O-GlcNAcylation of RBM14 contributes to elevated cellular O-GlcNAc through regulation of OGA protein stability.

Dysregulation of O-GlcNAcylation has emerged as a potential biomarker for several diseases, particularly cancer. The role of OGT (O-GlcNAc transferase) in maintaining O-GlcNAc homeostasis has been extensively studied; nevertheless, the regulation of OGA (O-GlcNAcase) in cancer remains elusive. Here, we demonstrated that the multifunctional protein RBM14 is a regulator of cellular O-GlcNAcylation. By investigating the correlation between elevated O-GlcNAcylation and increased RBM14 expression in lung cancer cells, we discovered that RBM14 promotes ubiquitin-dependent proteasomal degradation of OGA, ultimately mediating cellular O-GlcNAcylation levels. In addition, RBM14 itself is O-GlcNAcylated at serine 521, regulating its interaction with the E3 ligase TRIM33, consequently affecting OGA protein stability. Moreover, we demonstrated that mutation of serine 521 to alanine abrogated the oncogenic properties of RBM14. Collectively, our findings reveal a previously unknown mechanism for the regulation of OGA and suggest a potential therapeutic target for the treatment of cancers with dysregulated O-GlcNAcylation.

A surge of cytosolic calcium dysregulates lysosomal function and impairs autophagy flux during cupric chloride-induced neuronal death.

Autophagy is a degradative pathway that plays an important role in maintaining cellular homeostasis. Dysfunction of autophagy is associated with the progression of neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, and amyotrophic lateral sclerosis. Although one of the typical features of brain aging is an accumulation of redox-active metals that eventually lead to neurodegeneration, a plausible link between trace metal-induced neurodegeneration and dysregulated autophagy has not been clearly determined. Here, we used a cupric chloride-induced neurodegeneration model in MN9D dopaminergic neuronal cells along with ultrastructural and biochemical analyses to demonstrate impaired autophagic flux with accompanying lysosomal dysfunction. We found that a surge of cytosolic calcium was involved in cupric chloride-induced dysregulated autophagy. Consequently, buffering of cytosolic calcium by calbindin-D28K overexpression or co-treatment with the calcium chelator BAPTA attenuated the cupric chloride-induced impairment in autophagic flux by ameliorating dysregulation of lysosomal function. Thus, these events allowed the rescue of cells from cupric chloride-induced neuronal death. These phenomena were largely confirmed in cupric chloride-treated primary cultures of cortical neurons. Taken together, these results suggest that abnormal accumulation of trace metal elements and a resultant surge of cytosolic calcium leads to neuronal death by impairing autophagic flux at the lysosomal level.

SETD5 regulates the OGT-catalyzed O-GlcNAcylation of RNA polymerase II, which is involved in the stemness of colorectal cancer cells.

The dosage-dependent recruitment of RNA polymerase II (Pol II) at the promoters of genes related to neurodevelopment and stem cell maintenance is required for transcription by the fine-tuned expression of SET-domain-containing protein 5 (SETD5). Pol II O-GlcNAcylation by O-GlcNAc transferase (OGT) is critical for preinitiation complex formation and transcription cycling. SETD5 dysregulation has been linked to stem cell-like properties in some cancer types; however, the role of SETD5 in cancer cell stemness has not yet been determined. We here show that aberrant SETD5 overexpression induces stemness in colorectal cancer (CRC) cells. SETD5 overexpression causes the upregulation of PI3K-AKT pathway-related genes and cancer stem cell (CSC) markers such as CD133, Kruppel-like factor 4 (KLF4), and estrogen-related receptor beta (ESRRB), leading to the gain of stem cell-like phenotypes. Our findings also revealed a functional relationship between SETD5, OGT, and Pol II. OGT-catalyzed Pol II glycosylation depends on SETD5, and the SETD5-Pol II interaction weakens in OGT-depleted cells, suggesting a SETD5-OGT-Pol II interdependence. SETD5 deficiency reduces Pol II occupancy at PI3K-AKT pathway-related genes and CD133 promoters, suggesting a role for SETD5-mediated Pol II recruitment in gene regulation. Moreover, the SETD5 depletion nullified the SETD5-induced stemness of CRC cells and Pol II O-GlcNAcylation. These findings support the hypothesis that SETD5 mediates OGT-catalyzed O-GlcNAcylation of RNA Pol II, which is involved in cancer cell stemness gain via CSC marker gene upregulation.

