Polypeptide N-acetylgalactosaminyltransferase Research Articles

Indoxyl sulfate induces left ventricular hypertrophy via the AhR-FGF23-FGFR4 signaling pathway.

Patients with chronic kidney disease (CKD) have a high risk of left ventricular hypertrophy (LVH). Fibroblast growth factor 23 (FGF23) and indoxyl sulfate (IS) are associated with LVH in patients with CKD, but the interactions between these molecules remain unknown. We investigated whether IS contributes to LVH associated with FGF23 in cultured cardiomyocytes and CKD mice. In cultured rat cardiac myoblast H9c2 cells incubated with IS, mRNA levels of the LVH markers atrial natriuretic factor, brain natriuretic peptide, and β-myosin heavy chain were significantly upregulated. Levels of mRNA of the polypeptide N-acetylgalactosaminyltransferase 3 (GALNT3), which regulates FGF23 O-glycosylation, and FGF23 were also upregulated in H9c2 cells. Intact FGF23 protein expression and fibroblast growth factor receptor 4 (FGFR4) phosphorylation were increased in cell lysates by IS administration. In C57BL/6J mice with heminephrectomy, IS promoted LVH, whereas the inhibition of FGFR4 significantly reduced heart weight and left ventricular wall thickness in IS-treated groups. While there was no significant difference in serum FGF23 concentrations, cardiac FGF23 protein expression was markedly increased in IS-injected mice. GALNT3, hypoxia-inducible factor 1 alpha, and FGF23 protein expression was induced in H9c2 cells by IS treatment and suppressed by the inhibition of Aryl hydrocarbon receptor which is the receptor for IS. This study suggests that IS increases FGF23 protein expression via an increase in GALNT3 and hypoxia-inducible factor 1 alpha expression, and activates FGF23-FGFR4 signaling in cardiomyocytes, leading to LVH.

Polypeptide N-Acetylgalactosaminyltransferase 14 (GALNT14) as a Chemosensitivity-Related Biomarker for Osteosarcoma.

Osteosarcoma is the most common primary bone tumor. Polypeptide N-acetylgalactosaminyltransferase 14 (GALNT14), a member of the N-acetylgalactosaminyltransferase family, has been considered to be associated with various cancers. However, its role in osteosarcoma remains unknown. Here, we aimed to explore the expression and potential mechanism of GALNT14 in osteosarcoma through bioinformatics analysis and in vitro experiments. We investigated GALNT14 expression in osteosarcoma using GEO, the TIMER database, and clinical samples. Protein-protein interaction (PPI) network analysis on GALNT14 was performed by STRING. TARGET was used to identify differentially expressed genes (DEGs) between high and low GALNT14 expression. The correlation between GALNT14 and cuproptosis-related genes in osteosarcoma was analyzed by R language. The prognostic significance of GALNT14 was examined by Kaplan-Meier survival analysis. Additionally, we inhibited GALNT14 function in an osteosarcoma cell line by transfecting siRNA and subsequently explored the effect on drug sensitivity by CCK-8, clonogenic assay, and flow cytometry. GALNT14 was significantly elevated in osteosarcoma tissue, osteosarcoma cell lines, and metastatic osteosarcoma. PPI analysis revealed that GALNT14 was associated with MUC7, MUC13, MUC5AC, C1GALT1, MUC15, MUC16, MUC1, MUC4, MUC21, and MUC17. In the high GALNT14 expression group, we discovered 81 upregulated DEGs and 73 downregulated DEGs. Functional enrichment analysis of DEGs showed significant enrichment in the Wnt, TGF-β, Hippo, PI3K signaling pathways and cell adhesion molecules. Expression of cuproptosis-related genes was closely related in osteosarcoma, and GALNT14 expression was significantly positively correlated with FDX1, a key regulator of cuproptosis. Kaplan-Meier survival showed that GALNT14 was linked to poor overall survival and disease-free survival in osteosarcoma. In vitro experiments suggested that GALNT14 was associated with chemotherapy resistance in osteosarcoma. We identified a GALNT family gene, GALNT14, that was highly expressed in osteosarcoma. This gene was closely associated with metastasis, progression, cuproptosis-related genes, and chemosensitivity of osteosarcoma, and showed correlation with poor overall survival and disease-free survival in osteosarcoma.

