Pyramiding D-lactate dehydrogenase with the glyoxalase pathway enhances abiotic stress tolerance in plants.

Methylglyoxal is a common cytotoxic metabolite produced in plants during multiple biotic and abiotic stress. To mitigate the toxicity of MG, plants utilize the glyoxalase pathway comprising glyoxalase I (GLYI), glyoxalase II (GLYII), or glyoxalase III (GLYIII). GLYI and GLYII are the key enzymes of glyoxalase pathways that play an important role in abiotic stress tolerance. Earlier research showed that MG level is lower when both GLYI and GLYII are overexpressed together, compared to GLYI or GLYII single gene overexpressed transgenic plants. D-lactate dehydrogenase (D-LDH) is an integral part of MG detoxification which metabolizes the end product (D-lactate) of the glyoxalase pathway. In this study, two Arabidopsis transgenic lines were constructed using gene pyramiding technique: GLYI and GLYII overexpressed (G-I + II), and GLYI, GLYII, and D-LDH overexpressed (G-I + II + D) plants. G-I + II + D exhibits lower MG and D-lactate levels and enhanced abiotic stress tolerance than the G-I + II and wild-type plants. Further study explores the stress tolerance mechanism of G-I + II + D plants through the interplay of different regulators and plant hormones. This, in turn, modulates the expression of ABA-dependent stress-responsive genes like RAB18, RD22, and RD29B to generate adaptive responses during stress. Therefore, there might be a potential correlation between ABA and MG detoxification pathways. Furthermore, higher STY46, GPX3, and CAMTA1 transcripts were observed in G-I + II + D plants during abiotic stress. Thus, our findings suggest that G-I + II + D has significantly improved MG detoxification, reduced oxidative stress-induced damage, and provided a better protective mechanism against abiotic stresses than G-I + II or wild-type plants.

Association between GLO1 variants and gestational diabetes mellitus susceptibility in a Chinese population: a preliminary study.

Background: Glyoxalase 1 (GLO1) plays a crucial role in defending against glycation. Single nucleotide polymorphism (SNP) variants in the GLO1 gene may affect gene expression and alter enzyme activity. However, there have been limited studies evaluating the association between GLO1 and diabetes, especially gestational diabetes mellitus (GDM). Therefore, this study is the first to explore the association of GLO1 SNPs and GDM risk. Methods: The study included a total of 500 GDM patients and 502 control subjects. The SNPscan™ genotyping assay was used to genotype rs1781735, rs4746 and rs1130534. To assess the disparities in genotype, allele, and haplotype distributions and their correlation with GDM risk, the independent sample t-test, logistic regression, and chi-square test were employed during the data processing phase. Furthermore, one-way ANOVA was conducted to determine the differences in genotype and blood glucose and methylglyoxal(MG) levels. Results: Significant differences were observed in prepregnancy body mass index (pre-BMI), age, systolic blood pressure (SBP), diastolic blood pressure (DBP), and parity between GDM and healthy subjects (P < 0.05). After adjusting for these factors, GLO1 rs1130534 TA remained associated with an increased risk of GDM (TA vs. TT + AA: OR = 1.320; 95% CI: 1.008-1.728; P = 0.044), especially in the pre-BMI ≥ 24 subgroup (TA vs. TT + AA: OR = 2.424; 95% CI: 1.048-5.607; P = 0.039), with fasting glucose levels being significantly elevated in the TA genotype compared to the TT genotype (P < 0.05). Conversely, the GLO1 rs4746 TG was associated with a decreased risk of GDM (TG vs. TT: OR = 0.740; 95% CI: 0.548-0.999; P = 0.049; TG vs. TT + GG: OR = 0.740; 95% CI: 0.548-0.998; P = 0.048). Additionally, the haplotype T-G-T of rs1781735, rs4746 and rs1130534 was associated with a decreased risk of GDM among individuals with a pre-BMI ≥ 24 (OR = 0.423; 95% CI: 0.188-0.955; P = 0.038). Furthermore, the rs1781735 GG genotype was found to be more closely related to maternal MG accumulation and neonatal weight gain (P < 0.05). Conclusion: Our findings suggested that GLO1 rs1130534 was associated with an increased susceptibility to GDM and higher blood glucose levels, but GLO1 rs4746 was associated with a decreased risk of GDM. The rs1781735 has been associated with the accumulation of maternal MG and subsequent weight gain in neonates.

The genetic polymorphisms and activity of glyoxalase 1 as a risk factor for acute coronary syndrome in South Indians with type 2 diabetes mellitus.

