/ß-catenin signaling drives carcinogenesis. Tankyrases poly(ADP-ribosyl)ate and destabilize AXINs, ß-catenin repressors. Tankyrase inhibitors block WNT/ß-catenin signaling and colorectal cancer (CRC) growth. We previously reported that 'short' APC mutations, lacking all seven ß-catenin-binding 20-amino acid repeats (20-AARs), are potential predictive biomarkers for CRC cell sensitivity to tankyrase inhibitors. Meanwhile, 'Long' APC mutations, which possess more than one 20-AAR, do not predict inhibitor-resistant cells. Thus, additional biomarkers are needed to precisely predict the inhibitor sensitivity. METHODS: Using 47 CRC patient-derived cells (PDCs), we examined correlations between the sensitivity to tankyrase inhibitors (G007-LK and RK-582), driver mutations, and the expressions of signaling factors. NOD.CB17-Prkdcscid/J and BALB/c-nu/nu xenograft mice were treated with RK-582. RESULTS: Short APC mutant CRC cells exhibited high/intermediate sensitivities to tankyrase inhibitors in vitro and in vivo. Active ß-catenin levels correlated with inhibitor sensitivity in both short and long APC mutant PDCs. PIK3CA mutations, but not KRAS/BRAF mutations, were more frequent in inhibitor-resistant PDCs. Some wild-type APC PDCs showed inhibitor sensitivity in a ß-catenin-independent manner. CONCLUSIONS: APC/PIK3CA mutations and ß-catenin predict the sensitivity of APC-mutated CRC PDCs to tankyrase inhibitors. These observations may help inform the strategy of patient selection in future clinical trials of tankyrase inhibitors.


Colorectal Neoplasms , Tankyrases , Animals , Mice , Humans , Colorectal Neoplasms/drug therapy , Colorectal Neoplasms/genetics , Colorectal Neoplasms/metabolism , Tankyrases/genetics , Tankyrases/metabolism , Cell Line, Tumor , beta Catenin/genetics , beta Catenin/metabolism , Mice, Inbred NOD , Wnt Signaling Pathway/genetics , Biomarkers , Class I Phosphatidylinositol 3-Kinases/genetics , Class I Phosphatidylinositol 3-Kinases/metabolism
5.
Int J Mol Sci ; 24(23)2023 Nov 30.
Article En | MEDLINE | ID: mdl-38069287

Tankyrases, a versatile protein group within the poly(ADP-ribose) polymerase family, are essential for post-translational poly(ADP-ribosyl)ation, influencing various cellular functions and contributing to diseases, particularly cancer. Consequently, tankyrases have become important targets for anti-cancer drug development. Emerging approaches in drug discovery aim to disrupt interactions between tankyrases and their binding partners, which hinge on tankyrase-binding motifs (TBMs) within partner proteins and ankyrin repeat cluster domains within tankyrases. Our study addresses the challenge of identifying and ranking TBMs. We have conducted a comprehensive review of the existing literature, classifying TBMs into three distinct groups, each with its own scoring system. To facilitate this process, we introduce TBM Hunter-an accessible, web-based tool. This user-friendly platform provides a cost-free and efficient means to screen and assess potential TBMs within any given protein. TBM Hunter can handle individual proteins or lists of proteins simultaneously. Notably, our results demonstrate that TBM Hunter not only identifies known TBMs but also uncovers novel ones. In summary, our study offers an all-encompassing perspective on TBMs and presents an easy-to-use, precise, and free tool for identifying and evaluating potential TBMs in any protein, thereby enhancing research and drug development efforts focused on tankyrases.


