InteractiveFly: GeneBrief

tankyrase: Biological Overview | References

BIOLOGICAL OVERVIEW

Tankyrase (Tnks) transfers poly(ADP-ribose) on substrates. Whereas studies have highlighted the pivotal roles of Tnks in cancer, cherubism, systemic sclerosis, and viral infection, the requirement for Tnks under physiological contexts remains unclear. This study report that the loss of Tnks or its muscle-specific knockdown impairs lifespan, stress tolerance, and energy homeostasis in adult Drosophila. Tnks is a positive regulator in the JNK signaling pathway, and modest alterations in the activity of JNK signaling can strengthen or suppress the Tnks mutant phenotypes. JNK was identified as a direct substrate of Tnks. Although Tnks-dependent poly-ADP-ribosylation is tightly coupled to proteolysis in the proteasome, it was demonstrated that Tnks initiates degradation-independent ubiquitination on two lysine residues of JNK to promote its kinase activity and in vivo functions. This study uncovers a type of posttranslational modification of Tnks substrates and provides insights into Tnks-mediated physiological roles (Li, 2018).

Tankyrase (Tnks) belongs to the poly(ADP-ribose) polymerase (PARP) superfamily, which consists of 17 members in human. PARP-1 is the founding member of the family and has a critical role in the repair of DNA damage. The PARPs are characterized by a structurally similar PARP catalytic domain that successively transfers ADP-ribose from NAD+ onto substrate proteins. This post-translational modification is referred to as poly-ADP-ribosylation, or PARsylation. It has been shown that some PARPs actually catalyze mono-ADP-ribosylation rather than the polymerization of poly(ADP-ribose) chains. Because of this reason, the PARP family members have been renamed as ADP-ribosyltransferases diphtheria toxin-like (ARTDs). Tnks was identified as a telomere-associated protein that binds to the telomere-specific DNA binding protein TRF1. In addition to the PARP domain, Tnks contains two unique domains that distinguish it from other PARPs, including an Ankyrin repeat domain that is involved in the recruitment of substrate and a sterile alpha motif (SAM) that mediates oligomerization. Tnks is evolutionarily conserved in human, mouse, rat, chicken, C. elegans, and Drosophila. The human and mouse genome encodes two Tnks proteins (Tnk1/PARP5A/ARTD5 and Tnk2/PARP5B/ARTD6), whereas there is only one Tnks homolog in Drosophila (Li, 2018).

Tnks catalyzes PARsylation on its substrates and is involved in a variety of cellular processes, such as telomere homeostasis, cell-cycle progression, Wnt/β-catenin signaling, PI3K signaling, Hippo signaling, glucose metabolism, stress granule formation, and proteasome regulation. Various amino acid residues including lysine, arginine, aspartate, glutamate, asparagine, cysteine, phospho-serine, and diphthamide may serve as acceptors for PARsylation. In many cases, proteins modified by Tnks are subsequently poly-ubiquitinated and targeted for proteasomal degradation. For example, Tnks promotes telomere elongation by mediating the degradation of TRF1, a negative regulator of telomere length maintenance. Tnks PARsylates the β-catenin destruction complex component Axin, triggers its degradation, and thereby activates Wnt signaling. Tnks also regulates the stability of centrosomal P4.1-associated protein CPAP to limit centriole elongation during mitosis. On the other hand, the effects of PARsylation on some Tnks substrates, such as the nuclear mitotic apparatus protein NUMA, have not been elucidated, implying that alternative regulation following PARsylation may exist (Li, 2018).

Aberrant Tnks expression or activity has been implicated in a diversity of diseases including cancer, systemic sclerosis, cherubism, Herpes simplex, and Epstein-Barr viral infections. Particularly, the pro-oncogenic role of Tnks in many types of cancer strongly suggests a therapeutic benefit of Tnks inhibition. Whereas a substantial amount of studies have highlighted the important functions of Tnks under pathological conditions, the requirement for Tnks under physiological contexts is largely unexplored (Li, 2018).

This study investigate the physiological functions of Tnks during the adult stage in Drosophila with its mutant alleles and RNAi strains. The loss of Tnks was shown to impair lifespan, stress tolerance, and energy storage in adult flies. Ubiquitous or muscle-specific knockdown of Tnks causes defects similar to those observed in the mutant alleles. It was further shown that Tnks is specifically required for the activity of the c-Jun N-terminal kinase (JNK) and positively regulates the outputs of JNK signaling during organ development. In addition, mild reduction and slight increase in the activity of JNK signaling via genetic manipulations can strengthen and suppress the phenotypes of the Tnks mutants, respectively. Last, the results reveal for the first time that Tnks mediates degradation-independent ubiquitination on its substrate. Drosophila JNK was identified as a direct substrate protein of Tnks. The PARsylation by Tnks triggers K63-linked poly-ubiquitination on JNK, enhancing its kinase activity and maintaining its in vivo functions. Together, this study uncovers that Tnks is a positive regulator of JNK activity, mediates a novel type of post-translational modification, and functions through JNK signaling to affect lifespan, stress resistance, and energy homeostasis in Drosophila (Li, 2018).

PARsylation by Tnks appears to be tightly coupled to poly-ubiquitination and subsequent proteasome-dependent degradation, as observed for known Tnks substrates including Axin, TRF1, PTEN, 3BP2, CPAP, and AMOT family proteins. This study found that Tnks mediated PARsylation and poly-ubiquitination of Drosophila JNK (Bsk), however, without affecting its protein levels. This observation prompted investigation og the form of poly-ubiquitin chain assembled on Bsk in the presence of Tnks. Although all the seven lysine residues (K6, K11, K27, K29, K33, K48, and K63) and the N-terminal methionine residue in the ubiquitin can serve as the attachment site to generate poly-ubiquitin chain, K48-linked and K63-linked poly-ubiquitination are two predominant forms in cells. It is well-established that K48-linked poly-ubiquitination targets substrates to the 26S proteasome and promotes protein degradation, while K63-linked poly-ubiquitin chain performs non-proteolytic functions. Consistently, this study observed that Tnks promoted the assembly of K63-linked poly-ubiquitin chain on Bsk and increased its kinase activity. In contrast, Tnks did not affect K48-linked poly-ubiquitination of Bsk. Although the possibility cannot be completely that these modifications occur on a Bsk partner that tightly binds to Bsk even after stringent washing, this study reveals for the first time that Tnks mediates degradation-independent ubiquitination of its substrate, which may represent a novel type of modification induced by PARsylation. Indeed, it has been reported that PARsylation of some Tnks substrates such as the proteasome regulator PI31 and the mitotic spindle-pole protein NuMA may affect the activities of these proteins. It will be interesting to investigate whether similar modification occurs during these processes (Li, 2018).