GALNT3 protects against phosphate-induced calcification in vascular smooth muscle cells by enhancing active FGF23 and inhibiting the wnt/β-catenin signaling pathway

Vascular calcification (VC) acts as a notable risk factor in the cardiovascular system. Disorder of phosphorus (Pi) metabolism promotes VC. Recent findings show that polypeptide N-acetylgalactosaminyltransferase 3(GALNT3) is Pi responsive and with potent effects on Pi homeostasis. However, whether GALNT3 is involved in high Pi-induced VC remains unclear. The present study investigated the potential role of GALNT3 as a novel regulator of VC. In vitro, human aortic smooth muscle cells (HASMCs) calcification was induced by inorganic Pi, while in vivo, C57BL/6 J mice were used to determine the effects of GALNT3 on Vitamin D3-induced medial arterial calcification. Alizarin red staining, Von Kossa staining, calcium and alkaline phosphatase (ALP) activity were performed to test VC. We showed that expression of GALNT3 was increased in the calcified HASMCs and aortas of the calcified mice.In vitro, overexpression of GALNT3 increased the levels of active full-length FGF23, accompanied by suppression of the osteoblast-related factors (Runx2 and BMP2), and further inhibited the formation of calcified nodules. Moreover, the protein levels of Wnt3a and active β-catenin were determined and it was found that GALNT3 significantly inhibited their expression. LiCl, a Wnt/β-catenin signaling activator, was observed to reverse the protective effect of GALNT3 overexpression. The opposite results were observed in the GALNT3 knockdown cells. In vivo, overexpression of GALNT3 by adeno-associated virus decreased the serum Pi and slowed the formation of aortic calcification in the calcified mice. In conclusion, our results indicate that GALNT3 counteracts high Pi-induced osteoblastic differentiation of VSMCs and protects against the initiation and progression of VC by inhibiting the Wnt/β-catenin signaling pathway.

MiR-30c facilitates natural killer cell cytotoxicity to lung cancer through targeting GALNT7.

MicroRNAs (miRNAs) have been reported to play important roles in regulating natural killer (NK) cell cytotoxicity to cancer cells. This study aimed to investigate the effects and potential mechanism of miR-30c in regulating NK cell cytotoxicity to lung cancer cells. Primary NK cells were derived from the peripheral blood of lung cancer and normal participants. Exosomes were isolated and validated via transmission electron microscopy and nanoparticle tracking analysis. The levels of miR-30c, polypeptide N-acetylgalactosaminyltransferase 7 (GALNT7) and proteins in PI3K/AKT pathway were determined using quantitative real-time polymerase chain reaction or western blot. Tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ) levels and the cytotoxicity of effector NK cells to target lung cancer cells were measured via enzyme linked immunosorbent assay, cell apoptosis or xenograft experiments. The relationship between miR-30c and GALNT7 was analyzed by luciferase activity, RNA pull-down and RNA immunoprecipitation assays. And a xenograft mice model was established to verify the effect of miR-30c in regulating NK cell cytotoxicity to lung cancer cells in vivo. NK cell-derived exosomes carrying miR-30c, and miR-30c level was significantly downregulated in primary NK cells of lung cancer patients. MiR-30c overexpression promoted TNF-α and IFN-γ secretion and enhanced the cytotoxicity of interleukin 2 (IL-2)-treated NK cells to lung cancer cells, while knockdown of miR-30c played an opposite effect in regulating the cytotoxicity of NK cells to lung cancer cells. GALNT7 was a target of miR-30c and was negatively regulated by miR-30c. Besides, miR-30c targeted GALNT7 to exert its function in regulating NK cell cytotoxicity. Furthermore, GALNT7 prompted the activation of PI3K/AKT pathway in NK cells. Additionally, miR-30c overexpression enhanced NK cell cytotoxicity to lung cancer cells and inhibited tumor growth in vivo. miR-30c enhanced NK cell cytotoxicity to lung cancer cells via decreasing GALNT7 and inactivating the PI3K/AKT pathway, suggesting that regulating miR-30c expression maybe a promising approach for enhancing NK cell-based antitumor therapies.

Association of rs9679162 Genetic Polymorphism and Aberrant Expression of Polypeptide N-Acetylgalactosaminyltransferase 14 (GALNT14) in Head and Neck Cancer.