Innate mechanism of mucosal barrier erosion in the pathogenesis of acquired colitis.

The colonic mucosal barrier protects against infection, inflammation, and tissue ulceration. Composed primarily of Mucin-2, proteolytic erosion of this barrier is an invariant feature of colitis; however, the molecular mechanisms are not well understood. We have applied a recurrent food poisoning model of acquired inflammatory bowel disease using Salmonella enterica Typhimurium to investigate mucosal barrier erosion. Our findings reveal an innate Toll-like receptor 4-dependent mechanism activated by previous infection that induces Neu3 neuraminidase among colonic epithelial cells concurrent with increased Cathepsin-G protease secretion by Paneth cells. These anatomically separated host responses merge with the desialylation of nascent colonic Mucin-2 by Neu3 rendering the mucosal barrier susceptible to increased proteolytic breakdown by Cathepsin-G. Depletion of Cathepsin-G or Neu3 function using pharmacological inhibitors or genetic-null alleles protected against Mucin-2 proteolysis and barrier erosion and reduced the frequency and severity of colitis, revealing approaches to preserve and potentially restore the mucosal barrier.

Increased expression of sodium-glucose cotransporter 2 and O-GlcNAcylation in hepatocytes drives non-alcoholic steatohepatitis.

AIMS: Steatosis reducing effects of sodium-glucose cotransporter 2 (SGLT2) inhibitors in non-alcoholic steatohepatitis (NASH) has been consistently reported in humans, but their mechanism remains uncertain. In this study, we examined the expression of SGLT2 in human livers and investigated the crosstalk between SGLT2 inhibition and hepatic glucose uptake, intracellular O-GlcNAcylation, and autophagic regulation in NASH. MATERIALS AND METHODS: Human liver samples obtained from subjects with/without NASH were analyzed. For in vitro studies, human normal hepatocytes and hepatoma cells were treated with SGLT2 inhibitor under high-glucose and high-lipid conditions. NASH in vivo was induced by a high-fat, -fructose, and -cholesterol Amylin liver NASH (AMLN) diet for 10 weeks followed by an additional 10 weeks with/without SGLT2 inhibitor (empagliflozin 10 mg/kg/day). RESULTS: Liver samples from subjects with NASH were associated with increased SGLT2 and O-GlcNAcylation expression compared with controls. Under NASH condition (in vitro condition with high glucose and lipid), intracellular O-GlcNAcylation and inflammatory markers were increased in hepatocytes and SGLT2 expression was upregulated; SGLT2 inhibitor treatment blocked these changes by directly reducing hepatocellular glucose uptake. In addition, decreased intracellular O-GlcNAcylation by SGLT2 inhibitor promoted autophagic flux through AMPK-TFEB activation. In the AMLN diet-induced NASH mice model, SGLT2 inhibitor alleviated lipid accumulation, inflammation, and fibrosis through autophagy activation related to decreased SGLT2 expression and O-GlcNAcylation in the liver. CONCLUSIONS: This study firstly demonstrates increased SGLT2 expression in NASH and secondly reveals the novel effect of SGLT2 inhibition on NASH through autophagy activation mediated by inhibition of hepatocellular glucose uptake and consequently decreased intracellular O-GlcNAcylation.

O-GlcNAcylation of RIPK1 rescues red blood cells from necroptosis.

Necroptosis is a type of cell death with excessive inflammation and organ damage in various human diseases. Although abnormal necroptosis is common in patients with neurodegenerative, cardiovascular, and infectious diseases, the mechanisms by which O-GlcNAcylation contributes to the regulation of necroptotic cell death are poorly understood. In this study, we reveal that O-GlcNAcylation of RIPK1 (receptor-interacting protein kinase1) was decreased in erythrocytes of the mouse injected with lipopolysaccharide, resulting in the acceleration of erythrocyte necroptosis through increased formation of RIPK1-RIPK3 complex. Mechanistically, we discovered that O-GlcNAcylation of RIPK1 at serine 331 in human (corresponding to serine 332 in mouse) inhibits phosphorylation of RIPK1 at serine 166, which is necessary for the necroptotic activity of RIPK1 and suppresses the formation of the RIPK1-RIPK3 complex in Ripk1 -/- MEFs. Thus, our study demonstrates that RIPK1 O-GlcNAcylation serves as a checkpoint to suppress necroptotic signaling in erythrocytes.