JNK is an evolutionarily conserved stress-activated protein kinase (SAPK) and is one of the most versatile stress sensors in metazoans. The JNK signaling pathway adapts growth and metabolism to environmental conditions, and mediates stress tolerance, damage repair, and apoptosis. Therefore, JNK signaling has pivotal roles in regulating homeostasis, longevity, and organ development. This study reports an essential role of Tnks in regulating JNK signaling. As an initial hint, adult Tnks mutants are more sensitive to oxidative stress and to starvation, and live much shorter than the wild-type controls. Although several signaling pathways are known to coordinate these functions in Drosophila, this study observed that the loss of Tnks specifically decreased the activity of Bsk without affecting that of ERK, p38, Akt, and Nrf2 signaling. It was further found that Tnks was required for the outputs of JNK signaling during the development of eye, wing, and thorax. The knockdown of Tnks in the thorax strengthened the thoracic cleft phenotype caused by bsk knockdown. The loss of Tnks, or reducing its gene dosage, suppressed ectopic JNK signaling-induced phenotypes during the formation of compound eye and wing vein. In addition, Tnks overexpression in the wing disc activated puc expression in a bsk-dependent manner. Last, it was observed that bsk with mutations in Tnks-induced ubiquitination sites lacked the ability to rescue the lethality of the bsk^1/2 mutant and displayed impaired activity in regulating stress tolerance, climbing, energy storage, and organ development compared to wild-type bsk. This work thus reveals Tnks as a positive regulator of the JNK signaling pathway (Li, 2018).

Aberrant Tnks expression or activity has been implicated in a variety of disease states including cancer, cherubism, systemic sclerosis, and viral infection. The proposed roles of Tnks in telomere homeostasis, mitosis, and Wnt signaling have made it an attractive drug target in many types of cancer. Tnks is implied in a diversity of additional cellular processes, such as glucose metabolism, stress granule assembly, and proteasome regulation. However, the requirement for Tnks under physiological contexts remains poorly understood. While a double knockout of Tnks1 and Tnk2 is embryonically lethal in mice, Tnks mutant flies do not display any noticeable developmental defects. Although a previous study reported that Tnks-deficient mice exhibited substantially reduced adiposity, the underlying molecular mechanism is unclear. This work has focused on the physiological changes in Tnks mutant flies during the adult stage. It is reported that the loss of Tnks or its knockdown shortens lifespan, decreases climbing ability, reduces resistance to oxidative stress and starvation, and impairs energy storage in adult flies. Through tissue-specific knockdown, it is concluded that the above functions of Tnks are mainly mediated via its activity in the muscle. These physiological functions of Tnks are attributed to its regulation of JNK signaling activity, which is supported by several lines of evidence. First, the loss of Tnks specifically weakens Bsk activity and JNK signaling. Second, mild reduction in JNK signaling activity by removing one copy of bsk gene aggravates the phenotypes in Tnks mutants, whereas slightly elevated JNK signaling largely rescues these phenotypes. Third, this study shows that Tnks triggers PARsylation and K63-linked poly-ubiquitination of Bsk and enhances the kinase activity and in vivo functions of Bsk (Li, 2018).

Adherens junctions (AJ) are involved in cancer, infections and neurodegeneration. Still, their composition has not been completely disclosed. Poly(ADP-ribose) polymerases (PARPs) catalyze the synthesis of poly(ADP-ribose) (PAR) as a posttranslational modification. Four PARPs synthesize PAR, namely PARP-1/2 and Tankyrase-1/2 (TNKS). In the epithelial belt, AJ are accompanied by a PAR belt and a subcortical F-actin ring. F-actin depolymerization alters the AJ and PAR belts while PARP inhibitors prevent the assembly of the AJ belt and cortical actin. It was asked wondered which PARP synthesizes the belt and which is the PARylation target protein. Vinculin (VCL) participates in the anchorage of F-actin to the AJ, regulating its functions, and colocalized with the PAR belt. TNKS has been formerly involved in the assembly of epithelial cell junctions. TNKS poly(ADP-ribosylates) (PARylates) epithelial belt VCL, affecting its functions in AJ, including cell shape maintenance. In this work, some of the hypothesis predictions have been tested. It was demonstrated that: (1) VCL TBMs were conserved in vertebrate evolution while absent in C. elegans; (2) TNKS inhibitors disrupted the PAR belt synthesis, while PAR and an endogenous TNKS pool were associated to the plasma membrane; (3) a VCL pool was covalently PARylated; (4) transfection of MCF-7 cells leading to overexpression of Gg-VCL/*TBM induced mesenchymal-like cell shape changes. This last point deserves further investigation, bypassing the limits of the transient transfection and overexpression system. In fact, a 5th testable prediction would be that a single point mutation in VCL TBM-II under endogenous expression control would induce an epithelial to mesenchymal transition (EMT). To check this, a CRISPR/Cas9 substitution approach followed by migration, invasion, gene expression and chemo-resistance assays should be performed (Vilchez Larrea, 2021).

Axin proteolysis by Iduna is required for the regulation of stem cell proliferation and intestinal homeostasis in Drosophila

Self-renewal of intestinal stem cells is controlled by Wingless/Wnt-beta catenin signaling in both Drosophila and mammals. As Axin is a rate-limiting factor in Wingless signaling, its regulation is essential. Iduna (CG8786) is an evolutionarily conserved ubiquitin E3 ligase that has been identified as a crucial regulator for degradation of ADP-ribosylated Axin and, thus, of Wnt/beta-catenin signaling. However, its physiological significance remains to be demonstrated. This study generated loss-of-function mutants of Iduna to investigate its physiological role in Drosophila Genetic depletion of Iduna causes the accumulation of both Tankyrase and Axin. Increase of Axin protein in enterocytes non-autonomously enhanced stem cell divisions in the Drosophila midgut. Enterocytes secreted Unpaired proteins and thereby stimulated the activity of the JAK-STAT pathway in intestinal stem cells. A decrease in Axin gene expression suppressed the over-proliferation of stem cells and restored their numbers to normal levels in Iduna mutants. These findings suggest that Iduna-mediated regulation of Axin proteolysis is essential for tissue homeostasis in the Drosophila midgut (Gultekin, 2019).