OBJECTIVE: The individuals' genetic traits predispose them to a higher or lower risk of Type 2 diabetes mellitus (T2DM) and its complications, for example, acute coronary syndrome (ACS). As carbonyl stress is responsible for the pathogenesis and complications of T2DM, and glyoxalase 1 (GLO1) is the most crucial determinant of carbonyl stress, the study aimed to explore the association between GLO1 gene polymorphism, GLO1 activity in red blood cell (RBC), plasma methylglyoxal (MG) levels, and ACS risk in South Indian T2DM patients. METHODS: A total of 150 T2DM patients with ACS as cases and 150 T2DM patients without ACS as controls were recruited in a case-control study. The rs4746, rs1049346 and rs1130534 of the GLO1 gene were analysed using TaqMan allele discrimination assay. The RBC GLO1 activity and plasma MG levels were measured. RESULTS: Significantly lower RBC GLO1 activity and higher plasma MG levels were found in cases compared to controls (p < 0.001 and p = 0.008, respectively). The genotype and allele frequencies of rs1049346 significantly differed between cases and controls (p < 0.001). For rs1130534 and rs1049346, no significant difference was found. For rs1049346, the TT and CC genotypes were associated with higher (p = 0.002) and lower (p = 0.001) ACS risk, respectively, in various genetic models. The TT genotype of rs1049346 was associated with lower RBC GLO1 activity (p = 0.004) and higher MG level (p = 0.010). In haplotype analysis, higher ACS susceptibility with the TAT haplotype (p < 0.001) and lower ACS susceptibility with the TAC haplotype (p < 0.001) were observed. Also, lower RBC GLO1 activity was associated with the TAT haplotype (p = 0.002). CONCLUSIONS: The rs1049346 of the GLO1 gene may be associated with ACS risk in South Indian T2DM patients, and the T and C allele might be essential precipitating and protective factors, respectively.

Proline-mediated activation of glyoxalase II improve methylglyoxal detoxification in Oryza sativa L. under chromium injury: Clarification via vector analysis of enzymatic activities and gene expression.

Environmental factors affect plants in several ways including the excessive accumulation of methylglyoxal (MG), resulting in dysfunctions of many biological processes. Exogenous proline (Pro) application is one of the successful strategies to increase plant tolerance against various environmental stresses, including chromium (Cr). This study highlights the alleviation role of exogenous Pro on MG detoxification in rice plants induced by Cr(Vl) through modifying the expression of glyoxalase I (Gly I)- and glyoxalase II (Gly II)-related genes. The MG content in rice roots was significantly reduced by Pro application under Cr(VI) stress, however, there was little effect on the MG content in shoots. In this connection, the vector analysis was used to compare the involvement of Gly l and Gly II on MG detoxification in 'Cr(VI)' and 'Pro+Cr(VI)' treatments. Results exhibited that vector strength in rice roots increased with an increase in Cr concentrations, while there was a negligible difference in the shoots. The comparative analysis demonstrated that the vector strengths in roots under 'Pro+Cr(VI)' treatments were higher than 'Cr(VI)' treatments, suggesting that Pro improved Gly II activity more efficiently to reduce MG content in roots. Calculation of the gene expression variation factors (GEFs) indicated a positive effect of Pro application on the expression of Gly I and Gly ll-related genes, wherein a stronger impact was in roots than the shoots. Together, the vector analysis and gene expression data reveal that exogenous Pro chiefly improved Gly ll activity in rice roots which subsequently enhanced MG detoxification under Cr(VI) stress.

Overexpressing glyoxalase 1 attenuates acute hyperglycemia-exacerbated neurological deficits of ischemic stroke in mice.

Stress-induced hyperglycemia (SIH) is associated with poor functional recovery and high mortality in patients with acute ischemic stroke (AIS). However, intensive controlling of blood glucose by using insulin was not beneficial in patients with AIS and acute hyperglycemia. This study investigated the therapeutic effects of the overexpression of glyoxalase I (GLO1), a detoxifying enzyme of glycotoxins, on acute hyperglycemia-aggravated ischemic brain injury.  In the present study, adeno-associated viral (AAV)-mediated GLO1 overexpression reduced infarct volume and edema level but did not improve neurofunctional recovery in the mice with middle cerebral artery occlusion (MCAO). AAV-GLO1 infection significantly enhanced neurofunctional recovery in the MCAO mice with acute hyperglycemia but not in the mice with normoglycemia. Methylglyoxal (MG)-modified proteins expression significantly increased in the ipsilateral cortex of the MCAO mice with acute hyperglycemia. AAV-GLO1 infection attenuated the induction of MG-modified proteins, ER stress formation, and caspase 3/7 activation in MG-treated Neuro-2A cells, and reductions in synaptic plasticity and microglial activation were mitigated in the injured cortex of the MCAO mice with acute hyperglycemia. Treatment with ketotifen, a potent GLO1 stimulator, after surgery, alleviated neurofunctional deficits and ischemic brain damage in the MCAO mice with acute hyperglycemia.  Altogether, our data substantiate that, in ischemic brain injury, GLO1 overexpression can alleviate pathologic alterations caused by acute hyperglycemia. Upregulation of GLO1 may be a therapeutic strategy for alleviating SIH-aggravated poor functional outcomes in patients with AIS.

