Role Of Glyceraldehyde-3-phosphate Dehydrogenase Research Articles

Identification and expression profiling of GAPDH family genes involved in response to Sclerotinia sclerotiorum infection and phytohormones in Brassica napus.

Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is a crucial enzyme in glycolysis, an essential metabolic pathway for carbohydrate metabolism across all living organisms. Recent research indicates that phosphorylating GAPDH exhibits various moonlighting functions, contributing to plant growth and development, autophagy, drought tolerance, salt tolerance, and bacterial/viral diseases resistance. However, in rapeseed (Brassica napus), the role of GAPDHs in plant immune responses to fungal pathogens remains unexplored. In this study, 28 genes encoding GAPDH proteins were revealed in B. napus and classified into three distinct subclasses based on their protein structural and phylogenetic relationships. Whole-genome duplication plays a major role in the evolution of BnaGAPDHs. Synteny analyses revealed orthologous relationships, identifying 23, 26, and 26 BnaGAPDH genes with counterparts in Arabidopsis, Brassica rapa, and Brassica oleracea, respectively. The promoter regions of 12 BnaGAPDHs uncovered a spectrum of responsive elements to biotic and abiotic stresses, indicating their crucial role in plant stress resistance. Transcriptome analysis characterized the expression profiles of different BnaGAPDH genes during Sclerotinia sclerotiorum infection and hormonal treatment. Notably, BnaGAPDH17, BnaGAPDH20, BnaGAPDH21, and BnaGAPDH22 exhibited sensitivity to S. sclerotiorum infection, oxalic acid, hormone signals. Intriguingly, under standard physiological conditions, BnaGAPDH17, BnaGAPDH20, and BnaGAPDH22 are primarily localized in the cytoplasm and plasma membrane, with BnaGAPDH21 also detectable in the nucleus. Furthermore, the nuclear translocation of BnaGAPDH20 was observed under H2O2 treatment and S. sclerotiorum infection. These findings might provide a theoretical foundation for elucidating the functions of phosphorylating GAPDH.

Glyceraldehyde 3-Phosphate Dehydrogenase on the Surface of Candida albicans and Nakaseomyces glabratus Cells-A Moonlighting Protein That Binds Human Vitronectin and Plasminogen and Can Adsorb to Pathogenic Fungal Cells via Major Adhesins Als3 and Epa6.

Candida albicans and other closely related pathogenic yeast-like fungi carry on their surface numerous loosely adsorbed "moonlighting proteins"-proteins that play evolutionarily conserved intracellular functions but also appear on the cell surface and exhibit additional functions, e.g., contributing to attachment to host tissues. In the current work, we characterized this "moonlighting" role for glyceraldehyde 3-phosphate dehydrogenase (GAPDH, EC 1.2.1.12) of C. albicans and Nakaseomyces glabratus. GAPDH was directly visualized on the cell surface of both species and shown to play a significant part in the total capacity of fungal cells to bind two selected human host proteins-vitronectin and plasminogen. Using purified proteins, both host proteins were found to tightly interact with GAPDH, with dissociation constants in an order of 10-8 M, as determined by bio-layer interferometry and surface plasmon resonance measurements. It was also shown that exogenous GAPDH tightly adheres to the surface of candidal cells, suggesting that the cell surface location of this moonlighting protein may partly result from the readsorption of its soluble form, which may be present at an infection site (e.g., due to release from dying fungal cells). The major dedicated adhesins, covalently bound to the cell wall-agglutinin-like sequence protein 3 (Als3) and epithelial adhesin 6 (Epa6)-were suggested to serve as the docking platforms for GAPDH in C. albicans and N. glabratus, respectively.

Glyceraldehyde-3-phosphate dehydrogenase regulates activation of c-Jun N-terminal kinase under oxidative stress