Tankyrases , Tankyrases/metabolism , Ankyrin Repeat , Poly ADP Ribosylation
6.
Cell Rep ; 42(10): 113235, 2023 10 31.
Article En | MEDLINE | ID: mdl-37843976

Resolution of cohesion between sister telomeres in human cells depends on TRF1-mediated recruitment of the polyADP-ribosyltransferase tankyrase to telomeres. In human aged cells, due to insufficient recruitment of TRF1/tankyrase to shortened telomeres, sisters remain cohered in mitosis. This persistent cohesion plays a protective role, but the mechanism by which sisters remain cohered is not well understood. Here we show that telomere repeat-containing RNA (TERRA) holds sister telomeres together through RNA-DNA hybrid (R-loop) structures. We show that a tankyrase-interacting partner, the RNA-binding protein C19orf43, is required for repression of TERRA R-loops. Persistent telomere cohesion in C19orf43-depleted cells is counteracted by RNaseH1, confirming that RNA-DNA hybrids hold sisters together. Consistent with a protective role for persistent telomere cohesion, depletion of C19orf43 in aged cells reduces DNA damage and delays replicative senescence. We propose that the inherent inability of shortened telomeres to recruit R-loop-repressing machinery permits a controlled onset of senescence.


Tankyrases , Humans , Tankyrases/genetics , Tankyrases/metabolism , R-Loop Structures , Telomere/metabolism , Mitosis , RNA , DNA
7.
J Cell Sci ; 136(20)2023 10 15.
Article En | MEDLINE | ID: mdl-37721093

Axin (also known as AXIN1) is a central negative regulator of the proto-oncogenic Wnt/ß-catenin signaling pathway, as axin condensates provide a scaffold for the assembly of a multiprotein complex degrading ß-catenin. Axin, in turn, is degraded through tankyrase. Consequently, tankyrase small-molecule inhibitors block Wnt signaling by stabilizing axin, revealing potential for cancer therapy. Here, we discovered that axin is phosphorylated by casein kinase 1 alpha 1 (CSNK1A1, also known as CK1α) at an N-terminal casein kinase 1 consensus motif, and that this phosphorylation is antagonized by the catalytic subunit alpha of protein phosphatase 1 (PPP1CA, hereafter referred to as PP1). Axin condensates promoted phosphorylation by enriching CK1α over PP1. Importantly, the phosphorylation took place within the tankyrase-binding site, electrostatically and/or sterically hindering axin-tankyrase interaction, and counteracting tankyrase-mediated degradation of axin. Thus, the presented data propose a novel mechanism regulating axin stability, with implications for Wnt signaling, cancer therapy and self-organization of biomolecular condensates.


Neoplasms , Tankyrases , Humans , Axin Protein/metabolism , Phosphorylation , Tankyrases/metabolism , Biomolecular Condensates , beta Catenin/metabolism , Wnt Signaling Pathway
8.
Am J Hum Genet ; 110(9): 1470-1481, 2023 09 07.
Article En | MEDLINE | ID: mdl-37582359

Sclerosing skeletal dysplasias result from an imbalance between bone formation and resorption. We identified three homozygous, C-terminally truncating AXIN1 variants in seven individuals from four families affected by macrocephaly, cranial hyperostosis, and vertebral endplate sclerosis. Other frequent findings included hip dysplasia, heart malformations, variable developmental delay, and hematological anomalies. In line with AXIN1 being a central component of the ß-catenin destruction complex, analyses of primary and genome-edited cells harboring the truncating variants revealed enhanced basal canonical Wnt pathway activity. All three AXIN1-truncating variants resulted in reduced protein levels and impaired AXIN1 polymerization mediated by its C-terminal DIX domain but partially retained Wnt-inhibitory function upon overexpression. Addition of a tankyrase inhibitor attenuated Wnt overactivity in the AXIN1-mutant model systems. Our data suggest that AXIN1 coordinates the action of osteoblasts and osteoclasts and that tankyrase inhibitors can attenuate the effects of AXIN1 hypomorphic variants.


Hip Dislocation , Osteosclerosis , Tankyrases , Humans , Tankyrases/genetics , Tankyrases/metabolism , Axin Protein/genetics , Axin Protein/metabolism , Wnt Signaling Pathway/genetics , Osteosclerosis/genetics , beta Catenin/metabolism
9.
Naunyn Schmiedebergs Arch Pharmacol ; 396(12): 3363-3374, 2023 12.
Article En | MEDLINE | ID: mdl-37338576