A Context-Dependent Role for the RNF146 Ubiquitin Ligase in Wingless/Wnt Signaling in Drosophila

Aberrant activation of the Wnt signal transduction pathway triggers the development of colorectal cancer. The ADP-ribose polymerase Tankyrase (TNKS) mediates proteolysis of Axin, a negative regulator of Wnt signaling, and provides a promising therapeutic target for Wnt-driven diseases. Proteolysis of TNKS substrates is mediated through their ubiquitination by the poly-ADP-ribose (pADPr)-dependent RING-domain E3 ubiquitin ligase RNF146/Iduna. Like TNKS, RNF146 promotes Axin proteolysis and Wnt pathway activation in some cultured cell lines, but in contrast with TNKS, RNF146 is dispensable for Axin degradation in colorectal carcinoma cells. Thus the contexts in which RNF146 is essential for TNKS-mediated Axin destabilization and Wnt signaling remain uncertain. This study tested the requirement for RNF146 in TNKS-mediated Axin proteolysis and Wnt pathway activation in a range of in vivo settings. Using null mutants in Drosophila, genetic and biochemical evidence is provided that Rnf146 and Tnks function in the same proteolysis pathway in vivo. Furthermore, like Tnks, Drosophila Rnf146 promotes Wingless signaling in multiple developmental contexts by buffering Axin levels to ensure they remain below the threshold at which Wingless signaling is inhibited. However, in contrast with Tnks, Rnf146 is dispensable for Wingless target gene activation and the Wingless-dependent control of intestinal stem cell proliferation in the adult midgut during homeostasis. Together, these findings demonstrate that the requirement for Rnf146 in Tnks-mediated Axin proteolysis and Wingless pathway activation is dependent on physiological context, and suggest that in some cell types, functionally redundant pADPr-dependent E3 ligases or other compensatory mechanisms promote the Tnks-dependent proteolysis of Axin in both mammalian and Drosophila cells (Wang, 2019).

Poly(ADP-Ribose) Prevents Pathological Phase Separation of TDP-43 by Promoting Liquid Demixing and Stress Granule Localization

In amyotrophic lateral sclerosis (ALS) and frontotemporal degeneration (FTD), cytoplasmic aggregates of hyperphosphorylated TDP-43 accumulate and colocalize with some stress granule components, but how pathological TDP-43 aggregation is nucleated remains unknown. In Drosophila, it was established that downregulation of tankyrase, a poly(ADP-ribose) (PAR) polymerase, reduces TDP-43 accumulation in the cytoplasm and potently mitigates neurodegeneration. TDP-43 non-covalently binds to PAR via PAR-binding motifs embedded within its nuclear localization sequence. PAR binding promotes liquid-liquid phase separation of TDP-43 in vitro and is required for TDP-43 accumulation in stress granules in mammalian cells and neurons. Stress granule localization initially protects TDP-43 from disease-associated phosphorylation, but upon long-term stress, stress granules resolve, leaving behind aggregates of phosphorylated TDP-43. Finally, small-molecule inhibition of Tankyrase-1/2 in mammalian cells inhibits formation of cytoplasmic TDP-43 foci without affecting stress granule assembly. Thus, Tankyrase inhibition antagonizes TDP-43-associated pathology and neurodegeneration and could have therapeutic utility for ALS and FTD (McGurk, 2018).

Tankyrase regulates apoptosis by activating JNK signaling in Drosophila

Programmed cell death (PCD), or apoptosis, plays essential roles in various cellular and developmental processes, and deregulation of apoptosis causes many diseases. Thus, regulation of apoptotic process is very important. Drosophila tankyrase (DTNKS) is an evolutionarily conserved protein with poly (ADP-ribose) polymerase activity. In mammalian cells, tankyrases (TNKSs) have been reported to regulate cell death. To determine whether DTNKS plays function in inducing apoptosis in in vivo development, this study used Drosophila as a model system and generate transgenic flies expressing DTNKS. Ectopic expression of DTNKS promotes caspase-dependent apoptosis and knockdown of DTNKS by RNAi dramatically alleviates apoptotic defect caused by ectopic expression of pro-apoptotic proteins hid or rpr during eye development. Moreover, the result shows that ectopic expression of DTNKS triggers the activation of c-Jun N-terminal kinase (JNK) signaling, which is required for DTNKS-mediated apoptosis. Taken together, these findings have identified the role of DTNKS in regulating apoptosis by activating JNK signaling in Drosophila (Feng, 2018).

Wnt pathway activation by ADP-ribosylation

Wnt/β-catenin signal transduction directs metazoan development and is deregulated in numerous human congenital disorders and cancers. In the absence of Wnt stimulation, a multi-protein 'destruction complex', assembled by the scaffold protein Axin, targets the key transcriptional activator β-catenin for proteolysis. Axin is maintained at very low levels that limit destruction complex activity, a property that is currently being exploited in the development of novel therapeutics for Wnt-driven cancers. This study used an in vivo approach in Drosophila to determine how tightly basal Axin levels must be controlled for Wnt/Wingless pathway activation, and how Axin stability is regulated. For nearly all Wingless-driven developmental processes, a three- to four-fold increase in Axin was found to be insufficient to inhibit signaling, setting a lower-limit for the threshold level of Axin in the majority of in vivo contexts. Further, both the tumor suppressor Adenomatous polyposis coli (APC) and the ADP-ribose polymerase Tankyrase (Tnks) were found to have evolutionarily conserved roles in maintaining basal Axin levels below this in vivo threshold, and separable domains were defined in Axin that are important for APC- or Tnks-dependent destabilization. Together, these findings reveal that both APC and Tnks maintain basal Axin levels below a critical in vivo threshold to promote robust pathway activation following Wnt stimulation (Yang, 2016).

The Wnt/β-catenin signal transduction pathway directs fundamental processes during metazoan development and tissue homeostasis, whereas deregulation of Wnt signalling underlies numerous congenital disorders and carcinomas. Two multimeric protein complexes with opposing functions -- the cytoplasmic destruction complex and the plasma membrane-associated signalosome -- control the stability of the transcriptional co-factor β-catenin to coordinate the state of Wnt pathway activation. In the absence of Wnt stimulation, β-catenin is targeted for proteasomal degradation by the destruction complex, which includes the two tumour suppressors: Axin and Adenomatous polyposis coli (APC), and two kinases: casein kinase α (CK1α) and glycogen synthase kinase 3 (GSK3). Engagement of Wnt with its transmembrane receptors, Frizzled and low-density lipoprotein receptor-related protein 5/6 (herein LRP6), induces rapid LRP6 phosphorylation, recruitment of Axin to phospho-LRP6, and assembly of the signalosome, which includes two other Axin-associated components, GSK3 and Dishevelled (Dvl). Signalosome assembly results in the inhibition of β-catenin proteolysis; consequently stabilized β-catenin promotes the transcriptional regulation of Wnt pathway target genes (Yang, 2016).