Poly-ADP ribosylation of p21 by tankyrases promotes p21 degradation and regulates cell cycle progression.

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.

Mast4 determines the cell fate of MSCs for bone and cartilage development.

Mesenchymal stromal cells (MSCs) differentiation into different lineages is precisely controlled by signaling pathways. Given that protein kinases play a crucial role in signal transduction, here we show that Microtubule Associated Serine/Threonine Kinase Family Member 4 (Mast4) serves as an important mediator of TGF-ß and Wnt signal transduction in regulating chondro-osteogenic differentiation of MSCs. Suppression of Mast4 by TGF-ß1 led to increased Sox9 stability by blocking Mast4-induced Sox9 serine 494 phosphorylation and subsequent proteasomal degradation, ultimately enhancing chondrogenesis of MSCs. On the other hand, Mast4 protein, which stability was enhanced by Wnt-mediated inhibition of GSK-3ß and subsequent Smurf1 recruitment, promoted ß-catenin nuclear localization and Runx2 activity, increasing osteogenesis of MSCs. Consistently, Mast4-/- mice demonstrated excessive cartilage synthesis, while exhibiting osteoporotic phenotype. Interestingly, Mast4 depletion in MSCs facilitated cartilage formation and regeneration in vivo. Altogether, our findings uncover essential roles of Mast4 in determining the fate of MSC development into cartilage or bone.

Nephrin expression in human epidermal keratinocytes and its implication in poor wound closure.

Nephrin is a type-1 transmembrane protein and a component of the slit diaphragm renal-filtration barrier. It has several functions in actin remodeling and cell-cell adhesion. Nephrin is principally located in the kidney glomerulus, but several studies have reported that nephrin is found in the pancreas, brain, and placenta. However, nephrin expression and its role in human skin have not yet been reported. First, using single-cell RNA sequencing, immunohistochemistry, and immuno-electron microscopy, nephrin expression was confirmed in human-skin epidermal keratinocytes. Nephrin expression colocalized with the expression of zonula occludens-1 in keratinocytes and was closely related to keratinocyte cell density, proliferation, and migration. High glucose treatment decreased nephrin expression and compromised keratinocyte cell migration without yes-associated protein nuclear entry. This reduced cell migration under high glucose conditions was improved in nephrin-overexpressing keratinocytes. Nephrin was highly expressed on the margins of re-epithelized epidermis based on in vivo mice and ex vivo human skin wound models. The results demonstrate that nephrin is expressed in human-skin keratinocytes and functions in cell adhesion, proliferation, and migration. In conclusion, this study suggests that nephrin may have a variety of physiological roles in human skin.

Association of Metabolomic Change and Treatment Response in Patients with Non-Alcoholic Fatty Liver Disease.

Non-alcoholic fatty liver disease (NAFLD) is the major cause of chronic liver disease, yet cost-effective and non-invasive diagnostic tools to monitor the severity of the disease are lacking. We aimed to investigate the metabolomic changes in NAFLD associated with therapeutic responses. It was conducted in 63 patients with NAFLD who received either ezetimibe plus rosuvastatin or rosuvastatin monotherapy. The treatment response was determined by MRI performed at baseline and week 24. The metabolites were measured at baseline and week 12. In the combination group, a relative decrease in xanthine was associated with a good response to liver fat decrease, while a relative increase in choline was associated with a good response to liver stiffness. In the monotherapy group, the relative decreases in triglyceride (TG) 20:5_36:2, TG 18:1_38:6, acetylcarnitine (C2), fatty acid (FA) 18:2, FA 18:1, and docosahexaenoic acid were associated with a decrease in liver fat, while hexosylceramide (d18:2/16:0) and hippuric acid were associated with a decrease in liver stiffness. Models using the metabolite changes showed an AUC of >0.75 in receiver operating curve analysis for predicting an improvement in liver fat and stiffness. This approach revealed the physiological impact of drugs, suggesting the mechanism underlying the development of this disease.

O-GlcNAcylation: An Emerging Protein Modification Regulating the Hippo Pathway.

