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Key Points: Loss of β-catenin impairs in vivo and isolated muscle exercise/contraction-stimulated glucose uptake. β-Catenin is required for exercise-induced skeletal muscle actin cytoskeleton remodelling. β-Catenin 675 phosphorylation during exercise may be intensity dependent., Abstract: The conserved structural protein β-catenin is an emerging regulator of vesicle trafficking in multiple tissues and supports insulin-stimulated glucose transporter 4 (GLUT4) translocation in skeletal muscle by facilitating cortical actin remodelling. Actin remodelling may be a convergence point between insulin and exercise/contraction-stimulated glucose uptake. Here we investigated whether β-catenin is involved in regulating exercise/contraction-stimulated glucose uptake. We report that the muscle-specific deletion of β-catenin induced in adult mice (BCAT-mKO) impairs both exercise- and contraction (isolated muscle)-induced glucose uptake without affecting running performance or canonical exercise signalling pathways. Furthermore, high intensity exercise in mice and contraction of myotubes and isolated muscles led to the phosphorylation of β-catenin S675 , and this was impaired by Rac1 inhibition. Moderate intensity exercise in control and Rac1 muscle-specific knockout mice did not induce muscle β-catenin S675 phosphorylation, suggesting exercise intensity-dependent regulation of β-catenin S675 . Introduction of a non-phosphorylatable S675A mutant of β-catenin into myoblasts impaired GLUT4 translocation and actin remodelling stimulated by carbachol, a Rac1 and RhoA activator. Exercise-induced increases in cross-sectional phalloidin staining (F-actin marker) of gastrocnemius muscle was impaired in muscle from BCAT-mKO mice. Collectively our findings suggest that β-catenin is required for optimal glucose transport in muscle during exercise/contraction, potentially via facilitating actin cytoskeleton remodelling., (© 2021 The Authors. The Journal of Physiology © 2021 The Physiological Society.)

The presence of adherens junctions and the associated protein β-catenin are requirements for the development of glucose-stimulated insulin secretion (GSIS) in β-cells. Evidence indicates that modulation of β-catenin function in response to changes in glucose levels can modulate the levels of insulin secretion from β-cells but the role of β-catenin phosphorylation in this process has not been established. We find that a Ser552Ala version of β-catenin attenuates glucose-stimulated insulin secretion indicating a functional role for Ser552 phosphorylation of β-catenin in insulin secretion. This is associated with alterations F/G actin ratio but not the transcriptional activity of β-catenin. Both glucose and GLP-1 stimulated phosphorylation of the serine 552 residue on β-catenin. We investigated the possibility that an EPAC-PAK1 pathway might be involved in this phosphorylation event. We find that reduction in PAK1 levels using siRNA attenuates both glucose and GLP-1 stimulated phosphorylation of β-catenin Ser552 and the effects of these on insulin secretion in β-cell models. Furthermore, both the EPAC inhibitor ESI-09 and the PAK1 inhibitor IPA3 do the same in both β-cell models and mouse islets. Together this identifies phosphorylation of β-catenin at Ser552 as part of a cell signalling mechanism linking nutrient and hormonal regulation of β-catenin to modulation of insulin secretory capacity of β-cells and indicates this phosphorylation event is regulated downstream of EPAC and PAK1 in β-cells., (© 2021 The Author(s).)

Objective: Skeletal muscle glucose disposal following a meal is mediated through insulin-stimulated movement of the GLUT4-containing vesicles to the cell surface. The highly conserved scaffold-protein β-catenin is an emerging regulator of vesicle trafficking in other tissues. Here, we investigated the involvement of β-catenin in skeletal muscle insulin-stimulated glucose transport., Methods: Glucose homeostasis and transport was investigated in inducible muscle specific β-catenin knockout (BCAT-mKO) mice. The effect of β-catenin deletion and mutation of β-catenin serine 552 on signal transduction, glucose uptake and protein-protein interactions were determined in L6-G4-myc cells, and β-catenin insulin-responsive binding partners were identified via immunoprecipitation coupled to label-free proteomics., Results: Skeletal muscle specific deletion of β-catenin impaired whole-body insulin sensitivity and insulin-stimulated glucose uptake into muscle independent of canonical Wnt signalling. In response to insulin, β-catenin was phosphorylated at serine 552 in an Akt-dependent manner, and in L6-G4-myc cells, mutation of β-catenin S552 impaired insulin-induced actin-polymerisation, resulting in attenuated insulin-induced glucose transport and GLUT4 translocation. β-catenin was found to interact with M-cadherin in an insulin-dependent β-catenin S552 -phosphorylation dependent manner, and loss of M-cadherin in L6-G4-myc cells attenuated insulin-induced actin-polymerisation and glucose transport., Conclusions: Our data suggest that β-catenin is a novel mediator of glucose transport in skeletal muscle and may contribute to insulin-induced actin-cytoskeleton remodelling to support GLUT4 translocation., (Copyright © 2020 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