Simple SummaryNeoadjuvant chemotherapy was performed before surgery. Because the tumor itself and the surrounding vascular bed were not damaged, the chemotherapy we performed could have good drug delivery. After the operation, the volume of the tumor can be reduced to facilitate surgery or radiotherapy. However, neoadjuvant chemotherapy also delays the patient’s time to receive main therapy. The physician must make sure that it has a good response and does not allow disease progression in the patient during neoadjuvant chemotherapy. Therefore, predicting the treatment response of neoadjuvant chemotherapy can shorten the treatment time, reduce the harm of chemotherapy side effects, and avoid the occurrence of drug resistance. The results of this study showed that GALNT14-rs9679162 and mRNA expression were associated with post-treatment survival in head and neck cancer. It can be used as an indicator to predict the treatment response of neoadjuvant chemotherapy.The polypeptide N-Acetylgalactosaminyltransferase 14 (GALNT14) rs9679162 and mRNA expression were associated with treatment outcome in various cancers. However, the relation of GALNT14 and head and neck cancer were nuclear. A total of 199 patients with head and neck squamous cell carcinoma (HNSCC) were collected in this study, including oral SCC (OSCC), oropharyngeal SCC (OPSCC), laryngeal SCC (LSCC), and others. The DNA and RNA of cancer tissues were extracted using the TRI Reagent method. The rs9679162 was analyzed using polymerase chain reaction (PCR) and sequencing methods in 199 DNA specimens, and the mRNA expression was analyzed using quantitative reverse transcription PCR (RT-qPCR) methods in 68 paired RNA specimens of non-cancerous matched tissues (NCMT) and tumor tissues. The results showed that the genotype of TT, TG, and GG appeared at 30%, 44%, and 26%, respectively. Non-TT genotype or G alleotype were associated with alcohol, betel nut, and cigarette using among patients with OSCC, and it also affected the treatment and survival of patients with OSCC and LSCC. High GALNT14 mRNA expression levels increased lymphatic metastasis of patients with HNSCC, and treatment and survival in patients with OPSCC. Overall, the GALNT14-rs9679162 genotype and mRNA expression level can be used as indicators of HNSCC treatment prognosis.

A novel role for GALNAC-T2 dependent glycosylation in energy homeostasis

Background and Aims : GALNT2, encoding polypeptide N-acetylgalactosaminyltransferase 2 (GalNAc-T2), was initially discovered as a regulator of high-density lipoprotein metabolism. GalNAc-T2 is known to exert these effects through post-translational modification, i.e., O-linked glycosylation of secreted proteins including apolipoprotein C-III, angiopoietin-like 3 and phospholipid transfer protein with established roles in plasma lipid metabolism. More recently it has become clear that loss of GALNT2 in rodents, cattle, nonhuman primates and humans should be regarded as a novel congenital disorder of glycosylation which affects energy metabolism, a topic that has not yet been specifically addressed.

FRI546 - Polypeptide N-acetylgalactosaminyltransferase 14 mediated O-glycosylation on prohibitin-2 serine 161 modulates cell growth, migration and drug susceptibility in hepatocellular carcinoma

FRI546 - Polypeptide N-acetylgalactosaminyltransferase 14 mediated O-glycosylation on prohibitin-2 serine 161 modulates cell growth, migration and drug susceptibility in hepatocellular carcinoma

478-P: GALNT2 Variant Associates with Increased ANGPTL3 Circulation Level and Obesity