The balance between cellular proliferation and apoptosis and the regulation of cell differentiation must be established to maintain tissue homeostasis. These cellular responses involve the kinase cascade-mediated Hippo pathway as a crucial regulator. Hence, Hippo pathway dysregulation is implicated in diverse diseases, including cancer. O-GlcNAcylation is a non-canonical glycosylation that affects multiple signaling pathways through its interplay with phosphorylation in the nucleus and cytoplasm. An abnormal increase in the O-GlcNAcylation levels in various cancer cells is a potent factor in Hippo pathway dysregulation. Intriguingly, Hippo pathway dysregulation also disrupts O-GlcNAc homeostasis, leading to a persistent elevation of O-GlcNAcylation levels, which is potentially pathogenic in several diseases. Therefore, O-GlcNAcylation is gaining attention as a protein modification that regulates the Hippo pathway. This review presents a framework on how O-GlcNAcylation regulates the Hippo pathway and forms a self-perpetuating cycle with it. The pathological significance of this self-perpetuating cycle and clinical strategies for targeting O-GlcNAcylation that causes Hippo pathway dysregulation are also discussed.

Short Term Isocaloric Ketogenic Diet Modulates NLRP3 Inflammasome Via B-hydroxybutyrate and Fibroblast Growth Factor 21.

A ketogenic diet (KD) is known to have beneficial health effects. Various types of KD interventions have been applied to manage metabolic syndrome based on modification of diet parameters such as duration of intervention, macronutrient components, and total calories. Nevertheless, the beneficial health impact of isocaloric KD is largely unknown, especially in healthy subjects. The present study investigated the acute effects of a 3-day isocaloric KD. In this non-randomized intervention study, we recruited 15 healthy volunteers aged 24-38 years (7 men and 8 women) and placed them on an isocaloric KD restricting intake of carbohydrates but not energy (75% fat, 20% protein, 5% carbohydrate) for 3 days. Biochemical profiles and laboratory measurements were performed. Peripheral blood monocular cells were cultured, and measured cell stimulated cytokines. After short-term isocaloric KD, subjects lost body weight and serum free fatty acid levels were increased. These results accompanied elevated serum ß-hydroxybutyrate (BHB) concentration and fibroblast growth factor 21 (FGF21) levels and improved insulin sensitivity. Regarding the direct effect of BHB on inflammasome activation, interleukin-1ß (IL-1ß) and tumor necrosis factor-α secretion in response to adenosine triphosphate or palmitate stimulation in human macrophages decreased significantly after isocaloric KD. In ex-vivo experiments with macrophages, both FGF21 and BHB further reduced IL-1ß secretion compared to either BHB or FGF21 alone. The inhibitory effect of FGF21 on IL-1ß secretion was blunted with bafilomycin treatment, which blocked autophagy flux. In conclusion, isocaloric KD for 3 days is a promising approach to improve metabolic and inflammatory status. Clinical Trial Registration: clinicaltrials.gov (NCT02964572).

A GLP-1/GLP-2 receptor dual agonist to treat NASH: Targeting the gut-liver axis and microbiome.

BACKGROUND AND AIMS: Currently there is no Food and Drug Administration-approved drug to treat NAFLD and NASH, the rates of which are increasing worldwide. Although NAFLD/NASH are highly complex and heterogeneous conditions, most pharmacotherapy pipelines focus on a single mechanistic target. Considering the importance of the gut-liver axis in their pathogenesis, we investigated the therapeutic effect of a long-acting dual agonist of glucagon-like peptide (GLP)-1 and GLP-2 receptors in mice with NAFLD/NASH. APPROACH AND RESULTS: C57BL/6J mice were fed a choline-deficient high-fat diet/high fructose and sucrose solution. After 16 weeks, mice were randomly allocated to receive vehicle, GLP1-Fc, GLP2-Fc, or GLP1/2-Fc fusion (GLP1/2-Fc) subcutaneously every 2 days for 4 weeks. Body weight was monitored, insulin/glucose tolerance tests were performed, feces were collected, and microbiome profiles were analyzed. Immobilized cell systems were used to evaluate direct peptide effect. Immunohistochemistry, quantitative PCR, immunoblot analysis, tunnel assay, and biochemical assays were performed to assess drug effects on inflammation, hepatic fibrosis, cell death, and intestinal structures. The mice had well-developed NASH phenotypes. GLP1/2-Fc reduced body weight, glucose levels, hepatic triglyceride levels, and cellular apoptosis. It improved liver fibrosis, insulin sensitivity, and intestinal tight junctions, and increased microvillus height, crypt depth, and goblet cells of intestine compared with a vehicle group. Similar effects of GLP1/2-Fc were found in in vitro cell systems. GLP1/2-Fc also changed microbiome profiles. We applied fecal microbiota transplantation (FMT) gain further insight into the mechanism of GLP1/2-Fc-mediated protection. We confirmed that FMT exerted an additive effect on GLP1-Fc group, including the body weight change, liver weight, hepatic fat accumulation, inflammation, and hepatic fibrosis. CONCLUSIONS: A long-acting dual agonist of GLP-1 and GLP-2 receptors is a promising therapeutic strategy to treat NAFLD/NASH.