GLUT4 is unique among specialized glucose transporters in being exclusively expressed in muscle and adipocytes. In the absence of insulin the distribution of GLUT4 is preferentially intracellular and insulin stimulation results in the movement of GLUT4 containing vesicles to the plasma membrane. This process is responsible for the insulin stimulation of glucose uptake in muscle and fat. While signalling pathways triggering the translocation of GLUT4 are well understood, the mechanisms regulating the intracellular retention of GLUT4 are less well understood. Here we report a role for β-catenin in this process. In 3T3-L1 adipocytes in which β-catenin is depleted, the levels of GLUT4 at and near the plasma membrane rise in unstimulated cells while the subsequent increase in GLUT4 at the plasma membrane upon insulin stimulation is reduced. Small molecule approaches to acutely activate or inhibit β-catenin give results that support the results obtained with siRNA and these changes are accompanied by matching changes in glucose transport into these cells. Together these results indicate that β-catenin is a previously unrecognized regulator of the mechanisms that control the insulin sensitive pool of GLUT4 transporters inside these adipocyte cells., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The processes regulating glucose-stimulated insulin secretion (GSIS) and its modulation by incretins in pancreatic β-cells are only partly understood. Here we investigate the involvement of β-catenin in these processes. Reducing β-catenin levels using siRNA knockdown attenuated GSIS in a range of β-cell models and blocked the ability of GLP-1 agonists and the depolarizing agent KCl to potentiate this. This could be mimicked in both β-cell models and isolated islets by short-term exposure to the β-catenin inhibitory drug pyrvinium. In addition, short-term treatment with a drug that increases β-catenin levels results in an increase in insulin secretion. The timing of these effects suggests that β-catenin is required for the processes regulating trafficking and/or release of pre-existing insulin granules rather than for those regulated by gene expression. This was supported by the finding that the overexpression of the transcriptional co-activator of β-catenin, transcription factor 7-like 2 (TCF7L2), attenuated insulin secretion, consistent with the extra TCF7L2 translocating β-catenin from the plasma membrane pool to the nucleus. We show that β-catenin depletion disrupts the intracellular actin cytoskeleton, and by using total internal reflectance fluorescence (TIRF) microscopy, we found that β-catenin is required for the glucose- and incretin-induced depletion of insulin vesicles from near the plasma membrane. In conclusion, we find that β-catenin levels modulate Ca 2+ -dependent insulin exocytosis under conditions of glucose, GLP-1, or KCl stimulation through a role in modulating insulin secretory vesicle localization and/or fusion via actin remodeling. These findings also provide insights as to how the overexpression of TCF7L2 may attenuate insulin secretion., (© 2016 by The American Society for Biochemistry and Molecular Biology, Inc.)

Recently, it has been found that glucagon is able to activate the β-catenin signalling pathway leading to increased cyclin D1 and c-Myc expression in liver. Therefore the main aim of the present study is to determine whether the effect of glucagon activating β-catenin signalling leading to increased target gene expression is mediated through cAMP activation of PKA (protein kinase A). Primary rat hepatocytes were incubated with insulin, glucagon or adrenaline (epinephrine) and a range of inhibitors of PI3K (phosphoinositide 3-kinase), Wnt, mitochondrial uncoupler (niclosamide) or PKA inhibitors to dissect out the pathway leading to increased Ser(552) phosphorylation on β-catenin following glucagon exposure. In primary rat hepatocytes, we found that short exposure to glucagon or adrenaline caused a rapid increase in Ser(552) phosphorylation on β-catenin that leads to increased cyclin D1 and c-Myc expression. A range of PI3K and Wnt inhibitors were unable to block the effect of glucagon phosphorylating β-catenin. Interestingly, both niclosamide and the PKA inhibitor H89 blocked the glucagon effect on β-catenin signalling, leading to a reduction in target gene expression. Likewise, niclosamide inhibited cAMP levels and the direct addition of db-cAMP (dibutyryl-cAMP sodium salt) also resulted in Ser(552) phosphorylation of β-catenin. We have identified a new pathway via glucagon signalling that leads to increased β-catenin activity that can be reversed with the antihelminthic drug niclosamide, which has recently shown promise as a potential treatment of T2D (Type 2 diabetes). This novel finding could be useful in liver cancer treatment, particularly in the context of T2D with increased β-catenin activity., (© 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.)