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) acts as a sensor under oxidative stress, leading to induction of various biological responses. Given that mitogen-activated protein kinase (MAPK) signaling pathways mediate cellular responses to a wide variety of stimuli, including oxidative stress, here, we aimed to elucidate whether a cross-talk cascade between GAPDH and MAPKs occurs under oxidative stress. Of the three typical MAPKs investigated—extracellular signal-regulated kinase, p38, and c-Jun N-terminal kinase (JNK)—we found that hydrogen peroxide (H2O2)-induced JNK activation is significantly reduced in HEK293 cells treated with small-interfering (si)RNA targeting GAPDH. Co-immunoprecipitation with a GAPDH antibody further revealed protein–protein interactions between GAPDH and JNK in H2O2-stmulated cells. Notably, both JNK activation and these interactions depend on oxidation of the active-site cysteine (Cys152) in GAPDH, as demonstrated by rescue experiments with either exogenous wild-type GAPDH or the cysteine-substituted mutant (C152A) in endogenous GAPDH-knockdown HEK293 cells. Moreover, H2O2-induced translocation of Bcl-2-associated X protein (Bax) into mitochondria, which occurs downstream of JNK activation, is attenuated by endogenous GAPDH knockdown in HEK293 cells. These results suggest a novel role for GAPDH in the JNK signaling pathway under oxidative stress.

Mechanism of inactivation of glyceraldehyde-3-phosphate dehydrogenase in the presence of methylglyoxal

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is known to be one of the targets of methylglyoxal (MGO), a metabolite of glycolysis that increased in diabetes. However, the mechanism of GAPDH inactivation in the presence of MGO is unclear. The purpose of the work was to study the reaction of GAPDH with MGO and to identify the products of the reaction. It was shown that incubation of recombinant human GAPDH with MGO leads to irreversible inactivation of the enzyme, which is accompanied by a decrease in SH-group content by approximately 3.3 per tetramer GAPDH. MALDI-TOF MS analysis showed that the modification of GAPDH with MGO results in the oxidation of the catalytic cysteine residues (Cys152) to form cysteine-sulfinic acid. In addition, 2 arginine residues (R80 and R234) were identified that react with MGO to form hydroimidazolones. Incubation of SH-SY5Y neuroblastoma cells with MGO resulted in the inactivation of GAPDH and inhibition of glycolysis. The mechanism of GAPDH oxidation in the presence of MGO suggests the participation of superoxide anion, which is formed during the reaction of amino groups with methylglyoxal. The role of GAPDH in protection against the damaging effect of ROS in cells in the case of inefficiency of MGO removal by the GSH-dependent glyoxalase system is discussed.

GAPDH in neuroblastoma: Functions in metabolism and survival.

Neuroblastoma is a pediatric cancer of neural crest cells. It develops most frequently in nerve cells around the adrenal gland, although other locations are possible. Neuroblastomas rely on glycolysis as a source of energy and metabolites, and the enzymes that catalyze glycolysis are potential therapeutic targets for neuroblastoma. Furthermore, glycolysis provides a protective function against DNA damage, and there is evidence that glycolysis inhibitors may improve outcomes from other cancer treatments. This mini-review will focus on glyceraldehyde 3-phosphate dehydrogenase (GAPDH), one of the central enzymes in glycolysis. GAPDH has a key role in metabolism, catalyzing the sixth step in glycolysis and generating NADH. GAPDH also has a surprisingly diverse number of localizations, including the nucleus, where it performs multiple functions, and the plasma membrane. One membrane-associated function of GAPDH is stimulating glucose uptake, consistent with a role for GAPDH in energy and metabolite production. The plasma membrane localization of GAPDH and its role in glucose uptake have been verified in neuroblastoma. Membrane-associated GAPDH also participates in iron uptake, although this has not been tested in neuroblastoma. Finally, GAPDH activates autophagy through a nuclear complex with Sirtuin. This review will discuss these activities and their potential role in cancer metabolism, treatment and drug resistance.

Streptococcus agalactiae glyceraldehyde-3-phosphate dehydrogenase (GAPDH) elicits multiple cytokines from human cells and has a minor effect on bacterial persistence in the murine female reproductive tract