As a key component in both the destruction complex and the signalosome, Axin is tightly regulated. Under basal conditions, Axin is maintained at very low levels, and serves as the concentration-limiting scaffold for assembly of the destruction complex. Following Wnt exposure, the rapid association of phospho-Axin with phospho-LRP6 triggers Axin dephosphorylation, inducing a conformational change that inhibits Axin's interaction with both the destruction and signalosome complexes. Axin is subsequently degraded; however, Axin proteolysis occurs several hours after Wnt exposure, and thus does not regulate Axin's essential role during the initial activation of the Wnt pathway (Yang, 2016).

The mechanisms that rapidly reprogram Axin from inhibitory to stimulatory roles following Wnt exposure remain uncertain. In current models, Wnt stimulation induces Axin's dissociation from the destruction complex, thereby promoting its interaction with the signalosome. As Wnt stimulation induces Axin dephosphorylation, decreased phosphorylation was postulated to facilitate the dissociation of Axin from the destruction complex; however, recent work revealed that the interaction of Axin with LRP6 precedes Axin dephosphorylation, and that dephosphorylation serves to inhibit, rather than enhance this interaction (Kim, 2013) Furthermore, some findings have challenged prevailing models, providing evidence that Axin's interaction with the destruction complex is not diminished upon Wnt stimulation. Thus, whereas the rapid switch in Axin function following Wnt stimulation is essential for the activation of signalling, the underlying mechanisms remain uncertain (Yang, 2016).

During investigation of this critical process, an unanticipated role was discovered for the ADP-ribose polymerase Tankyrase (Tnks) in the reprogramming of Axin activity following Wnt exposure. As Tnks-mediated ADP-ribosylation is known to target Axin for proteolysis, small molecule Tnks inhibitors have become lead candidates for development in the therapeutic targeting of Wnt-driven cancers. This study identified a novel mechanism through which Tnks regulates Axin: by promoting Axin's central role in rapid Wnt pathway activation. Wnt stimulation was found to modulate Axin levels biphasically in both Drosophila and human cells. Unexpectedly, Axin is rapidly stabilized following Wnt stimulation, before its ultimate proteolysis hours later. In an evolutionarily conserved process, the ADP-ribosylated pool of Axin is preferentially increased immediately following Wnt exposure. ADP-ribosylation enhances Axin's association with phospho-LRP6, providing a mechanistic basis for the rapid switch in Axin function following Wnt stimulation. These results thus indicate that Tnks inhibition not only increases basal Axin levels, but also impedes the Wnt-dependent interaction between Axin and LRP6, suggesting a basis for the potency of Tnks inhibitors in Wnt-driven cancers. Thus, Tnks not only targets Axin for proteolysis independently of Wnt stimulation, but also promotes Axin's central role in Wnt pathway activation, which may be relevant to the context-dependent activation of Wnt signalling and the treatment of Wnt-driven cancers with Tnks inhibitors (Yang, 2016).

Wnt exposure induces biphasic regulation in the level of Axin, and a large increase in the level of ADP-ribosylated Axin immediately after stimulation. ADP-ribosylation enhances the interaction of Axin with phospho-LRP6, and promotes the activation of Wnt signalling. These findings lead to three major revisions of the current model for the role of Tnks in the activation of the Wnt pathway. First, Tnks serves bifunctional roles under basal conditions and after stimulation, revealing a remarkable economy and coordination of pathway components. Second, the results provide a mechanistic basis for the rapid reprogramming of Axin function in response to Wnt stimulation, and thereby reveal an unanticipated role for Tnks in this process. These findings suggest that Wnt exposure either rapidly increases the ADP-ribosylation of Axin or inhibits the targeting of ADP-ribosylated Axin for proteasomal degradation, through mechanisms yet to be elucidated. Finally, pharmacologic inactivation of Tnks was shown to diminish the interaction of Axin with LRP6, revealing a previously unknown mechanism through which small molecule Tnks inhibitors disrupt Wnt signalling, distinct from their known role in stabilizing the destruction complex inhibitors (Yang, 2016).

In the absence of Wnt stimulation, the concentration-limiting levels of Axin regulate its scaffold function in the destruction complex. As components of the destruction complex participate in other signalling pathways, the low levels of Axin were proposed to maintain modularity of the Wnt pathway. The new findings indicate that Axin levels are not only regulated in the absence of Wnt, but also regulated biphasically following Wnt stimulation. This sequential modulation of Axin divides activation of the pathway into an early, fast phase and a delayed long-term phase. During embryogenesis, the earliest expression of Wg triggers the rapid appearance of Axin in segmental stripes, which is a novel hallmark for the initial activation of the pathway. The findings reveal that Wnt exposure induces a rapid increase in the total level of Axin, and importantly, a preferential increase in the level of the ADP-ribosylated Axin. The early Axin stripes are absent in Tnks null mutant embryos and are also absent when the Tnks binding domain in Axin is deleted. Therefore, it is proposed that Axin ADP-ribosylation contributes to Axin stabilization and to the rapid response to Wg stimulation (Yang, 2016).

It is postulated that the initial increase in levels of ADP-ribosylated Axin jump-starts the response to Wnt stimulation by enhancing the Axin-LRP6 interaction, whereas the subsequent decrease in Axin levels prolongs the duration of signalling by reducing destruction complex assembly. Thus, Wnt stimulation induces rapid increases in the levels of not only cytoplasmic β-catenin, but also ADP-ribosylated Axin. Previous work that coupled mathematical modelling with experimental analysis revealed that several Wnt signalling systems were responsive to the relative change in β-catenin levels, rather than their absolute value. This dependence was proposed to impart robustness and resistance to noise and cellular variation. The current data raise the possibility that a similar principle applies to changes in Axin levels on the Axin-LRP6 interaction, as the marked increase in ADP-ribosylated Axin levels following Wnt stimulation is evolutionarily conserved. Thus, the relative change in levels of ADP-ribosylated Axin may promote signalling following Wnt exposure by facilitating the fold change in β-catenin levels (Yang, 2016).