In the last 20 years the prevalence of metabolic disorders, in particular type 2 diabetes (T2D), has more than doubled. Recently, a strong link between T2D and cancer, in particularly liver cancer has been reported. However, the mechanism connecting the development of type 2 diabetes and cancer remains unknown. One of the biggest drivers of liver cancer is alterations in the Wnt/β-catenin pathway. In this study, we aimed to identify the effect of glucagon on β-catenin in the isolated rat liver. We found glucagon, which is substantially raised in patients with T2D, rapidly phosphorylates β-catenin on serine 552 that is associated with increased expression of genes cyclin D1 (CCND1) and c-Myc (MYC), which are known to be involved in liver cancer. This finding may explain the increased risk of liver cancer in people with T2D.

Adiponectin is an adipokine that can suppress the proliferation of various human carcinoma cells. Although its anti-tumor activities have been suggested by many clinical investigations and animal studies, the underlying mechanisms are not fully characterized. In MMTV-polyomavirus middle T antigen (MMTV-PyVT) transgenic mice models, reduced- or complete loss-of-adiponectin expression promotes mammary tumor development. The present study demonstrated that while tumor development in control MMTV-PyVT mice is associated with a progressively decreased circulating cholesterol concentration, adiponectin deficient MMTV-PyVT mice showed significantly elevated total- and low density lipoprotein (LDL)-cholesterol levels. Cholesterol contents in tumors derived from adiponectin deficient mice were dramatically augmented. High fat high cholesterol diet further accelerated the tumor development in adiponectin deficient PyVT mice. The protein levels of LDL receptor (LDLR) were found to be upregulated in adiponectin-deficient tumor cells. In human breast carcinoma cells, treatment with LDL-cholesterol or overexpressing LDLR elevates nuclear beta-catenin activity and facilitates tumor cell proliferation. On the other hand, adiponectin decreased LDLR protein expression in breast cancer cells and inhibited LDL-cholesterol-induced tumor cell proliferation. Both in vivo and in vitro evidence demonstrated a stimulatory effect of adiponectin on autophagy process, which mediated the down-regulation of LDLR. Adiponectin-induced reduction of LDLR was blocked by treatment with a specific inhibitor of autophagy, 3-methyladenine. In conclusion, the study demonstrates that adiponectin elicits tumor suppressive effects by modulating cholesterol homeostasis and LDLR expression in breast cancer cells, which is at least in part attributed to its role in promoting autophagic flux.

Pancreatic β-cells are highly responsive to changes in glucose, but the mechanisms involved are only partially understood. There is increasing evidence that the β-catenin signalling pathway plays an important role in regulating β-cell function, but the mechanisms regulating β-catenin signalling in these cells is not well understood. In the present study we show that β-catenin levels and downstream signalling are regulated by changes in glucose levels in INS-1E and β-TC6-F7 β-cell models. We found a glucose-dependent increase in levels of β-catenin in the cytoplasm and nucleus of INS-1E cells. Expression of cyclin D1 also increased with glucose and required the presence of β-catenin. This was associated with an increase in phosphorylation of β-catenin on Ser552, which is known to stabilize the molecule and increase its transcriptional activity. In a search for possible signalling intermediates we found forskolin and cell-permeable cAMP analogues recapitulated the glucose effects, suggesting a role for cAMP and PKA (cAMP-dependent protein kinase/protein kinase A) downstream of glucose. Furthermore, glucose caused sustained increases in cAMP. Two different inhibitors of adenylate cyclase and PKA signalling blocked the effects of glucose, whereas siRNA (small interfering RNA) knockdown of PKA blocked the effects of glucose on β-catenin signalling. Finally, reducing β-catenin levels with either siRNA or pyrvinium impaired glucose- and KCl-stimulated insulin secretion. Taken together the results of the present study define a pathway by which changes in glucose levels can regulate β-catenin using a mechanism which involves cAMP production and the activation of PKA. This identifies a pathway that may be important in glucose-dependent regulation of gene expression and insulin secretion in β-cells.