Background: Polypeptide N-Acetylgalactosaminyltransferase 2 (GALNT2) gene has been associated with serum lipid levels, insulin resistance, and adipogenesis. One of its variants, rs4846914 has been shown to associate with Triglycerides and HDL levels in global genome-wide association studies. ANGPTL proteins have emerged as important regulators of lipid metabolism through its regulation of lipoprotein lipase activity. In this study we aimed to evaluate the association between GALNT2 variant with plasma level of ANGPTL3, 8 and apolipoproteins in individuals of Arab ethnicity. Methods: GALNT2 rs4846914 variant was genotyped in a cohort of 278 Arab individuals from Kuwait. Plasma levels of ANGPTL3 and 8 were measured by ELISA. Apolipoproteins were measured by Luminex multiplexing assay. Significance of association signals was assessed using Bonferroni-corrected P-values. Results: The GALNT2 rs4846914_G allele was associated with increased ANGPTL3 but not ANGPTL8 plasma levels. It was associated with obesity status with an odds ratio of 1.45 (CI:1.024-2.055) (P-value =0.036) . It also associated with increased levels of ApoC1 (P-values ≤0.006) and lower levels of HDL (P-values ≤0.05) . Levels of HDL were found to be mediated by interactions between genotypes (AG+GG) at rs4846914 and measures of percentage body fat (PBF) , apolipoproteins A1a, C1 and B48. Conclusions: Association between GALNT2 variant and lipid metabolism could be mediated through the increased ANGPTL3 protein level. Disclosure A.T.Thangavel: None. M.Abu-farha: None. P.Hebbar: None. M.G.Qaddoumi: None. F.Almulla: None. J.Abubaker: None.

Long Non-Coding RNA AL139385.1 as a Novel Prognostic Biomarker in Lung Adenocarcinoma.

Lung adenocarcinoma (LUAD) is the most common histological lung cancer, and it is the leading cause of cancer-related deaths worldwide. LncRNA-AL139385.1 (ENSG00000275880) is a novel lncRNA that is abnormally expressed in various cancer types including LUAD. However, the underlying biological function and potential mechanisms of AL139385.1 driving the progression of LUAD remain unclear. In this study, we investigated the role of AL139385.1 in LUAD and found that DNA hypomethylation was positively correlated with AL139385.1 expression in LUAD. Moreover, we uncover that the expression of AL139385.1 in LUAD tissues was significantly higher than that of AL139385.1 expression in adjacent normal tissues. Kaplan–Meier survival analysis showed that patients with higher AL139385.1 expression correlated with adverse overall survival and progression-free survival. Receiver operating characteristic (ROC) curve analysis showed that the area under the curve (AUC) value of AL139385.1 was 0.808. Correlation analysis showed that AL139385.1 expression was associated with immune infiltration in LUAD. We also found that AL139385.1 was upregulated in LUAD cancer tissues and cell lines. Knockdown of AL139385.1 significantly inhibited cell proliferation and migration abilities of LUAD. Finally, we constructed a ceRNA network that includes hsa-miR-532-5p and four mRNAs (GALNT3, CYCS, EIF5A, and ITGB4) specific to AL139385.1 in LUAD. Subsequent Kaplan–Meier survival analysis suggested that polypeptide N-acetylgalactosaminyltransferase 3 (GALNT3), cytochrome c, somatic (CYCS), eukaryotic translation initiation factor 5A (EIF5A), and integrin subunit beta 4 (ITGB4), were potential prognostic biomarkers for patients with LUAD. In conclusion, this finding provides possible mechanisms underlying the abnormal upregulation of AL139385.1 as well as a comprehensive view of the AL139385.1-mediated competing endogenous RNAs (ceRNA) network in LUAD, thereby highlighting its potential role in diagnosis and therapy.

GALNTs: master regulators of metastasis-associated epithelial-mesenchymal transition (EMT)?

In humans, the UDP-N-α-D galactosamine:polypeptide N-acetylgalactosaminyltransferases family (ppGalNAc-Ts, GalNAc-Ts or GALNTs) comprises 20 isoenzymes. They are responsible for the initial synthesis of α-GalNAc1,3-O-Ser/Thr, or Tn antigen, at initiation of mucin type O-linked glycosylation. This structure is normally extended by the further sequential action of glycosytransferases to build more complex linear or branched O-linked structures, but in cancers it is frequently left unelaborated, and its presence is often associated with poor patient prognosis. Altered levels of GALNT expression or distribution have also been extensively reported in a wide range of cancers. These changes would be predicted to result in marked alterations in GalNAc O-linked glycosylation, including altered levels of site specific O-linked glycosylation and changes in the glycan structures formed, including, potentially, exposure of truncated O-glycans such as Tn antigen. Many reports have demonstrated that altered levels of specific GALNTs have prognostic significance in cancers, or shown that they are associated with changes in cell behaviour, including proliferation, migration, invasion or growth and metastasis in animal models. We have previously reviewed how deregulation of GALNTs in several epithelial cancers is a feature of different stages metastasis. Here we consider evidence that changes in GALNT expression, and therefore consequent alterations in GalNAc O-linked glycosylation, may directly influence molecules implicated in aspects of epithelial-mesenchymal transition (EMT), a fundamental aspect of cancer metastasis, during which epithelial cancer cells lose their cell-cell junctions, apical-basal polarity and adhesive interactions with basement membrane and become mesenchymal, with a spindle-shaped morphology and increased migratory capacity.