The current findings have relevance for the context-specific in vivo roles of Tnks in Wnt signalling suggested in previous studies. Tnks inhibition disrupts Wnt signalling in a number of cultured cell lines, but in vivo studies in several model organisms suggested that the requirement for Tnks in promoting Wnt signalling is restricted to specific cell types or developmental stages. In mice, functional redundancy exists between the two Tnks homologues, such that Tnks single mutants are viable and fertile, whereas double mutants display embryonic lethality without overt Wnt-related phenotypes. However, a missense mutation in the TBD of Axin2 that is predicted to disrupt ADP-ribosylation resulted in either activating or inhibiting effects on Wnt signalling that were dependent on developmental stage. Tnks inhibitors resulted in the same paradoxical effects, suggesting complex roles in mouse embryonic development. Analogously, treatment of fish with Tnks inhibitors resulted in no observed defects in Wnt-mediated processes during development; however, the regeneration of injured fins in adults, a process that requires Wnt signalling, was disrupted (Yang, 2016).

Similarly, the finding that Drosophila Tnks null mutants are viable (Wang, 2016a; Wang, 2016b; Feng, 2014) was unexpected, as Tnks is highly evolutionarily conserved, and no other Tnks homologues exist in fly genomes. Nonetheless, the current studies reveal that a less than twofold increase in Axin levels uncovers the importance of Tnks in promoting Wg signalling during embryogenesis. Therefore, it is postulated that Tnks loss can be compensated during development unless Axin levels are increased, but that the inhibition of Wg signalling resulting from Tnks inactivation cannot be attributed solely to increased Axin levels. Furthermore, Drosophila Tnks is essential for Wg target gene activation in the adult intestine, and exclusively within regions of the gradient where Wg is present at relatively low concentration. Thus, the context-specific roles of Tnks observed in different model organisms may reflect the mechanisms described in this study, which reveal that the Wnt-induced association of Axin with LRP6 occurs even in the absence of Axin ADP-ribosylation, but is markedly enhanced in its presence. It is postulated that by enhancing this interaction, Tnks-dependent ADP-ribosylation of Axin serves to amplify the initial response to Wnt stimulation, and thus is essential in a subset of in vivo contexts (Yang, 2016).

The recent discovery that Tnks enhances signalling in Wnt-driven cancers has raised the possibility that Tnks inhibitors will offer a promising new therapeutic option. Indeed, preclinical studies have supported this possibility. Tnks inhibitors were thought previously to disrupt Wnt signalling solely by increasing the basal levels of Axin, and thus by increasing destruction complex activity. However, the current findings indicate that the degree to which the basal level of Axin increases following Tnks inactivation is not sufficient to disrupt Wnt signalling in some in vivo contexts. Instead, the results reveal that Tnks inhibition simultaneously disrupts signalling at two critical and functionally distinct steps: by promoting activity of the destruction complex and by diminishing an important step in signalosome assembly: the Wnt-induced interaction between LRP6 and Axin. On the basis of these findings, it is proposed that the efficacy of Tnks inhibitors results from their combined action at both of these steps, providing a rationale for their use in the treatment of a broad range of Wnt-driven cancers. Therefore, these results suggest that in contrast with the current focus on tumours in which attenuation of the destruction complex aberrantly activates Wnt signalling (such as those lacking APC), the preclinical testing of Tnks inhibitors could be expanded to include cancers that are dependent on pathway activation by Wnt stimulation. These include the colorectal, gastric, ovarian and pancreatic cancers that harbour inactivating mutations in RNF43, a negative Wnt feedback regulator that promotes degradation of the Wnt co-receptors Frizzled and LRP6 (Yang, 2016).

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

The poly(ADP-ribose) polymerase (PARP) Tankyrase (TNKS and TNKS2) is paramount to Wnt-beta-catenin signaling and a promising therapeutic target in Wnt-dependent cancers. The pool of active beta-catenin is normally limited by destruction complexes, whose assembly depends on the polymeric master scaffolding protein AXIN. Tankyrase, which poly(ADP-ribosyl)ates and thereby destabilizes AXIN, also can polymerize, but the relevance of these polymers has remained unclear. This study reports crystal structures of the polymerizing TNKS and TNKS2 sterile alpha motif (SAM) domains, revealing versatile head-to-tail interactions. Biochemical studies informed by these structures demonstrate that polymerization is required for Tankyrase to drive beta-catenin-dependent transcription. The polymeric state supports PARP activity and allows Tankyrase to effectively access destruction complexes through enabling avidity-dependent AXIN binding. This study provides an example for regulated signal transduction in non-membrane-enclosed compartments (signalosomes), and it points to novel potential strategies to inhibit Tankyrase function in oncogenic Wnt signaling (Mariotti, 2016).

Wnt/Wingless Pathway Activation Is Promoted by a Critical Threshold of Axin Maintained by the Tumor Suppressor APC and the ADP-Ribose Polymerase Tankyrase

Wnt/β-catenin signal transduction directs metazoan development and is deregulated in numerous human congenital disorders and cancers. In the absence of Wnt stimulation, a multi-protein "destruction complex", assembled by the scaffold protein Axin, targets the key transcriptional activator β-catenin for proteolysis. Axin is maintained at very low levels that limit destruction complex activity, a property that is currently being exploited in the development of novel therapeutics for Wnt-driven cancers. This study used an in vivo approach in Drosophila to determine how tightly basal Axin levels must be controlled for Wnt/Wingless pathway activation, and how Axin stability is regulated. For nearly all Wingless-driven developmental processes, a three- to four-fold increase in Axin was found to be insufficient to inhibit signaling, setting a lower-limit for the threshold level of Axin in the majority of in vivo contexts. Further, both the tumor suppressor Adenomatous polyposis coli (APC) and the ADP-ribose polymerase Tankyrase (Tnks) were found to have evolutionarily conserved roles in maintaining basal Axin levels below this in vivo threshold, and separable domains were defined in Axin that are important for APC- or Tnks-dependent destabilization. Together, these findings reveal that both APC and Tnks maintain basal Axin levels below a critical in vivo threshold to promote robust pathway activation following Wnt stimulation (Wang, 2016).