ABSTRACT Streptococcus agalactiae glyceraldehyde 3-phosphate dehydrogenase (GAPDH), encoded by gapC, is a glycolytic enzyme that is associated with virulence and immune-mediated protection. However, the role of GAPDH in cellular cytokine responses to S. agalactiae, bacterial phagocytosis and colonization of the female reproductive tract, a central host niche, is unknown. We expressed and studied purified recombinant GAPDH (rGAPDH) of S. agalactiae in cytokine elicitation assays with human monocyte-derived macrophage, epithelial cell, and polymorphonuclear leukocyte (PMN) co-culture infection models. We also generated a S. agalactiae mutant that over-expresses GAPDH (oeGAPDH) from gapC using a constitutively active promoter, and analyzed the mutant in murine macrophage antibiotic protection assays and in virulence assays in vivo, using a colonization model that is based on experimental infection of the reproductive tract in female mice. Human cell co-cultures produced interleukin (IL)-1β, IL-6, macrophage inflammatory protein (MIP)-1, tumor necrosis factor (TNF)-α and IL-10 within 24 h of exposure to rGAPDH. PMNs were required for several of these cytokine responses. However, over-expression of GAPDH in S. agalactiae did not significantly affect measures of phagocytic uptake compared to an empty vector control. In contrast, oeGAPDH-S. agalactiae showed a small but statistically significant attenuation for persistence in the reproductive tract of female mice during the chronic phase of infection (10–28 days post-inoculation), relative to the vector control. We conclude that S. agalactiae GAPDH elicits production of multiple cytokines from human cells, and over-expression of GAPDH renders the bacterium more susceptible to host clearance in the female reproductive tract. One-sentence summary: This study shows Streptococcus agalactiae glyceraldehyde 3-phosphate dehydrogenase, an enzyme that functions in glycolysis, gluconeogenesis and virulence, modifies phagocytosis outcomes, including cytokine synthesis, and affects bacterial persistence in the female reproductive tract.

Glyceraldehyde-3-phosphate dehydrogenase and glutamine synthetase inhibition in the presence of pro-inflammatory cytokines contribute to the metabolic imbalance of diabetic retinopathy

Diabetic retinopathy (DR) is the leading cause of vision impairment in working age adults. In addition to hyperglycemia, retinal inflammation is an important driving factor for DR development. Although DR is clinically described as diabetes-induced damage to the retinal blood vessels, several studies have reported that metabolic dysregulation occurs in the retina prior to the development of microvascular damage. The two most commonly affected metabolic pathways in diabetic conditions are glycolysis and the glutamate pathway. We investigated the role of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and glutamine synthetase (GS) in an in-vitro model of DR incorporating high glucose and pro-inflammatory cytokines. We found that GAPDH and GS enzyme activity were not significantly affected in hyperglycemic conditions or after exposure to cytokines alone, but were significantly decreased in the DR model. This confirmed that pro-inflammatory cytokines IL-1β and TNFα enhance the hyperglycemic metabolic deficit. We further investigated metabolite and amino acid levels after specific pharmacological inhibition of GAPDH or GS in the absence/presence of pro-inflammatory cytokines. The results indicate that GAPDH inhibition increased glucose and addition of cytokines increased lactate and ATP levels and reduced glutamate levels. GS inhibition did not alter retinal metabolite levels but the addition of cytokines increased ATP levels and caused glutamate accumulation in Müller cells. We conclude that it is the action of pro-inflammatory cytokines concomitantly with the inhibition of the glycolytic or GS mediated glutamate recycling that contribute to metabolic dysregulation in DR. Therefore, in the absence of good glycemic control, therapeutic interventions aimed at regulating inflammation may prevent the onset of early metabolic imbalance in DR.

Products of S-nitrosylation of glyceraldehyde-3-phosphate dehydrogenase: Relation between S-nitrosylation and oxidation

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is one of the major targets of NO in cells, especially in neurodegenerative diseases. S-Nitrosylation of GAPDH is accompanied by its translocation into the nucleus with subsequent apoptosis. The product of GAPDH modification by NO is considered to be S-nitrosylated GAPDH (GAPDH-SNO). However, this has not been confirmed by direct methods.Products of GAPDH modification in the presence of the NO donor diethylamine NONOate were analyzed by MALDI- and ESI- mass spectrometry methods.The adduct between GAPDH and dimedone was detected by MALDI-MS analysis after incubation of S-nitrosylated GAPDH with dimedone, which points to the formation of cysteine-sulfenic acid (GAPDH-SOH) in the protein. Analysis of the protein hydrolysate revealed the incorporation of dimedone into the catalytic residue Cys150. An additional peak that corresponded to GAPDH-SNO was detected by ESI-MS analysis in GAPDH after the incubation with the NO donor. The content of GAPDH-SNO and GAPDH-SOH in the modified GAPDH was evaluated by different approaches and constituted 2.3 and 0.7 mol per mol GAPDH, respectively. A small fraction of GAPDH was irreversibly inactivated after NO treatment, suggesting that a minor part of the products includes cysteine-sulfinic or cysteine-sulfonic acids.The main products of GAPDH modification by NO are GAPDH-SNO and GAPDH-SOH that is presumably formed due to the hydrolysis of GAPDH-SNO.The obtained results are important for understanding the molecular mechanism of redox regulation of cell functions and the role of GAPDH in the development of neurodegenerative disorders.

Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol

BackgroundClostridium thermocellum is a promising microorganism for conversion of cellulosic biomass to biofuel, without added enzymes; however, the low ethanol titer produced by strains developed thus far is an obstacle to industrial application.ResultsHere, we analyzed changes in the relative concentration of intracellular metabolites in response to gradual addition of ethanol to growing cultures. For C. thermocellum, we observed that ethanol tolerance, in experiments with gradual ethanol addition, was twofold higher than previously observed in response to a stepwise increase in the ethanol concentration, and appears to be due to a mechanism other than mutation. As ethanol concentrations increased, we found accumulation of metabolites upstream of the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) reaction and depletion of metabolites downstream of that reaction. This pattern was not observed in the more ethanol-tolerant organism Thermoanaerobacterium saccharolyticum. We hypothesize that the Gapdh enzyme may have different properties in the two organisms. Our hypothesis is supported by enzyme assays showing greater sensitivity of the C. thermocellum enzyme to high levels of NADH, and by the increase in ethanol tolerance and production when the T. saccharolyticum gapdh was expressed in C. thermocellum.ConclusionsWe have demonstrated that a metabolic bottleneck occurs at the GAPDH reaction when the growth of C. thermocellum is inhibited by high levels of ethanol. We then showed that this bottleneck could be relieved by expression of the gapdh gene from T. saccharolyticum. This enzyme is a promising target for future metabolic engineering work.

Lipid transfer protein CaMBP10 binds to glyceraldehyde-3-phosphate dehydrogenase and modulates its activity

Lipid transfer proteins (LTPs) are a protein family found in plants with a variety of functions. In addition to lipid binding, LTPs also bind to calmodulin and Ca2+-dependent protein kinase (CDPK), which are calcium signal transducers. For the first time, we identified glyceraldehyde-3- phosphate dehydrogenase (GAPDH) as a novel binding protein of LTP-CaMBP10 in Chinese cabbage. This binding was confirmed using multiple biochemical approaches. The effects of this interaction on GAPDH activity were assessed for both recombinant and endogenous GAPDH proteins. LTP-CaMBP10 does not appear to affect nicotinamide adenine dinucleotide (NAD)-dependent GAPDH activity. In contrast, it significantly suppresses nicotinamide adenine dinucleotide phosphate (NADPH) consumption by GAPDH in a dosage-dependent manner. This result indicated a specific role of GAPDH in regulating LTP functions and implicating crosstalk between LTP-dependent and GAPDH-dependent biological events.

Abstract 386: The Role of GAPDH in the Cardio-metabolic Response to Ischemia

Fundamental changes in nutrient metabolism underlie many disease processes, including obesity, diabetes, and cancer. Metabolic derangements are also a hallmark of heart disease, both in the context of failure and ischemia, and likely contributors to its etiology. For example, while the healthy heart favors fats as a fuel, failing hearts will consume more glucose; but it is unclear if this is pathological or protective. It is very likely that manipulation of metabolic pathways can affect, and possibly augment and protect, heart function. Yet no drug therapy exists that targets cardiac metabolism. Staples of ischemic disease management largely focus on anti-platelet, cholesterol-lowering, and beta-adrenergic blocking regimens. Heart failure medicines focus on maladaptive neuro-hormonal responses, a strategy several decades old. Our goal here is to delineate exactly how critical aspects of metabolism are altered in cardiac ischemia, and to identify new targets of pharmacological intervention. Metabolism largely remains unexplored because of its intricate network of regulation, compounded by the fact that variations in this network will often escape traditional methods of gene expression profiling. But by using high throughput metabolomic and proteomic platforms, we have striking evidence that the glycolytic enzyme glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is instrumental in the heart’s response to ischemia. Dynamic, real-time metabolomic flux using heavy carbon isotope labeling points toward a partial shift of glycolytic intermediates toward lipid synthesis during ischemic stress in cardiomyocytes, precisely at the step catalyzed by GAPDH. Expansive phospho-proteomic analyses from Langendorff ex vivo hanging heart experiments show that this shift in metabolism may be triggered by an inhibitory phosphorylation of GAPDH. Our hypothesis that glycolysis is in part rerouted toward triglyceride synthesis is a revolutionary departure from decades-old perceptions of energy management. Modulating GAPDH function, either directly or indirectly via partner proteins, may offer insight into untapped areas of therapy. Our early attempts with chemicals that inhibit GAPDH already show heart cells with improved survival under ischemia.