A novel role for GalNAc-T2 dependent glycosylation in energy homeostasis

ObjectiveGALNT2, encoding polypeptide N-acetylgalactosaminyltransferase 2 (GalNAc-T2), was initially discovered as a regulator of high-density lipoprotein metabolism. GalNAc-T2 is known to exert these effects through post-translational modification, i.e., O-linked glycosylation of secreted proteins with established roles in plasma lipid metabolism. It has recently become clear that loss of GALNT2 in rodents, cattle, nonhuman primates, and humans should be regarded as a novel congenital disorder of glycosylation that affects development and body weight. The role of GALNT2 in metabolic abnormalities other than plasma lipids, including insulin sensitivity and energy homeostasis, is poorly understood. MethodsGWAS data from the UK Biobank was used to study variation in the GALNT2 locus beyond changes in high-density lipoprotein metabolism. Experimental data were obtained through studies in Galnt2−/− mice and wild-type littermates on both control and high-fat diet. ResultsFirst, we uncovered associations between GALNT2 gene variation, adiposity, and body mass index in humans. In mice, we identify the insulin receptor as a novel substrate of GalNAc-T2 and demonstrate that Galnt2−/− mice exhibit decreased adiposity, alterations in insulin signaling and a shift in energy substrate utilization in the inactive phase. ConclusionsThis study identifies a novel role for GALNT2 in energy homeostasis, and our findings suggest that the local effects of GalNAc-T2 are mediated through posttranslational modification of the insulin receptor.

Abstract 149: Hyperphosphatemic familial tumoral calcinosis due to homozygous mutation at Leu366Arg: First case report

Background: Hyperphosphatemic familial tumoral calcinosis (HFTC) is a rare disabling autosomal recessive metabolic disorder characterized by hyperphosphatemia and manifestations ranging from progressive deposition of calcium phosphate crystals in the skin, soft tissues and bone, and hyperostosis. The most common cause of HFTC is related to loss-of-function mutations in GALNT3 (Polypeptide N-Acetylgalactosaminyl transferase 3) gene causing defective glycosylation of FGF23.Material and Methods: A 16 year boy presented with lump in the left gluteal area and hyperphosphatemia. Serum ionized calcium, alkaline phosphate, intact PTH, 25-OH vitamin D, serum creatinine and C terminal FGF 23, magnetic resonance imaging of hip, dental CBCT , bone mineral density at lumbar spine and hip by dual x-ray absorptiometry (DEXA) BMD, and 99Tc MDP bone scan, followed by next generation sequencing (NGS) and Sanger's sequencing.Results: A 16 year otherwise healthy boy presented with hard painless lump in the left gluteal area and hyperphosphatemia for last one year. His parents had third degree consanguinity with ancestral roots in Middle East. Father has small calcinosis in the right hand and mother was asymptomatic. Child had no associated bone pain, proximal muscle weakness, fracture or constitutional symptoms. S. phosphorus was 6.98mg/dl (normal: 3.5–5.5) with inappropriately increased TRP (95%) and TMP-GFR (7.2). Serum ionized calcium, alkaline phosphate, intact PTH, 25-OH vitamin D and serum creatinine were normal. MRI showed 5.7x3.7x3 cm lobulated heterogenous hypointensity near left greater trochanter. This mass was excised and showed tissue calcification. 99Tc-MDP bone scan and bone density parameters were normal. CBCT scan maxilla and mandible showed generalized hypercementosis with irregular root morphology. Serum C-terminal FGF 23 was 2401 RU/ml (normal: <150). NGS revealed normal FGF23 and Alpha Klotho genes. Exon 5 of GALNT3 gene showed homozygous mutation at Leu 366 Arg. GALNT3 Leu 366 Arg mutation causing HFTC has been described as compound heterozygous.Conclusion: This is the first case report of homozygous mutation at Leu366Arg causing HFTC.