The ADP-ribose polymerase Tankyrase regulates adult intestinal stem cell proliferation during homeostasis in Drosophila

Wnt/beta-catenin signaling controls intestinal stem cell (ISC) proliferation, and is aberrantly activated in colorectal cancer. Inhibitors of the ADP-ribose polymerase Tankyrase (Tnks) have become lead therapeutic candidates for Wnt-driven cancers, following the recent discovery that Tnks targets Axin, a negative regulator of Wnt signaling, for proteolysis. Initial reports indicated that Tnks is important for Wnt pathway activation in cultured human cell lines. However, the requirement for Tnks in physiological settings has been less clear, as subsequent studies in mice, fish and flies suggested that Tnks was either entirely dispensable for Wnt-dependent processes in vivo, or alternatively, had tissue-specific roles. Using null alleles, this study demonstrated that the regulation of Axin by the highly conserved Drosophila Tnks homolog is essential for the control of ISC proliferation. Furthermore, in the adult intestine, where activity of the Wingless pathway is graded and peaks at each compartmental boundary, Tnks is dispensable for signaling in regions where pathway activity is high, but essential where pathway activity is relatively low. Finally, as observed previously for Wingless pathway components, Tnks activity in absorptive enterocytes controls the proliferation of neighboring ISCs non-autonomously by regulating JAK/STAT signaling. These findings reveal the requirement for Tnks in the control of ISC proliferation and suggest an essential role in the amplification of Wnt signaling, with relevance for development, homeostasis and cancer (Wang, 2016b).

Dual roles for membrane association of Drosophila Axin in Wnt signaling

Axin, a concentration-limiting scaffold protein, facilitates assembly of a "destruction complex" that prevents Wnt signaling in the unstimulated state and a plasma membrane-associated "signalosome" that activates signaling following Wnt stimulation. In the classical model, Axin is cytoplasmic under basal conditions, but relocates to the cell membrane after Wnt exposure. This study analyzed the subcellular distribution of endogenous Drosophila Axin in vivo and found that a pool of Axin localizes to cell membrane proximal puncta even in the absence of Wnt stimulation. Axin localization in these puncta is dependent on the destruction complex component Adenomatous polyposis coli (Apc). In the unstimulated state, the membrane association of Axin increases its Tankyrase-dependent ADP-ribosylation and consequent proteasomal degradation to control its basal levels. Furthermore, Wnt stimulation does not result in a bulk redistribution of Axin from cytoplasmic to membrane pools, but causes an initial increase of Axin in both of these pools, with concomitant changes in two post-translational modifications, followed by Axin proteolysis hours later. Finally, the ADP-ribosylated Axin that increases rapidly following Wnt stimulation is membrane associated. The study concludes that even in the unstimulated state, a pool of Axin forms membrane-proximal puncta that are dependent on Apc, and that membrane association regulates both Axin levels and Axin's role in the rapid activation of signaling that follows Wnt exposure (Wang, 2016).

Polymerase Enzyme Tankyrase Antagonizes Activity of the beta-Catenin Destruction Complex through ADP-ribosylation of Axin and APC2

Most colon cancer cases are initiated by truncating mutations in the tumor suppressor, adenomatous polyposis coli (APC). APC is a critical negative regulator of the Wnt signaling pathway that participates in a multi-protein "destruction complex" to target the key effector protein beta-catenin for ubiquitin-mediated proteolysis. Prior work has established that the poly(ADP-ribose) polymerase (PARP) enzyme Tankyrase (TNKS) antagonizes destruction complex activity by promoting degradation of the scaffold protein Axin, and recent work suggests that TNKS inhibition is a promising cancer therapy. This study performed a yeast two-hybrid (Y2H) screen and uncovered TNKS as a putative binding partner of Drosophila APC2, suggesting that TNKS may play multiple roles in destruction complex regulation. TNKS binds a C-terminal RPQPSG motif in Drosophila APC2, and this motif is conserved in human APC2, but not human APC1. In addition, it was found that APC2 can recruit TNKS into the beta-catenin destruction complex, placing the APC2/TNKS interaction at the correct intracellular location to regulate beta-catenin proteolysis. It was further shown that TNKS directly PARylates both Drosophila Axin and APC2, but that PARylation does not globally regulate APC2 protein levels as it does for Axin. Moreover, TNKS inhibition in colon cancer cells decreases beta-catenin signaling, cannot be explained solely through Axin stabilization. Instead, these findings suggest that TNKS regulates destruction complex activity at the level of both Axin and APC2, providing further mechanistic insight into TNKS inhibition as a potential Wnt pathway cancer therapy (Croy, 2016).

The Drosophila tankyrase regulates Wg signaling depending on the concentration of Daxin

The canonical Wnt signaling pathway plays critical roles during development and homeostasis. Dysregulation of this pathway can lead to many human diseases, including cancers. A key process in this pathway consists of regulation of beta-catenin concentration through an Axin-recruited destruction complex. Previous studies have demonstrated a role for Tankyrase (TNKS), a protein with poly (ADP-ribose) polymerase, in the regulation of Axin levels in human cancers. However, the role of TNKS in development is still unclear. This study generated a Drosophila tankyrase (DTNKS) mutant and provided compelling evidence that DTNKS is involved in the degradation of Drosophila Axin (Daxin). Daxin was shown to physically interact with DTNKS, and its protein levels are elevated in the absence of DTNKS in eye discs. In S2 cells, DTNKS suppressed the levels of Daxin. Surprisingly, it was found that Daxin in turn down-regulated DTNKS protein level. In vivo study showed that DTNKS regulated Wg signaling and wing patterning at a high Daxin protein level, but not at a normal level. Taken together, these findings identified a conserved role of DTNKS in regulating Daxin levels, and thereby Wg (Feng, 2014).

Proteasome regulation by ADP-ribosylation

Protein degradation by the ubiquitin-proteasome system is central to cell homeostasis and survival. Defects in this process are associated with diseases such as cancer and neurodegenerative disorders. The 26S proteasome is a large protease complex that degrades ubiquitinated proteins. This study shows that ADP-ribosylation promotes 26S proteasome activity in both Drosophila and human cells. The ADP-ribosyltransferase tankyrase (TNKS) and the 19S assembly chaperones dp27 and dS5b were identified as direct binding partners of the proteasome regulator PI31. TNKS-mediated ADP-ribosylation of PI31 drastically reduces its affinity for 20S proteasome alpha subunits to relieve 20S repression by PI31. Additionally, PI31 modification increases binding to and sequestration of dp27 and dS5b from 19S regulatory particles, promoting 26S assembly. Inhibition of TNKS by either RNAi or a small-molecule inhibitor, XAV939, blocks this process to reduce 26S assembly. These results unravel a mechanism of proteasome regulation that can be targeted with existing small-molecule inhibitors (Cho-Park, 2013).