Tankyrase 1 (TNKS1) and tankyrase 2 (TNKS2) enzymes belong to the poly (ADP-ribose) polymerase (PARP) family participates in process of poly-ADP-ribosylation of different target proteins which leads to ubiquitin-mediated proteasomal degradation. Tankyrases are also involved in the pathophysiology of many diseases, especially cancer. Their functions include cell cycle homeostasis (primarily in mitosis), telomere maintenance, Wnt signaling pathway regulation, and insulin signaling (particularly GLUT4 translocation). Studies have implicated that genetic changes, mutations in the tankyrase coding sequence, or up regulation and down regulation of tankyrase are reflected in the numerous disease conditions. Investigations are pursued to develop putative molecules that target tankyrase in various diseases such as cancer, obesity, osteoarthritis, fibrosis, cherubism, and diabetes, thereby providing a new therapeutic treatment option. In the present review, we described the structure and function of tankyrase along with its role in different disease conditions. Furthermore, we also presented cumulative experimental evidences of different drugs acting on tankyrase.


Neoplasms , Tankyrases , Humans , Tankyrases/metabolism , Wnt Signaling Pathway , Mitosis
10.
Cancer Gene Ther ; 30(8): 1144-1155, 2023 08.
Article En | MEDLINE | ID: mdl-37248434

The tumor suppressor gene WWOX is localized in an unstable chromosomal region and its expression is decreased or absent in several types of cancer. A low expression of WWOX is associated with a poor prognosis in breast cancer (BC). It has recently been shown that WWOX contributes to genome stability through its role in the DNA damage response (DDR). In breast cancer cells, WWOX inhibits homologous recombination (HR), and thus promotes the repair of DNA double-stranded breaks (DSBs) by non-homologous end joining (NHEJ). The fine-tuning modulation of HR activity is crucial. Its under or overstimulation inducing genome alterations that can induce cancer. MERIT40 is a positive regulator of the DDR. This protein is indispensable for the function of the multi-protein complex BRCA1-A, which suppresses excessive HR activity. MERIT40 also recruits Tankyrase, a positive regulator of HR, to the DSBs to stimulate DNA repair. Here, we identified MERIT40 as a new molecular partner of WWOX. We demonstrated that WWOX inhibited excessive HR activity induced by overexpression of MERIT40. We showed that WWOX impaired the MERIT40-Tankyrase interaction preventing the role of the complex on DSBs. Furthermore, we found that MERIT40 is overexpressed in BC and that this overexpression is associated to a poor prognosis. These results strongly suggest that WWOX, through its interaction with MERIT40, prevents the deleterious impact of excessive HR on BC development by inhibiting MERIT40-Tankyrase association. This inhibitory effect of WWOX would oppose MERIT40-dependent BC development.


Breast Neoplasms , Homologous Recombination , Female , Humans , Breast Neoplasms/genetics , DNA Breaks, Double-Stranded , DNA Repair , Tankyrases/genetics , Tankyrases/metabolism , Tumor Suppressor Proteins/genetics , Tumor Suppressor Proteins/metabolism , WW Domain-Containing Oxidoreductase/genetics , WW Domain-Containing Oxidoreductase/metabolism
11.
J Cancer Res Clin Oncol ; 149(10): 7103-7112, 2023 Aug.
Article En | MEDLINE | ID: mdl-36871092

PURPOSE: Chronic myeloid leukemia (CML) is a myeloproliferative neoplasm characterized by recurrent genetic aberration in leukemic stem cells, namely Philadelphia chromosome caused by reciprocal translocation t(9;22)(q34;q11). In our study, we analyzed the telomeric complex expression and function in the molecular pathogenesis of CML. METHODS: We employed CD34+ primary leukemic cells, comprising both leukemic stem and progenitor cell populations, isolated from peripheral blood or bone marrow of CML patients in chronic and blastic phase to analyze the telomere length and telomeric-associated proteins. RESULTS: The reduction in telomere length during disease progression was correlated with increased expression of BCR::ABL1 transcript and the dynamic changes were neither associated with the enzymatic activity of telomerase nor with gene copy number and expression of telomerase subunits. Increased expression of BCR::ABL1 was positively correlated with expression of TRF2, RAP1, TPP1, DKC1, TNKS1, and TNKS2 genes. CONCLUSIONS: The dynamics of telomere length changes in CD34+ CML cells is dependent on the expression level of BCR::ABL, which promotes the expression of certain shelterins including RAP1 and TRF2, as well as TNKS, and TNKS2, and results in telomere shortening regardless of telomerase activity. Our results may allow better understanding of the mechanisms responsible for the genomic instability of leukemic cells and CML progression.