GALNT8 suppresses breast cancer cell metastasis potential by regulating EGFR O-GalNAcylation

Breast cancer represents the most lethal malignancy that threatens the health of females. Metastasis is the fatal hallmark of breast cancer, and current effective therapeutic targets of metastasis are still lacking. Aberrant O-GalNAcylation, which is attributed to alteration of polypeptide N-acetylgalactosaminyl transferases (GALNTs), has been implicated in cancer metastasis. However, GALNTs that drive metastasis in breast cancer and their underlying mechanisms are largely unclear. In the present study, a negative correlation between GALNT8 and the prognosis of breast cancer patients was observed in multiple groups of Gene Expression Omnibus (GEO) datasets. We then constructed a stable GALNT8 knockdown MCF7 cell line and performed transcriptome analysis using RNA sequencing, which revealed that the expression of multiple migration-related genes was changed. GALNT8 was identified as a regulator of epithelial-mesenchymal transition (EMT) markers, including E-cadherin, N-cadherin, ZO-1 and vimentin. Moreover, loss- and gain-of-function GALNT8 assays demonstrated that this glycosyltransferase inhibited the metastatic potential of breast cancer cells. Interestingly, the O-GalNAcylation of EGFR, which is the key factor related to the metastasis cascade, was impacted by GALNT8. Furthermore, our results suggested that the GALNT8-mediated O-GalNAcylation led to the suppression of the EGFR signaling pathway and metastatic potential in breast cancer cells. These results suggested that GALNT8 acts as a tumor suppressor, represses tumor metastasis and inhibits the EMT process through the EGFR signaling pathway. This finding may provide insight into the mechanism by which aberrant O-glycosylation modulates breast cancer metastasis.

Novel Quinic Acid Glycerates from Tussilago farfara Inhibit Polypeptide GalNAc-Transferase.

The discovery of a bioactive inhibitor tool for human polypeptide N-acetylgalactosaminyl transferases (GalNAc-Ts), the initiating enzyme for mucin-type O-glycosylation, remains challenging. In the present study, we identified an array of quinic acid derivatives, including four new glycerates (1-4) from Tussilago farfara, a traditional Chinese medicinal plant, as active inhibitors of GalNAc-T2 using a combined screening approach with a cell-based T2-specific sensor and purified enzyme assay. These inhibitors dose-dependently inhibited human GalNAc-T2 but did not affect O-linked N-acetylglucosamine transferase (OGT), the other type of glycosyltransferase. Importantly, they are not cytotoxic and retain inhibitory activity in cells lacking elongated O-glycans, which are eliminated by the CRISPR/Cas9 gene editing tool. A structure-activity relationship study unveiled a novel quinic acid-caffeic acid conjugate pharmacophore that directs inhibition. Overall, these new natural product inhibitors could serve as a basis for developing an inhibitor tool for GalNAc-T2.

Brain‐region‐specific, glycosylation‐related transcriptomic alterations in Alzheimer's disease

Glycosylation aberrations have been found to be associated with Alzheimer's Disease (AD). In this study, three large public datasets that include RNA-seq data were used to investigate which glycosylation pathways are differentially expressed in AD patients in the brain. RNA-Seq data for six brain regions from hundreds of control, AD, and other neurodegenerative disease cases from Mayo RNASeq, MSBB, and ROSMAP were analyzed. The differential expression of glycosylation-related mRNAs between control and disease groups was analyzed for each brain region using the negative binomial model and a quasi-likelihood F-test using the edgeR package. The TMM method was used for normalization and the Benjamini-Hochberg method was used for multiple testing correction. The total number of genes involved in glycosylation or proteoglycans for each brain region varied from 457 to 467. The number of genes found to be differentially expressed exclusively in controls vs. AD varied between brain regions, with the dorsolateral prefrontal cortex (DLPFC), anterior prefrontal cortex (aPFC), primary auditory cortex, and temporal cortex (TC) having 83, 105, 61, and 99 differentially expressed genes respectively, while in Broca's area and the perirhinal cortex, only 12 and 27 genes were differential respectively. Polypeptide N-acetylgalactosaminyltransferase like 6 (GALNTL6), a gene involved in the initiation of O-glycosylation, was found differentially expressed exclusively in AD in four brain regions (adjusted p-value): DLPFC (p=2.88e-6), TC (p=0.00346), aPFC (p=0.000507) and primary auditory cortex (p=0.00446). GALNTL6 was down-regulated in DLPFC (logFC=-0.29) and TC (logFC=-0.40) and up-regulated in aPFC (logFC=0.24) and primary auditory cortex (logFC=0.27). Alpha-2,3-sialyltransferase 3 (ST3GAL3) and alpha-2,6-sialyltransferase 2 (ST6GAL2) are involved in the sialylation of both N- and O-glycans, and their mRNA was exclusively down-regulated in AD in DLPFC (ST3GAL3, logFC=-0.10, adjusted p-value=0.00275; ST6GAL2, logFC=-0.24, adjusted p-value=6.43-e-06) and TC (ST3GAL3, logFC=-0.22, adjusted p-value=0.000250; ST6GAL2, logFC=-0.41, adjusted p-value=1.30e-06). mRNAs for other O-glycan biosynthesis genes, such as GALNT17, POMT2, GXYLT1, and B3GAT2 were altered in AD in multiple brain regions. Interestingly several enzymes involved in O- and N- glycosylation were significantly down regulated in the dorsolateral prefrontal cortex and temporal cortex, areas significantly impaired in AD. These findings may facilitate future development of AD therapeutics.