O-GlcNAcylated p53 in the liver modulates hepatic glucose production.

p53 regulates several signaling pathways to maintain the metabolic homeostasis of cells and modulates the cellular response to stress. Deficiency or excess of nutrients causes cellular metabolic stress, and we hypothesized that p53 could be linked to glucose maintenance. We show here that upon starvation hepatic p53 is stabilized by O-GlcNAcylation and plays an essential role in the physiological regulation of glucose homeostasis. More specifically, p53 binds to PCK1 promoter and regulates its transcriptional activation, thereby controlling hepatic glucose production. Mice lacking p53 in the liver show a reduced gluconeogenic response during calorie restriction. Glucagon, adrenaline and glucocorticoids augment protein levels of p53, and administration of these hormones to p53 deficient human hepatocytes and to liver-specific p53 deficient mice fails to increase glucose levels. Moreover, insulin decreases p53 levels, and over-expression of p53 impairs insulin sensitivity. Finally, protein levels of p53, as well as genes responsible of O-GlcNAcylation are elevated in the liver of type 2 diabetic patients and positively correlate with glucose and HOMA-IR. Overall these results indicate that the O-GlcNAcylation of p53 plays an unsuspected key role regulating in vivo glucose homeostasis.

Neu3 neuraminidase induction triggers intestinal inflammation and colitis in a model of recurrent human food-poisoning.

Intestinal inflammation is the underlying basis of colitis and the inflammatory bowel diseases. These syndromes originate from genetic and environmental factors that remain to be fully identified. Infections are possible disease triggers, including recurrent human food-poisoning by the common foodborne pathogen Salmonella enterica Typhimurium (ST), which in laboratory mice causes progressive intestinal inflammation leading to an enduring colitis. In this colitis model, disease onset has been linked to Toll-like receptor-4-dependent induction of intestinal neuraminidase activity, leading to the desialylation, reduced half-life, and acquired deficiency of anti-inflammatory intestinal alkaline phosphatase (IAP). Neuraminidase (Neu) inhibition protected against disease onset; however, the source and identity of the Neu enzyme(s) responsible remained unknown. Herein, we report that the mammalian Neu3 neuraminidase is responsible for intestinal IAP desialylation and deficiency. Absence of Neu3 thereby prevented the accumulation of lipopolysaccharide-phosphate and inflammatory cytokine expression in providing protection against the development of severe colitis.

O-GlcNAc stabilizes SMAD4 by inhibiting GSK-3ß-mediated proteasomal degradation.

O-linked ß-N-acetylglucosamine (O-GlcNAc) is a post-translational modification which occurs on the hydroxyl group of serine or threonine residues of nucleocytoplasmic proteins. It has been reported that the presence of this single sugar motif regulates various biological events by altering the fate of target proteins, such as their function, localization, and degradation. This study identified SMAD4 as a novel O-GlcNAc-modified protein. SMAD4 is a component of the SMAD transcriptional complex, a major regulator of the signaling pathway for the transforming growth factor-ß (TGF-ß). TGF-ß is a powerful promoter of cancer EMT and metastasis. This study showed that the amount of SMAD4 proteins changes according to cellular O-GlcNAc levels in human lung cancer cells. This observation was made based on the prolonged half-life of SMAD4 proteins. The mechanism behind this interaction was that O-GlcNAc impeded interactions between SMAD4 and GSK-3ß which promote proteasomal degradation of SMAD4. In addition, O-GlcNAc modification on SMAD4 Thr63 was responsible for stabilization. As a result, defects in O-GlcNAcylation on SMAD4 Thr63 attenuated the reporter activity of luciferase, the TGF-ß-responsive SMAD binding element (SBE). This study's findings imply that cellular O-GlcNAc may regulate the TGF-ß/SMAD signaling pathway by stabilizing SMAD4.