Metformin inhibits methylglyoxal-induced retinal pigment epithelial cell death and retinopathy via AMPK-dependent mechanisms: Reversing mitochondrial dysfunction and upregulating glyoxalase 1.

Diabetic retinopathy (DR) is a major cause of blindness in adult, and the accumulation of advanced glycation end products (AGEs) is a major pathologic event in DR. Methylglyoxal (MGO), a highly reactive dicarbonyl compound, is a precursor of AGEs. Although the therapeutic potential of metformin for retinopathy disorders has recently been elucidated, possibly through AMPK activation, it remains unknown how metformin directly affects the MGO-induced stress response in retinal pigment epithelial cells. Therefore, in this study, we compared the effects of metformin and the AMPK activator A769662 on MGO-induced DR in mice, as well as evaluated cytotoxicity, mitochondrial dynamic changes and dysfunction in ARPE-19 cells. We found MGO can induce mitochondrial ROS production and mitochondrial membrane potential loss, but reduce cytosolic ROS level in ARPE-19 cells. Although these effects of MGO can be reversed by both metformin and A769662, we demonstrated that reduction of mitochondrial ROS production rather than restoration of cytosolic ROS level contributes to cell protective effects of metformin and A769662. Moreover, MGO inhibits AMPK activity, reduces LC3II accumulation, and suppresses protein and gene expressions of MFN1, PGC-1α and TFAM, leading to mitochondrial fission, inhibition of mitochondrial biogenesis and autophagy. In contrast, these events of MGO were reversed by metformin in an AMPK-dependent manner as evidenced by the effects of compound C and AMPK silencing. In addition, we observed an AMPK-dependent upregulation of glyoxalase 1, a ubiquitous cellular enzyme that participates in the detoxification of MGO. In intravitreal drug-treated mice, we found that AMPK activators can reverse the MGO-induced cotton wool spots, macular edema and retinal damage. Functional, histological and optical coherence tomography analysis support the protective actions of both agents against MGO-elicited retinal damage. Metformin and A769662 via AMPK activation exert a strong protection against MGO-induced retinal pigment epithelial cell death and retinopathy. Therefore, metformin and AMPK activator can be therapeutic agents for DR.

Genome-Wide Identification and Functional Characterization of Stress Related Glyoxalase Genes in Brassica napus L.

Rapeseed (Brassica napus L.) is not only one of the most important oil crops in the world, but it is also an important vegetable crop with a high value nutrients and metabolites. However, rapeseed is often severely damaged by adverse stresses, such as low temperature, pathogen infection and so on. Glyoxalase I (GLYI) and glyoxalase II (GLYII) are two enzymes responsible for the detoxification of a cytotoxic metabolite methylglyoxal (MG) into the nontoxic S-D-lactoylglutathione, which plays crucial roles in stress tolerance in plants. Considering the important roles of glyoxalases, the GLY gene families have been analyzed in higher plans, such as rice, soybean and Chinese cabbage; however, little is known about the presence, distribution, localizations and expression of glyoxalase genes in rapeseed, a young allotetraploid. In this study, a total of 35 BnaGLYI and 30 BnaGLYII genes were identified in the B. napus genome and were clustered into six and eight subfamilies, respectively. The classification, chromosomal distribution, gene structure and conserved motif were identified or predicted. BnaGLYI and BnaGLYII proteins were mainly localized in chloroplast and cytoplasm. By using publicly available RNA-seq data and a quantitative real-time PCR analysis (qRT-PCR), the expression profiling of these genes of different tissues was demonstrated in different developmental stages as well as under stresses. The results indicated that their expression profiles varied among different tissues. Some members are highly expressed in specific tissues, BnaGLYI11 and BnaGLYI27 expressed in flowers and germinating seed. At the same time, the two genes were significantly up-regulated under heat, cold and freezing stresses. Notably, a number of BnaGLY genes showed responses to Plasmodiophora brassicae infection. Overexpression of BnGLYI11 gene in Arabidopsis thaliana seedlings confirmed that this gene conferred freezing tolerance. This study provides insight of the BnaGLYI and BnaGLYII gene families in allotetraploid B. napus and their roles in stress resistance, and important information and gene resources for developing stress resistant vegetable and rapeseed oil.