GALNTL6 Rs558129: A Novel Polymorphism for Swimming Performance?

The enzyme polypeptide N-acetylgalactosaminyltransferase like 6, encoded by the GALANTL6 gene, plays a role in the gut microbiome regarding regulation of short-chain fatty acids and their anti-inflammatory and resynthesis functions. It was hypothesized that the T allele of the GALNTL6 rs558129 polymorphism could have a positive effect on anaerobic metabolism. Thus, this study was performed to investigate the association between GALNTL6 rs558129 polymorphism and athletic performance in swimmers. A total of 147 Polish short distance (SDS) and 49 long distance swimmers (LDS) of national or international competitive levels and 379 controls were genotyped using the real-time polymerase chain reaction (real-time PCR). We found that the carriers of the T allele (CT+TT) had a 1.56 times higher chance of being SDS (odds ratio (OR): 95%CI 1.06-2.29) than the CC homozygotes. The T allele was overrepresented in the SDS compared with controls (33.7% vs. 25.7%, p = 0.025, OR 1.40, 95% CI 1.04-1.87), but no statistically significant differences were found for LDS. This study provides evidence for an association between the GALNTL6 rs558129 polymorphism and short distance swimming athlete status. Although more replication studies are needed, the preliminary data suggest an opportunity to use the analysis of GALNTL6 polymorphism along with other variants of candidate genes and standard phenotypic assessment in power-oriented sports selection.

Role of GALNT4 in protecting against cardiac hypertrophy through ASK1 signaling pathway

Pathological myocardial hypertrophy is regulated by multiple pathways. However, its underlying pathogenesis has not been fully explored. The goal of this work was to elucidate the function of polypeptide N-acetylgalactosaminyltransferase 4 (GALNT4) in myocardial hypertrophy and its underlying mechanism of action. We illustrated that GALNT4 was upregulated in the models of hypertrophy. Two cardiac hypertrophy models were established through partial transection of the aorta in GALNT4-knockout (GALNT4-KO) mice and adeno-associated virus 9-GALNT4 (AAV9-GALNT4) mice. The GALNT4-KO mice demonstrated accelerated cardiac hypertrophy, dysfunction, and fibrosis, whereas the opposite phenotype was observed in AAV9-GALNT4 mice. Similarly, GALNT4 overexpression mitigated the degree of phenylephrine-induced cardiomyocyte hypertrophy in vitro whereas GALNT4 knockdown aggravated the hypertrophy. In terms of mechanism, GALNT4 deficiency increased the phosphorylation and activation of ASK1 and its downstream targets (JNK and p38), whereas GALNT4 overexpression inhibited activation of the ASK1 pathway. Furthermore, we demonstrated that GALNT4 can directly bind to ASK1 inhibiting its N-terminally mediated dimerization and the subsequent phosphorylation of ASK1. Finally, an ASK1 inhibitor (iASK1) was able to reverse the effects of GALNT4 in vitro. In summary, GALNT4 may serve as a new regulatory factor and therapeutic target by blocking the activation of the ASK1 signaling cascade.