Abstract 2367: High expression of GAPDH is relevant to lung adenocarcinoma with low expression of the tumor suppressor gene Deleted in Liver Cancer-1 (DLC1)

Abstract Although glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is often considered to be a stable housekeeping marker gene, we recently determined that high levels of GAPDH mRNA expression in solid tumors was highly correlated with the expression of a group of genes, most of which are classified as cell cycle-dependent (Wang et al. PLoS One, 2013). In the current project, we have further evaluated the role of GAPDH in lung adenocarcinoma. Using data from an integrated microarray gene expression dataset and The Cancer Genome Atlas (TCGA) Project, we have now found a correlation between high mRNA expression of GAPDH and low expression of the tumor suppressor gene Deleted in Liver Cancer-1 (DLC1). DLC1 encodes a protein with strong RhoGAP (GTPase activating protein) activity and is inactivated in various human malignancies. In lung adenocarcinoma, low DLC1 expression occurred frequently together with up-regulation of GAPDH. The correlation coefficient of GAPDH and DLC1 expression in the integrated gene expression dataset was -0.6. In addition, high GAPDH and low DLC1 expression in lung cancer was associated with unfavorable prognosis. The combination of DLC1 expression level and GAPDH level improved the predictive prognostic power compared with that of the individual gene level alone. Pathway analysis indicated that low DLC1 in lung cancer was accompanied by increased expression of genes implicated in glycolysis and reduced expression of fructose-1,6-bisphosphatase (FBP1) in gluconeogenesis. Our findings suggest that GAPDH may contribute to the regulation of cell cycle genes and of tumor suppressor genes in cancer. Citation Format: Dunrui Wang, Guangxian Zhang. High expression of GAPDH is relevant to lung adenocarcinoma with low expression of the tumor suppressor gene Deleted in Liver Cancer-1 (DLC1). [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2367. doi:10.1158/1538-7445.AM2014-2367

The glyceraldehyde 3-phosphate dehydrogenase gene (GAPDH) is regulated by myeloid zinc finger 1 (MZF-1) and is induced by calcitriol

Recently, new tissue-specific functions for glyceraldehyde 3-phosphate dehydrogenase (GAPDH) have been discovered, aside from its archetypal function in glycolysis. This casts doubt on the legitimacy of using GAPDH as a normalization control for gene expression analysis. We report the binding of the myeloid zinc finger-1 (MZF-1) transcription factor to the human GAPDH promoter. Furthermore, we show that up-regulation of MZF-1 by 1,25-dihydroxyvitamin D3 (calcitriol) induces GAPDH in HS-5 stromal fibroblasts, while knockdown of MZF1 by shRNA leads to a concomitant reduction in GAPDH expression. This argues that MZF-1 regulates GAPDH, indicating a role for GAPDH in calcitriol-mediated signaling.

Identification of novel GAPDH-derived antimicrobial peptides secreted by Saccharomyces cerevisiae and involved in wine microbial interactions

Saccharomyces cerevisiae plays a primordial role in alcoholic fermentation and has a vast worldwide application in the production of fuel-ethanol, food and beverages. The dominance of S. cerevisiae over other microbial species during alcoholic fermentations has been traditionally ascribed to its higher ethanol tolerance. However, recent studies suggested that other phenomena, such as microbial interactions mediated by killer-like toxins, might play an important role. Here we show that S. cerevisiae secretes antimicrobial peptides (AMPs) during alcoholic fermentation that are active against a wide variety of wine-related yeasts (e.g. Dekkera bruxellensis) and bacteria (e.g. Oenococcus oeni). Mass spectrometry analyses revealed that these AMPs correspond to fragments of the S. cerevisiae glyceraldehyde 3-phosphate dehydrogenase (GAPDH) protein. The involvement of GAPDH-derived peptides in wine microbial interactions was further sustained by results obtained in mixed cultures performed with S. cerevisiae single mutants deleted in each of the GAPDH codifying genes (TDH1-3) and also with a S. cerevisiae mutant deleted in the YCA1 gene, which codifies the apoptosis-involved enzyme metacaspase. These findings are discussed in the context of wine microbial interactions, biopreservation potential and the role of GAPDH in the defence system of S. cerevisiae.