Leukemia, Myelogenous, Chronic, BCR-ABL Positive , Tankyrases , Telomerase , Humans , Bone Marrow/metabolism , Cell Cycle Proteins/genetics , Fusion Proteins, bcr-abl/genetics , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/pathology , Nuclear Proteins/genetics , Tankyrases/genetics , Tankyrases/metabolism , Telomerase/genetics , Telomerase/metabolism , Telomere/metabolism
12.
Trends Mol Med ; 29(5): 390-405, 2023 05.
Article En | MEDLINE | ID: mdl-36948987

Modification of proteins by ADP-ribose (PARsylation) is catalyzed by the poly(ADP-ribose) polymerase (PARP) family of enzymes exemplified by PARP1, which controls chromatin organization and DNA repair. Additionally, PARsylation induces ubiquitylation and proteasomal degradation of its substrates because PARsylation creates a recognition site for E3-ubiquitin ligase. The steady-state levels of the adaptor protein SH3-domain binding protein 2 (3BP2) is negatively regulated by tankyrase (PARP5), which coordinates ubiquitylation of 3BP2 by the E3-ligase ring finger protein 146 (RNF146). 3BP2 missense mutations uncouple 3BP2 from tankyrase-mediated negative regulation and cause Cherubism, an autosomal dominant autoinflammatory disorder associated with craniofacial dysmorphia. In this review, we summarize the diverse biological processes, including bone dynamics, metabolism, and Toll-like receptor (TLR) signaling controlled by tankyrase-mediated PARsylation of 3BP2, and highlight the therapeutic potential of this pathway.


Cherubism , Tankyrases , Humans , Tankyrases/genetics , Tankyrases/chemistry , Tankyrases/metabolism , Cherubism/genetics , Cherubism/metabolism , Ubiquitination , Signal Transduction , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/metabolism
13.
Adv Sci (Weinh) ; 10(10): e2207693, 2023 04.
Article En | MEDLINE | ID: mdl-36703617

Transferrin receptor (TFRC) is the major mediator for iron entry into a cell. Under excessive iron conditions, TFRC is expected to be reduced to lower iron uptake and toxicity. However, the mechanism whereby TFRC expression is maintained at high levels in iron-enriched cancer cells and the contribution of TFRC to cancer development are enigmatic. Here the work shows TFRC is induced by adenomatous polyposis coli (APC) gene loss-driven ß-catenin activation in colorectal cancer, whereas TFRC-mediated intratumoral iron accumulation potentiates ß-catenin signaling by directly enhancing the activity of tankyrase. Disruption of TFRC leads to a reduction of colonic iron levels and iron-dependent tankyrase activity, which caused stabilization of axis inhibition protein 2 (AXIN2) and subsequent repression of the ß-catenin/c-Myc/E2F Transcription Factor 1/DNA polymerase delta1 (POLD1) axis. POLD1 knockdown, iron chelation, and TFRC disruption increase DNA replication stress, DNA damage response, apoptosis, and reduce colon tumor growth. Importantly, a combination of iron chelators and DNA damaging agents increases DNA damage response and reduces colon tumor cell growth. TFRC-mediated iron import is at the center of a novel feed-forward loop that facilitates colonic epithelial cell survival. This discovery may provide novel strategies for colorectal cancer therapy.