Role of lactoyl-glutathione lyase of Salmonella in the colonization of plants under salinity stress.

Salmonella, a foodborne human pathogen, can colonize the members of the kingdom Plantae. However, the basis of the persistence of Salmonella in plants is largely unknown. Plants encounter various biotic and abiotic stress agents in soil. We conjectured that methylglyoxal (MG), one of the common metabolites that accumulate in plants during both biotic and abiotic stress, plays a role in regulating the plant-Salmonella interaction. The interaction of Salmonella Typhimurium with plants under salinity stress was investigated. It was observed that wild-type Salmonella Typhimurium can efficiently colonize the root, but mutant bacteria lacking MG detoxifying enzyme, lactoyl-glutathione lyase (Lgl), showed lower colonization in roots exclusively under salinity stress. This colonization defect is due to the poor viability of the mutated bacterial strains under these conditions. This is the first report to prove the role of MG-detoxification genes in the colonization of stressed plants and highlights the possible involvement of metabolic genes in the evolution of the plant-associated life of Salmonella.

Loss of glyoxalase 2 alters the glucose metabolism in zebrafish.

Glyoxalase 2 is the second enzyme of the glyoxalase system, catalyzing the detoxification of methylglyoxal to d-lactate via SD-Lactoylglutathione. Recent in vitro studies have suggested Glo2 as a regulator of glycolysis, but if Glo2 regulates glucose homeostasis and related organ specific functions in vivo has not yet been evaluated. Therefore, a CRISPR-Cas9 knockout of glo2 in zebrafish was created and analyzed. Consistent with its function in methylglyoxal detoxification, SD-Lactoylglutathione, but not methylglyoxal accumulated in glo2-/- larvae, without altering the glutathione metabolism or affecting longevity. Adult glo2-/- livers displayed a reduced hexose concentration and a reduced postprandial P70-S6 kinase activation, but upstream postprandial AKT phosphorylation remained unchanged. In contrast, glo2-/- skeletal muscle remained metabolically intact, possibly compensating for the dysfunctional liver through increased glucose uptake and glycolytic activity. glo2-/- zebrafish maintained euglycemia and showed no damage of the retinal vasculature, kidney, liver and skeletal muscle. In conclusion, the data identified Glo2 as a regulator of cellular energy metabolism in liver and skeletal muscle, but the redox state and reactive metabolite accumulation were not affected by the loss of Glo2.

H3K27 acetylation activated long noncoding RNA RP11-162G10.5 promotes breast cancer progression via the YBX1/GLO1 axis.

PURPOSE: Long noncoding RNAs (lncRNAs) orchestrate critical roles in human tumorigenesis. However, the regulatory mechanism of lncRNAs in tissue-specific expressions in breast cancer (BC) remains poorly understood. This study aims to investigate lncRNA role and mechanisms in BC. METHODS: RNA sequencing was used to explore differentially expressed lncRNAs in BC and adjacent tissues. H3K27 acetylation (H3K27ac) chromatin immune-precipitation sequencing (ChIP-seq) data of BC cells from the GEO dataset (GSE85158) was retrieved to identify the H3K27ac activated lncRNAs that were involved in tumorigenesis. RP11-162G10.5 was selected as the target lncRNA for further functional and mechanism study. RESULTS: In this study, we identified a novel lncRNA RP11-162G10.5, whose overexpression was specifically driven by H3K27ac in luminal breast cancer. And increased RP11-162G10.5 in BC is correlated with poor patient outcomes. RP11-162G10.5 promotes tumor cell proliferation in vitro and in vivo. Mechanistically, RP11-162G10.5 recruits transcriptional factor YBX1 to the GLO1 promoter, consequently activating GLO1 transcription to modulate the progression of BC. CONCLUSIONS: Our findings suggest that the histone modification-activated lncRNA contributes to the oncogenesis of BC. Also, our data reveal a role for RP11-162G10.5 in BC tumorigenesis and may supply a strategy for targeting the RP11-162G10.5 as a potential biomarker and a therapeutic target for breast cancer patients.