Properties and functional diversity of glyceraldehyde-3-phosphate dehydrogenase

For a long time glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was considered a classical glycolytic protein of little interest. It was also used as a model protein for analysis of protein structure and enzyme mechanisms. However, recent evidence demonstrates that GAPDH from mammalian cells displays a number of diverse activities unrelated to its glycolytic function. This enzyme is an example of moonlighting protein. Dehydrogenase participates in membrane fusion, microtubule assembly, vesicular transport, and the maintenance of DNA integrity. New and novel studies indicate that enzyme is directly involved in transcriptional, posttranscriptional gene regulation, and the maintenance of chromatin structure. Furthermore, other studies also indicate a role of GAPDH in apoptosis, and age-related neurodegenerative disease e.g. Alzheimer's, Huntington's and Parkinson's diseases. This work describes the structure and localization of GAPDH in cells as well as the latest discoveries on the multifunctional properties of the enzyme.

Direct Inhibition of Tombusvirus Plus-Strand RNA Synthesis by a Dominant Negative Mutant of a Host Metabolic Enzyme, Glyceraldehyde-3-Phosphate Dehydrogenase, in Yeast and Plants

The replication of plus-strand RNA viruses depends on many cellular factors. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is an abundant metabolic enzyme that is recruited to the replicase complex of Tomato bushy stunt virus (TBSV) and affects asymmetric viral RNA synthesis. To further our understanding on the role of GAPDH in TBSV replication, we used an in vitro TBSV replication assay based on recombinant p33 and p92(pol) viral replication proteins and cell-free yeast extract. We found that the addition of purified recombinant GAPDH to the cell extract prepared from GAPDH-depleted yeast results in increased plus-strand RNA synthesis and asymmetric production of viral RNAs. Our data also demonstrate that GAPDH interacts with p92(pol) viral replication protein, which may facilitate the recruitment of GAPDH into the viral replicase complex in the yeast model host. In addition, we have identified a dominant negative mutant of GAPDH, which inhibits RNA synthesis and RNA recruitment in vitro. Moreover, this mutant also exhibits strong suppression of tombusvirus accumulation in yeast and in virus-infected Nicotiana benthamiana. Overall, the obtained data support the model that the co-opted GAPDH plays a direct role in TBSV replication by stimulating plus-strand synthesis by the viral replicase.

Nuclear GAPDH: changing the fate of Müller cells in diabetes

Müller cells, the primary glial cells are a crucial component of the retinal tissue performing a wide range of functions including maintaining the blood-retinal barrier. Several studies suggest that diabetes leads to Müller cell dysfunction and loss. The pathophysiology of hyperglycemia-induced cellular injury of Müller cells remains only poorly understood. Recently, the concept that translocation of the predominantly cytosolic glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) to the nucleus and its accumulation in this cellular compartment alters transcriptional events associated with cell death induction has gained major interest. High glucose conditions induce nuclear translocation and accumulation of GAPDH in the nucleus of Müller cells in vivo and in vitro. With regards to Müller cell dysfunction, the effects of nuclear accumulation of GAPDH are multifaceted. Considering the functional versatility of GAPDH including gene regulation, DNA repair, telomere protection, etc., it is of immense importance to explore possible GAPDH actions to unravel the mysteries around the role of GAPDH in hyperglycemia-induced cellular changes in order to develop novel therapeutic strategies. Therefore, this review focuses on the molecular events associated with the nuclear translocation of GAPDH and how it affects the fate of Müller cells in diabetes.

The role of glyceraldehyde 3-phosphate dehydrogenase (GapA-1) in Neisseria meningitidis adherence to human cells