Cancer cell death using metabolic glycan labelling techniques.

Herein we describe a method for inducing cancer cell death, which relies on the use of a H2O2-responsive glycan metabolic precursor in conjunction with antibody-dependent cellular cytotoxicity (ADCC) or photodynamic therapy (PDT).

Mutual regulation between OGT and XIAP to control colon cancer cell growth and invasion.

O-GlcNAc transferase (OGT) is an enzyme that catalyzes the O-GlcNAc modification of nucleocytoplasmic proteins and is highly expressed in many types of cancer. However, the mechanism regulating its expression in cancer cells is not well understood. This study shows that OGT is a substrate of the E3 ubiquitin ligase X-linked inhibitor of apoptosis (XIAP) which plays an important role in cancer pathogenesis. Although LSD2 histone demethylase has already been reported as an E3 ubiquitin ligase in lung cancer cells, we identified XIAP as the main E3 ubiquitin ligase in colon cancer cells. Interestingly, OGT catalyzes the O-GlcNAc modification of XIAP at serine 406 and this modification is required for the E3 ubiquitin ligase activity of XIAP toward specifically OGT. Moreover, O-GlcNAcylation of XIAP suppresses colon cancer cell growth and invasion by promoting the proteasomal degradation of OGT. Therefore, our findings regarding the reciprocal regulation of OGT and XIAP provide a novel molecular mechanism for controlling cancer growth and invasion regulated by OGT and O-GlcNAc modification.

LDL receptor-related protein LRP6 senses nutrient levels and regulates Hippo signaling.

Controlled cell growth and proliferation are essential for tissue homeostasis and development. Wnt and Hippo signaling are well known as positive and negative regulators of cell proliferation, respectively. The regulation of Hippo signaling by the Wnt pathway has been shown, but how and which components of Wnt signaling are involved in the activation of Hippo signaling during nutrient starvation are unknown. Here, we report that a reduction in the level of low-density lipoprotein receptor-related protein 6 (LRP6) during nutrient starvation induces phosphorylation and cytoplasmic localization of YAP, inhibiting YAP-dependent transcription. Phosphorylation of YAP via loss of LRP6 is mediated by large tumor suppressor kinases 1/2 (LATS1/2) and Merlin. We found that O-GlcNAcylation of LRP6 was reduced, and the overall amount of LRP6 was decreased via endocytosis-mediated lysosomal degradation during nutrient starvation. Merlin binds to LRP6; when LRP6 is less O-GlcNAcylated, Merlin dissociates from it and becomes capable of interacting with LATS1 to induce phosphorylation of YAP. Our data suggest that LRP6 has unexpected roles as a nutrient sensor and Hippo signaling regulator.

O-GlcNAcylation of Mef2c regulates myoblast differentiation.

Unlike other types of glycosylation, O-GlcNAcylation is a single glycosylation which occurs exclusively in the nucleus and cytosol. O-GlcNAcylation underlie metabolic diseases, including diabetes and obesity. Furthermore, O-GlcNAcylation affects different oncogenic processes such as osteoblast differentiation, adipogenesis and hematopoiesis. Emerging evidence suggests that skeletal muscle differentiation is also regulated by O-GlcNAcylation, but the detailed molecular mechanism has not been fully elucidated. In this study, we showed that hyper-O-GlcNAcylation reduced the expression of myogenin, a transcription factor critical for terminal muscle development, in C2C12 myoblasts differentiation by O-GlcNAcylation on Thr9 of myocyte-specific enhancer factor 2c. Furthermore, we showed that O-GlcNAcylation on Mef2c inhibited its DNA binding affinity to myogenin promoter. Taken together, we demonstrated that hyper-O-GlcNAcylation attenuates skeletal muscle differentiation by increased O-GlcNAcylation on Mef2c, which downregulates its DNA binding affinity.