Colonic Neoplasms , Tankyrases , Humans , beta Catenin/metabolism , Iron/metabolism , Tankyrases/metabolism , Colonic Neoplasms/genetics , Carcinogenesis/genetics , Cell Transformation, Neoplastic , Receptors, Transferrin/genetics , Receptors, Transferrin/metabolism
14.
Eur J Med Chem ; 246: 114980, 2023 Jan 15.
Article En | MEDLINE | ID: mdl-36495630

DNA-encoded chemical libraries (DECLs) interrogate the interactions of a target of interest with vast numbers of molecules. DECLs hence provide abundant information about the chemical ligand space for therapeutic targets, and there is considerable interest in methods for exploiting DECL screening data to predict novel ligands. Here we introduce one such approach and demonstrate its feasibility using the cancer-related poly-(ADP-ribose)transferase tankyrase 1 (TNKS1) as a model target. First, DECL affinity selections resulted in structurally diverse TNKS1 inhibitors with high potency including compound 2 with an IC50 value of 0.8 nM. Additionally, TNKS1 hits from four DECLs were translated into pharmacophore models, which were exploited in combination with docking-based screening to identify TNKS1 ligand candidates in databases of commercially available compounds. This computational strategy afforded TNKS1 inhibitors that are outside the chemical space covered by the DECLs and yielded the drug-like lead compound 12 with an IC50 value of 22 nM. The study further provided insights in the reliability of screening data and the effect of library design on hit compounds. In particular, the study revealed that while in general DECL screening data are in good agreement with off-DNA ligand binding, unpredictable interactions of the DNA-attachment linker with the target protein contribute to the noise in the affinity selection data.


Small Molecule Libraries , Tankyrases , Small Molecule Libraries/chemistry , Pharmacophore , Tankyrases/metabolism , Ligands , Reproducibility of Results , DNA/metabolism
15.
Biochem J ; 479(22): 2379-2394, 2022 11 30.
Article En | MEDLINE | ID: mdl-36383218

p21WAF1/Cip1 acts as a key negative regulator of cell cycle progression, which can form complexes with cyclin-dependent kinases together with specific cyclins to induce cell cycle arrest at specific stages. p21 protein levels have been shown to be regulated primarily through phosphorylation and ubiquitination during various stages of the cell cycle. Although phosphorylation and ubiquitin-dependent proteasomal degradation of p21 have been well established, other post-translational modifications that contribute to regulation of p21 stability and function remain to be further elucidated. Here, we show that p21 degradation and its function are controlled by tankyrases, which are members of the poly(ADP-ribose) polymerase (PARP) protein family. p21 interacts with tankyrases via newly defined tankyrase-binding motifs and is PARylated by tankyrases in vitro and in vivo, suggesting that PARylation is a new post-translational modification of p21. Up-regulation of tankyrases induces ubiquitin-dependent proteasomal degradation of p21 through an E3 ligase RNF146, thus promoting cell cycle progression in the G1/S phase transition. On the contrary, inhibition of tankyrases by knockdown or inhibitor treatment stabilizes p21 protein and leads to cell cycle arrest in the G1 phase. Together, our data demonstrate that tankyrase may function as a new molecular regulator that controls the protein levels of p21 through PARylation-dependent proteasomal degradation. Hence, a novel function of the tankyrase-p21 axis may represent a new avenue for regulating cell cycle progression.


Tankyrases , Tankyrases/chemistry , Tankyrases/metabolism , Poly ADP Ribosylation , Ubiquitination , Cell Cycle , Ubiquitins/metabolism
16.
Development ; 149(24)2022 12 15.
Article En | MEDLINE | ID: mdl-36398796

Propagation of human naïve pluripotent stem cells (nPSCs) relies on the inhibition of MEK/ERK signalling. However, MEK/ERK inhibition also promotes differentiation into trophectoderm (TE). Therefore, robust self-renewal requires suppression of TE fate. Tankyrase inhibition using XAV939 has been shown to stabilise human nPSCs and is implicated in TE suppression. Here, we dissect the mechanism of this effect. Tankyrase inhibition is known to block canonical Wnt/ß-catenin signalling. However, we show that nPSCs depleted of ß-catenin remain dependent on XAV939. Rather than inhibiting Wnt, we found that XAV939 prevents TE induction by reducing activation of YAP, a co-factor of TE-inducing TEAD transcription factors. Tankyrase inhibition stabilises angiomotin, which limits nuclear accumulation of YAP. Upon deletion of angiomotin-family members AMOT and AMOTL2, nuclear YAP increases and XAV939 fails to prevent TE induction. Expression of constitutively active YAP similarly precipitates TE differentiation. Conversely, nPSCs lacking YAP1 or its paralog TAZ (WWTR1) resist TE differentiation and self-renewal efficiently without XAV939. These findings explain the distinct requirement for tankyrase inhibition in human but not in mouse nPSCs and highlight the pivotal role of YAP activity in human naïve pluripotency and TE differentiation. This article has an associated 'The people behind the papers' interview.