BackgroundGlyceraldehyde 3-phosphate dehydrogenases (GAPDHs) are cytoplasmic glycolytic enzymes, which although lacking identifiable secretion signals, have also been found localized to the surface of several bacteria (and some eukaryotic organisms); where in some cases they have been shown to contribute to the colonization and invasion of host tissues. Neisseria meningitidis is an obligate human nasopharyngeal commensal which can cause life-threatening infections including septicaemia and meningitis. N. meningitidis has two genes, gapA-1 and gapA-2, encoding GAPDH enzymes. GapA-1 has previously been shown to be up-regulated on bacterial contact with host epithelial cells and is accessible to antibodies on the surface of capsule-permeabilized meningococcal cells. The aims of this study were: 1) to determine whether GapA-1 was expressed across different strains of N. meningitidis; 2) to determine whether GapA-1 surface accessibility to antibodies was dependant on the presence of capsule; 3) to determine whether GapA-1 can influence the interaction of meningococci and host cells, particularly in the key stages of adhesion and invasion.ResultsIn this study, expression of GapA-1 was shown to be well conserved across diverse isolates of Neisseria species. Flow cytometry confirmed that GapA-1 could be detected on the cell surface, but only in a siaD-knockout (capsule-deficient) background, suggesting that GapA-1 is inaccessible to antibody in in vitro-grown encapsulated meningococci. The role of GapA-1 in meningococcal pathogenesis was addressed by mutational analysis and functional complementation. Loss of GapA-1 did not affect the growth of the bacterium in vitro. However, a GapA-1 deficient mutant showed a significant reduction in adhesion to human epithelial and endothelial cells compared to the wild-type and complemented mutant. A similar reduction in adhesion levels was also apparent between a siaD-deficient meningococcal strain and an isogenic siaD gapA-1 double mutant.ConclusionsOur data demonstrates that meningococcal GapA-1 is a constitutively-expressed, highly-conserved surface-exposed protein which is antibody-accessible only in the absence of capsule. Mutation of GapA-1 does not affect the in vitro growth rate of N. meningitidis, but significantly affects the ability of the organism to adhere to human epithelial and endothelial cells in a capsule-independent process suggesting a role in the pathogenesis of meningococcal infection.

Glyceraldehyde-3-Phosphate Dehydrogenase Regulates Endothelin-1 Expression by a Novel, Redox-Sensitive Mechanism Involving mRNA Stability

The regulation of the synthesis of the endothelial-derived vasoconstrictor endothelin-1 (ET-1) is a complex process encompassing transcriptional as well as mRNA stability mechanisms. We have described recently the existence of a mechanism for the control of ET-1 expression based on the mRNA-destabilizing capacity of specific cytosolic proteins through interaction with AU-rich elements (AREs) present in the 3' untranslated region of the gene. We now identify glyceraldehyde-3'-phosphate dehydrogenase (GAPDH) as a protein which binds to the AREs and is responsible for the destabilization of the mRNA. Oxidant stress alters the binding of GAPDH to the mRNA and its capacity to modulate ET-1 expression, a phenomenon occurring through specific S glutathionylation of the catalytically active residue Cys 152. Finally, we provide data consistent with a role for GAPDH in mRNA unwinding, yielding this molecule more prone to degradation. In contrast, S-thiolated GAPDH appears unable to modify mRNA unwinding, thus facilitating enhanced stability. Taken together, these results describe a novel, redox-based mechanism regulating mRNA stability and add a new facet to the panoply of GAPDH cellular homeostatic actions.

Role of glyceraldehyde 3-phosphate dehydrogenase in the development and progression of diabetic retinopathy.

OBJECTIVE—Mitochondrial superoxide levels are elevated in the retina in diabetes, and manganese superoxide dismutase overexpression prevents the development of retinopathy. Superoxide inhibits glyceraldehyde-3-phosphate dehydrogenase (GAPDH), which activates major pathways implicated in diabetic complications, including advanced glycation end products (AGEs), protein kinase C, and hexosamine pathway. Our aim is to investigate the role of GAPDH in the development and progression of diabetic retinopathy and to elucidate the mechanism.RESEARCH DESIGN AND METHODS—Rats with streptozotocin-induced diabetes were in a state of poor control (GHb >11%) for 12 months, good control (GHb <7) soon after induction of diabetes, or poor control for 6 months with 6 months’ good control. Retinal GAPDH, its ribosylation and nitration, AGEs, and PKC activation were determined and correlated with microvascular histopathology.RESULTS—In rats with poor control, retinal GAPDH activity and expressions were subnormal with increased ribosylation and nitration (25–30%). GAPDH activity was subnormal in both cytosol and nuclear fractions, but its protein expression and nitration were significantly elevated in nuclear fraction. Reinstitution of good control failed to protect inactivation of GAPDH, its covalent modification, and translocation to the nucleus. PKC, AGEs, and hexosamine pathways remained activated, and microvascular histopathology was unchanged. However, GAPDH and its translocation in good control rats were similar to those in normal rats.CONCLUSIONS—GAPDH plays a significant role in the development of diabetic retinopathy and its progression after cessation of hyperglycemia. Thus, therapies targeted toward preventing its inhibition may inhibit development of diabetic retinopathy and arrest its progression.