Drosophila poly(ADP-ribose) glycohydrolase mediates chromatin structure and SIR2-dependent silencing

Protein ADP ribosylation catalyzed by cellular poly(ADP-ribose) polymerases (PARPs) and tankyrases modulates chromatin structure, telomere elongation, DNA repair, and the transcription of genes involved in stress resistance, hormone responses, and immunity. Using Drosophila genetic tools, this study characterized the expression and function of poly(ADP-ribose) glycohydrolase (PARG), the primary enzyme responsible for degrading protein-bound ADP-ribose moieties. Strongly increasing or decreasing PARG levels mimics the effects of Parp mutation, supporting PARG's postulated roles in vivo both in removing ADP-ribose adducts and in facilitating multiple activity cycles by individual PARP molecules. PARP is largely absent from euchromatin in PARG mutants, but accumulates in large nuclear bodies that may be involved in protein recycling. Reducing the level of either PARG or the silencing protein SIR2 weakens copia transcriptional repression. In the absence of PARG, SIR2 is mislocalized and hypermodified. It is proposed that PARP and PARG promote chromatin silencing at least in part by regulating the localization and function of SIR2 and possibly other nuclear proteins (Tulin, 2006).

Tankyrase-mediated ADP-ribosylation is a regulator of TNF-induced death

Tumor necrosis factor (TNF) is a key component of the innate immune response. Upon binding to its receptor, TNFR1, it promotes production of other cytokines via a membrane-bound complex 1 or induces cell death via a cytosolic complex 2. To understand how TNF-induced cell death is regulated, mass spectrometry was performed of complex 2 and tankyrase-1 was identified as a native component that, upon a death stimulus, mediates complex 2 poly-ADP-ribosylation (PARylation). PARylation promotes recruitment of the E3 ligase RNF146, resulting in proteasomal degradation of complex 2, thereby limiting cell death. Expression of the ADP-ribose-binding/hydrolyzing severe acute respiratory syndrome coronavirus 2 macrodomain sensitizes cells to TNF-induced death via abolishing complex 2 PARylation. This suggests that disruption of ADP-ribosylation during an infection can prime a cell to retaliate with an inflammatory cell death (Liu, 2002).

Loss of ATRX suppresses resolution of telomere cohesion to control recombination in ALT cancer cells

The chromatin-remodeler ATRX is frequently lost in cancer cells that use ALT (alternative lengthening of telomeres) for telomere maintenance, but its function in telomere recombination is unknown. This study shows that loss of ATRX suppresses the timely resolution of sister telomere cohesion that normally occurs prior to mitosis. In the absence of ATRX, the histone variant macroH2A1.1 binds to the poly(ADP-ribose) polymerase tankyrase 1, preventing it from localizing to telomeres and resolving cohesion. The resulting persistent telomere cohesion promotes recombination between sister telomeres, while it suppresses inappropriate recombination between non-sisters. Forced resolution of sister telomere cohesion induces excessive recombination between non-homologs, genomic instability, and impaired cell growth, indicating the ATRX-macroH2A1.1-tankyrase axis as a potential therapeutic target in ALT tumors (Ramamoorthy, 2015).

Tankyrase Regulates Neurite Outgrowth through Poly(ADP-ribosyl)ation-Dependent Activation of beta-Catenin Signaling

Poly(ADP-ribosyl)ation is a post-translational modification of proteins by transferring poly(ADP-ribose) (PAR) to acceptor proteins by the action of poly(ADP-ribose) polymerase (PARP). Two tankyrase (TNKS) isoforms, TNK1 and TNK2 (TNKS1/2), are ubiquitously expressed in mammalian cells and participate in diverse cellular functions, including wnt/beta-catenin signaling, telomere maintenance, glucose metabolism and mitosis regulation. For wnt/beta-catenin signaling, TNKS1/2 catalyze poly(ADP-ribosyl)ation of Axin, a key component of the beta-catenin degradation complex, which allows Axin's ubiquitination and subsequent degradation, thereby activating beta-catenin signaling. This study focused on the functions of TNKS1/2 in neuronal development. In primary hippocampal neurons, TNKS1/2 were detected in the soma and neurites, where they co-localized with PAR signals. Treatment with XAV939, a selective TNKS1/2 inhibitor, suppressed neurite outgrowth and synapse formation. In addition, XAV939 also suppressed norepinephrine uptake in PC12 cells, a rat pheochromocytoma cell line. These effects likely resulted from the inhibition of beta-catenin signaling through the stabilization of Axin, which suggests TNKS1/2 enhance Axin degradation by modifying its poly(ADP-ribosyl)ation, thereby stabilizing wnt/beta-catenin signaling and, in turn, promoting neurite outgrowth and synapse formation (Mashimo, 2022).

The zinc-binding motif in tankyrases is required for the structural integrity of the catalytic ADP-ribosyltransferase domain

Tankyrases are ADP-ribosylating enzymes that regulate many physiological processes in the cell and are considered promising drug targets for cancer and fibrotic diseases. The catalytic ADP-ribosyltransferase domain of tankyrases contains a unique zinc-binding motif of unknown function. Recently, this motif was suggested to be involved in the catalytic activity of tankyrases. This work was set out to study the effect of the zinc-binding motif on the activity, stability and structure of human tankyrases. Mutants of human tankyrase (TNKS) 1 and TNKS2 were generated, abolishing the zinc-binding capabilities, and characterized the proteins biochemically and biophysically in vitro. A crystal structure of TNKS2 was generated, in which the zinc ion was oxidatively removed. This work shows that the zinc-binding motif in tankyrases is a crucial structural element which is particularly important for the structural integrity of the acceptor site. While mutation of the motif rendered TNKS1 inactive, probably due to introduction of major structural defects, the TNKS2 mutant remained active and displayed an altered activity profile compared to the wild-type (Sowa, 2022).

Tankyrase-1-mediated degradation of Golgin45 regulates glycosyltransferase trafficking and protein glycosylation in Rab2-GTP-dependent manner

Altered glycosylation plays an important role during development and is also a hallmark of increased tumorigenicity and metastatic potentials of several cancers. This study reports that Tankyrase-1 (TNKS1) controls protein glycosylation by Poly-ADP-ribosylation (PARylation) of a Golgi structural protein, Golgin45, at the Golgi. TNKS1 is a Golgi-localized peripheral membrane protein that plays various roles throughout the cell, ranging from telomere maintenance to Glut4 trafficking. This study indicates that TNKS1 localization to the Golgi apparatus is mediated by Golgin45. TNKS1-dependent control of Golgin45 protein stability influences protein glycosylation, as shown by Glycomic analysis. Further, FRAP experiments indicated that Golgin45 protein level modulates Golgi glycosyltransferease trafficking in Rab2-GTP-dependent manner. Taken together, these results suggest that TNKS1-dependent regulation of Golgin45 may provide a molecular underpinning for altered glycosylation at the Golgi during development or oncogenic transformation (Yue, 2021).