Angiomotins , Pluripotent Stem Cells , Tankyrases , YAP-Signaling Proteins , Animals , Humans , Mice , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/metabolism , beta Catenin/metabolism , Mitogen-Activated Protein Kinase Kinases/metabolism , Tankyrases/metabolism , Wnt Signaling Pathway , Pluripotent Stem Cells/cytology
17.
Nature ; 612(7938): 162-169, 2022 12.
Article En | MEDLINE | ID: mdl-36418402

The poly-ADP-ribosyltransferase tankyrase (TNKS, TNKS2) controls a wide range of disease-relevant cellular processes, including WNT-ß-catenin signalling, telomere length maintenance, Hippo signalling, DNA damage repair and glucose homeostasis1,2. This has incentivized the development of tankyrase inhibitors. Notwithstanding, our knowledge of the mechanisms that control tankyrase activity has remained limited. Both catalytic and non-catalytic functions of tankyrase depend on its filamentous polymerization3-5. Here we report the cryo-electron microscopy reconstruction of a filament formed by a minimal active unit of tankyrase, comprising the polymerizing sterile alpha motif (SAM) domain and its adjacent catalytic domain. The SAM domain forms a novel antiparallel double helix, positioning the protruding catalytic domains for recurring head-to-head and tail-to-tail interactions. The head interactions are highly conserved among tankyrases and induce an allosteric switch in the active site within the catalytic domain to promote catalysis. Although the tail interactions have a limited effect on catalysis, they are essential to tankyrase function in WNT-ß-catenin signalling. This work reveals a novel SAM domain polymerization mode, illustrates how supramolecular assembly controls catalytic and non-catalytic functions, provides important structural insights into the regulation of a non-DNA-dependent poly-ADP-ribosyltransferase and will guide future efforts to modulate tankyrase and decipher its contribution to disease mechanisms.


Biocatalysis , Cryoelectron Microscopy , Polymerization , Tankyrases , beta Catenin , Tankyrases/chemistry , Tankyrases/metabolism , Tankyrases/ultrastructure , Enzyme Activation , Catalytic Domain , Wnt Signaling Pathway , Amino Acid Motifs
18.
Biomolecules ; 12(11)2022 11 15.
Article En | MEDLINE | ID: mdl-36421702

Tankyrases are poly-ADP-ribosyltransferases that regulate many crucial and diverse cellular processes in humans such as Wnt signaling, telomere homeostasis, mitotic spindle formation and glucose metabolism. While tankyrases are present in most animals, functional differences across species may exist. In this work, we confirm the widespread distribution of tankyrases throughout the branches of multicellular animal life and identify the single-celled choanoflagellates as earliest origin of tankyrases. We further show that the sequences and structural aspects of TNKSs are well-conserved even between distantly related species. We also experimentally characterized an anciently diverged tankyrase homolog from the sponge Amphimedon queenslandica and show that the basic functional aspects, such as poly-ADP-ribosylation activity and interaction with the canonical tankyrase binding peptide motif, are conserved. Conversely, the presence of tankyrase binding motifs in orthologs of confirmed interaction partners varies greatly between species, indicating that tankyrases may have different sets of interaction partners depending on the animal lineage. Overall, our analysis suggests a remarkable degree of conservation for tankyrases, and that their regulatory functions in cells have likely changed considerably throughout evolution.