Investigation of sequon engineering for improved O-glycosylation by the human polypeptide N-acetylgalactosaminyl transferase T2 isozyme and two orthologues.

We have been developing bacterial expression systems for human mucin-type O-glycosylation on therapeutic proteins, which is initiated by the addition of α-linked GalNAc to serine or threonine residues by enzymes in the GT-27 family of glycosyltransferases. Substrate preference across different isoforms of this enzyme is influenced by isoform-specific amino acid sequences at the site of glycosylation, which we have exploited to engineer production of Core 1 glycan structures in bacteria on human therapeutic proteins. Using RP-HPLC with a novel phenyl bonded phase to resolve intact protein glycoforms, the effect of sequon mutation on O-glycosylation initiation was examined through in vitro modification of the naturally O-glycosylated human interferon α-2b, and a sequon engineered human growth hormone. As part of the development of our glycan engineering in the bacterial expression system we are surveying various orthologues of critical enzymes to ensure complete glycosylation. Here we present an in vitro enzyme kinetic profile of three related GT-27 orthologues on natural and engineered sequons in recombinant human interferon α2b and human growth hormone where we show a significant change in kinetic properties with the amino acid changes. It was found that optimizing the protein substrate amino acid sequence using Isoform Specific O-Glycosylation Prediction (ISOGlyP, http://isoglyp.utep.edu/index.php) resulted in a measurable increase in kcat/KM, thus improving glycosylation efficiency. We showed that the Drosophila orthologue showed superior activity with our human growth hormone designed sequons compared with the human enzyme.

LncRNA PSMA3-AS1 promotes cell proliferation, migration, and invasion in ovarian cancer by activating the PI3K/Akt pathway via the miR-378a-3p/GALNT3 axis.

The crucial roles of the long noncoding RNAs (lncRNAs) in the development of ovarian cancer (OC) have been extensively studied. According to the prediction result from the Kaplan-Meier Plotter database, high expression of lncRNA proteasome subunit α type-3 antisense RNA1 (PSMA3-AS1) is associated with the poor prognosis in patients with OC. Thus, the study aimed to investigate the role of lncRNA PSMA3-AS1 in OC. Reverse transcription quantitative polymerase chain reaction analysis revealed that PSMA3-AS1 expression was significantly upregulated in OC cells and tissues. PSMA3-AS1 silencing inhibited OC cell proliferation, migration, and invasion, as shown by results of cell counting kit-8, colony formation, wound healing, and Transwell assays, respectively. Additionally, PSMA3-AS1 deficiency suppressed tumor growth in vivo. Mechanistically, luciferase reporter and RNA pulldown assays implied that PSMA3-AS1 served as a competing endogenous RNA for miR-378a-3p to upregulate the expression of polypeptide N-acetylgalactosaminyltransferase 3 (GALNT3). GALNT3 was a target gene of miR-378a-3p in OC. Moreover, PSMA3-AS1 activated the PI3K/Akt pathway by upregulating GALNT3 expression. Overall, PSMA3-AS1 promotes OC cell proliferation, migration, invasion, and xenograft tumor growth by activating the PI3K/Akt pathway via the miR-378a-3p/GALNT3 axis.

Polypeptide N-acetylgalactosaminyltransferase-Associated Phenotypes in Mammals.

Mucin-type O-glycosylation involves the attachment of glycans to an initial O-linked N-acetylgalactosamine (GalNAc) on serine and threonine residues on proteins. This process in mammals is initiated and regulated by a large family of 20 UDP-GalNAc: polypeptide N-acetylgalactosaminyltransferases (GalNAc-Ts) (EC 2.4.1.41). The enzymes are encoded by a large gene family (GALNTs). Two of these genes, GALNT2 and GALNT3, are known as monogenic autosomal recessive inherited disease genes with well characterized phenotypes, whereas a broad spectrum of phenotypes is associated with the remaining 18 genes. Until recently, the overlapping functionality of the 20 members of the enzyme family has hindered characterizing the specific biological roles of individual enzymes. However, recent evidence suggests that these enzymes do not have full functional redundancy and may serve specific purposes that are found in the different phenotypes described. Here, we summarize the current knowledge of GALNT and associated phenotypes.