Search PubMed for articles about Drosophila Tankyrase

Cho-Park, P. F. and Steller, H. (2013). Proteasome regulation by ADP-ribosylation. Cell 153(3): 614-627. PubMed ID: 23622245

Croy, H. E., Fuller, C. N., Giannotti, J., Robinson, P., Foley, A. V. A., Yamulla, R. J., Cosgriff, S., Greaves, B. D., von Kleeck, R. A., An, H. H., Powers, C. M., Tran, J. K., Tocker, A. M., Jacob, K. D., Davis, B. K. and Roberts, D. M. (2016). The Poly(ADP-ribose) Polymerase Enzyme Tankyrase Antagonizes Activity of the beta-Catenin Destruction Complex through ADP-ribosylation of Axin and APC2. J Biol Chem 291(24): 12747-12760. PubMed ID: 27068743

Feng, Y., Li, Z., Lv, L., Du, A., Lin, Z., Ye, X., Lin, Y. and Lin, X. (2018). Tankyrase regulates apoptosis by activating JNK signaling in Drosophila. Biochem Biophys Res Commun 503(4): 2234-2239. PubMed ID: 29953853

Gultekin, Y. and Steller, H. (2019). Axin proteolysis by Iduna is required for the regulation of stem cell proliferation and intestinal homeostasis in Drosophila. Development 146(6). PubMed ID: 30796047

Li, P., Huang, P., Li, X., Yin, D., Ma, Z., Wang, H. and Song, H. (2018). Tankyrase mediates K63-linked ubiquitination of JNK to confer stress tolerance and influence lifespan in Drosophila. Cell Rep 25(2): 437-448. PubMed ID: 30304683

Liu, L., Sandow, J. J,, ..., Lalaoui, N., Silke, J. (2022). Tankyrase-mediated ADP-ribosylation is a regulator of TNF-induced death. Sci Adv. 8(19):eabh2332. PubMed ID: 35544574

Mariotti, L., Templeton, C. M., Ranes, M., Paracuellos, P., Cronin, N., Beuron, F., Morris, E. and Guettler, S. (2016). Tankyrase Requires SAM Domain-Dependent Polymerization to Support Wnt-beta-Catenin Signaling. Mol Cell 63(3): 498-513. PubMed ID: 27494558

Mashimo, M., Kita, M., Uno, A., Nii, M., Ishihara, M., Honda, T., Gotoh-Kinoshita, Y., Nomura, A., Nakamura, H., Murayama, T., Kizu, R. and Fujii, T. (2022). Tankyrase Regulates Neurite Outgrowth through Poly(ADP-ribosyl)ation-Dependent Activation of beta-Catenin Signaling. Int J Mol Sci 23(5). PubMed ID: 35269974

McGurk, L., Gomes, E., Guo, L., Mojsilovic-Petrovic, J., Tran, V., Kalb, R. G., Shorter, J. and Bonini, N. M. (2018). Poly(ADP-Ribose) Prevents Pathological Phase Separation of TDP-43 by Promoting Liquid Demixing and Stress Granule Localization. Mol Cell 71(5): 703-717. PubMed ID: 30100264

Ramamoorthy, M. and Smith, S. (2015). Loss of ATRX Suppresses Resolution of Telomere Cohesion to Control Recombination in ALT Cancer Cells. Cancer Cell 28(3): 357-369. PubMed ID: 26373281

Sowa, S. T. and Lehtio, L. (2022). The zinc-binding motif in tankyrases is required for the structural integrity of the catalytic ADP-ribosyltransferase domain. Open Biol 12(3): 210365. PubMed ID: 35317661

Tulin, A., Naumova, N. M., Menon, A. K. and Spradling, A. C. (2006). Drosophila poly(ADP-ribose) glycohydrolase mediates chromatin structure and SIR2-dependent silencing. Genetics 172(1): 363-371. PubMed ID: 16219773

Vilchez Larrea, S., Valsecchi, W. M., Fernandez Villamil, S. H. and Lafon Hughes, L. I. (2021). First body of evidence suggesting a role of a tankyrase-binding motif (TBM) of vinculin (VCL) in epithelial cells. PeerJ 9: e11442. PubMed ID: 34123588

Wang, Z., Tacchelly-Benites, O., Yang, E. and Ahmed, Y. (2016a). Dual roles for membrane association of Drosophila Axin in Wnt signaling. PLoS Genet 12: e1006494. PubMed ID: 27959917

Wang, Z., Tacchelly-Benites, O., Yang, E., Thorne, C. A., Nojima, H., Lee, E. and Ahmed, Y. (2016a). Wnt/Wingless Pathway Activation Is Promoted by a Critical Threshold of Axin Maintained by the Tumor Suppressor APC and the ADP-Ribose Polymerase Tankyrase. Genetics 203(1): 269-281. PubMed ID: 26975665

Wang, Z., Tian, A., Benchabane, H., Tacchelly-Benites, O., Yang, E., Nojima, H. and Ahmed, Y. (2016b). The ADP-ribose polymerase Tankyrase regulates adult intestinal stem cell proliferation during homeostasis in Drosophila. Development 143: 1710-1720. PubMed ID: 27190037

Wang, Z., Tacchelly-Benites, O., Noble, G. P., Johnson, M. K., Gagne, J. P., Poirier, G. G. and Ahmed, Y. (2019). A Context-Dependent Role for the RNF146 Ubiquitin Ligase in Wingless/Wnt Signaling in Drosophila. Genetics 211(3): 913-923. PubMed ID: 30593492

Yang, E., Tacchelly-Benites, O., Wang, Z., Randall, M. P., Tian, A., Benchabane, H., Freemantle, S., Pikielny, C., Tolwinski, N. S., Lee, E. and Ahmed, Y. (2016). Wnt pathway activation by ADP-ribosylation. Nat Commun 7: 11430. PubMed ID: 27138857

Yue, X., Tiwari, N., Zhu, L., Ngo, H. D. T., Lim, J. M., Gim, B., Jing, S., Wang, Y., Qian, Y. and Lee, I. (2021). Tankyrase-1-mediated degradation of Golgin45 regulates glycosyltransferase trafficking and protein glycosylation in Rab2-GTP-dependent manner. Commun Biol 4(1): 1370. PubMed ID: 34876695

date revised: 10 May 2022

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