Tankyrases , Humans , Animals , Tankyrases/genetics , Tankyrases/chemistry , Tankyrases/metabolism , Telomere Homeostasis , Wnt Signaling Pathway
19.
Int J Mol Sci ; 23(17)2022 Sep 02.
Article En | MEDLINE | ID: mdl-36077457

Hyperactive poly(ADP-ribose) polymerases (PARP) promote ischemic heart failure (IHF) after myocardial infarction (MI). However, the role of tankyrases (TNKSs), members of the PARP family, in pathogenesis of IHF remains unknown. We investigated the expression and activation of TNKSs in myocardium of IHF patients and MI rats. We explored the cardioprotective effect of TNKS inhibition in an isoproterenol-induced zebrafish HF model. In IHF patients, we observed elevated TNKS2 and DICER and concomitant upregulation of miR-34a-5p and miR-21-5p in non-infarcted myocardium. In a rat MI model, we found augmented TNKS2 and DICER in the border and infarct areas at the early stage of post-MI. We also observed consistently increased TNKS1 in the border and infarct areas and destabilized AXIN in the infarct area from 4 weeks onward, which in turn triggered Wnt/ß-catenin signaling. In an isoproterenol-induced HF zebrafish model, inhibition of TNKS activity with XAV939, a TNKSs-specific inhibitor, protected against ventricular dilatation and cardiac dysfunction and abrogated overactivation of Wnt/ß-catenin signaling and dysregulation of miR-34a-5p induced by isoproterenol. Our study unravels a potential role of TNKSs in the pathogenesis of IHF by regulating Wnt/ß-catenin signaling and possibly modulating miRNAs and highlights the pharmacotherapeutic potential of TNKS inhibition for prevention of IHF.


Heart Failure , MicroRNAs , Tankyrases , Animals , Dilatation , Heart Failure/drug therapy , Isoproterenol/pharmacology , MicroRNAs/genetics , Rats , Tankyrases/antagonists & inhibitors , Tankyrases/metabolism , Wnt Signaling Pathway , Zebrafish/metabolism , beta Catenin/metabolism
20.
Int J Mol Sci ; 23(18)2022 Sep 17.
Article En | MEDLINE | ID: mdl-36142793

Deregulation of signalling pathways that regulate cell growth, survival, metabolism, and migration can frequently lead to the progression of cancer. Brain tumours are a large group of malignancies characterised by inter- and intratumoral heterogeneity, with glioblastoma (GBM) being the most aggressive and fatal. The present study aimed to characterise the expression of cancer pathway-related genes (n = 84) in glial tumour cell lines (A172, SW1088, and T98G). The transcriptomic data obtained by the qRT-PCR method were compared to different control groups, and the most appropriate control for subsequent interpretation of the obtained results was chosen. We analysed three widely used control groups (non-glioma cells) in glioblastoma research: Human Dermal Fibroblasts (HDFa), Normal Human Astrocytes (NHA), and commercially available mRNAs extracted from healthy human brain tissues (hRNA). The gene expression profiles of individual glioblastoma cell lines may vary due to the selection of a different control group to correlate with. Moreover, we present the original multicriterial decision making (MCDM) for the possible characterization of gene expression profiles. We observed deregulation of 75 genes out of 78 tested in the A172 cell line, while T98G and SW1088 cells exhibited changes in 72 genes. By comparing the delta cycle threshold value of the tumour groups to the mean value of the three controls, only changes in the expression of 26 genes belonging to the following pathways were identified: angiogenesis FGF2; apoptosis APAF1, CFLAR, XIAP; cellular senescence BM1, ETS2, IGFBP5, IGFBP7, SOD1, TBX2; DNA damage and repair ERCC5, PPP1R15A; epithelial to mesenchymal transition SNAI3, SOX10; hypoxia ADM, ARNT, LDHA; metabolism ATP5A1, COX5A, CPT2, PFKL, UQCRFS1; telomeres and telomerase PINX1, TINF2, TNKS, and TNKS2. We identified a human astrocyte cell line and normal human brain tissue as the appropriate control group for an in vitro model, despite the small sample size. A different method of assessing gene expression levels produced the same disparities, highlighting the need for caution when interpreting the accuracy of tumorigenesis markers.


Brain Neoplasms , Glioblastoma , Tankyrases , Telomerase , Brain Neoplasms/metabolism , Cell Cycle Proteins/metabolism , Cell Line, Tumor , Epithelial-Mesenchymal Transition , Fibroblast Growth Factor 2/metabolism , Gene Expression Regulation, Neoplastic , Glioblastoma/metabolism , Humans , Superoxide Dismutase-1/genetics , Tankyrases/metabolism , Telomerase/metabolism , Tumor Suppressor Proteins/genetics
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