Role of glyoxalase I and II in somatic and spermatogenic testicular cells during the postnatal development of the domestic cat.

Like humans, many felid species suffer from teratozoospermia and frequently produce low numbers of normal spermatozoa. Male fertility can be affected by oxidative and dicarbonyl stress. Because of the high level of glycolytic activity in testes, reactive dicarbonyl metabolites may arise as side-products of glycolysis; their generation is further promoted by oxidative stress. Alpha-oxoaldehydes, including methylglyoxal (MG), are reactive dicarbonyl metabolites and substrates for the formation of advanced glycation end products. Elevated levels of both may lead to dicarbonyl stress and cause cellular dysfunction. However, MG and other α-oxoaldehydes can be converted to less dangerous molecules via the glyoxalase pathway. In this pathway, α-oxoaldehydes react with glutathione (GSH), forming a thioacetal, which becomes metabolized by glyoxalase I (GLO I) to S-D-lactoyl-glutathione (SLG). Glyoxalase II (GLO II) converts SLG to d-lactate upon the release of GSH. Nothing is known about the glyoxalase system in the feline testis and its capacity to mitigate an excess of dicarbonyl metabolites. To study whether GLO I and GLO II are present and have a specific function in the testis of the domestic cat, the gene expression of both enzymes were analyzed in testis samples of different developmental stages (prepubertal, pubertal, postpubertal). Furthermore, the presence of GLO I and GLO II proteins was investigated via immunohistochemistry. The GLO I gene expression does not change between developmental stages. Immunohistochemistry revealed strong signals for GLO I in the cytoplasm and nuclei of Sertoli and Leydig cells during all developmental stages. GLO I was described as catalyzing the rate-limiting step in the glyoxalase pathway. This implies a function on the part of this enzyme of sustaining the homeostasis of somatic testicular cells. For GLO II, we observed stage-dependent mRNA expression, which was significantly increased after puberty. In accordance with this observation, clear immunohistochemical GLO II signals were observed in nuclei of individual germ cells. The most intense signals were visible in spermatocytes. The different localizations of the strong GLO I and GLO II signals indicate that GLO II, in addition to the classical glyoxalase pathway, may have additional functions in meiotic germ cells, for example, providing lactate as an energy substrate and/or GSH as an antioxidant. Moreover, protein functions may be modulated via S-glutathionylation.

Viridiplantae-specific GLXI and GLXII isoforms co-evolved and detoxify glucosone in planta.

Reactive carbonyl species (RCS) such as methylglyoxal (MGO) and glyoxal (GO) are highly reactive, unwanted side-products of cellular metabolism maintained at harmless intracellular levels by specific scavenging mechanisms.MGO and GO are metabolized through the glyoxalase (GLX) system, which consists of two enzymes acting in sequence, GLXI and GLXII. While plant genomes encode a number of different GLX isoforms, their specific functions and how they arose during evolution are unclear. Here, we used Arabidopsis (Arabidopsis thaliana) as a model species to investigate the evolutionary history of GLXI and GLXII in plants and whether the GLX system can protect plant cells from the toxicity of RCS other than MGO and GO. We show that plants possess two GLX systems of different evolutionary origins and with distinct structural and functional properties. The first system is shared by all eukaryotes, scavenges MGO and GO, especially during seedling establishment, and features Zn2+-type GLXI proteins with a metal cofactor preference that were present in the last eukaryotic common ancestor. GLXI and GLXII of the second system, featuring Ni2+-type GLXI, were acquired by the last common ancestor of Viridiplantae through horizontal gene transfer from proteobacteria and can together metabolize keto-D-glucose (KDG, glucosone), a glucose-derived RCS, to D-gluconate. When plants displaying loss-of-function of a Viridiplantae-specific GLXI were grown in KDG, D-gluconate levels were reduced to 10%-15% of those in the wild type, while KDG levels showed an increase of 48%-67%. In contrast to bacterial GLXI homologs, which are active as dimers, plant Ni2+-type GLXI proteins contain a domain duplication, are active as monomers, and have a modified second active site. The acquisition and neofunctionalization of a structurally, biochemically, and functionally distinct GLX system indicates that Viridiplantae are under strong selection to detoxify diverse RCS.

A glutathione-independent DJ-1/PfpI domain-containing tomato glyoxalaseIII2, SlGLYIII2, confers enhanced tolerance under salt and osmotic stresses.

In plants, glyoxalase enzymes are activated under stress conditions to mitigate the toxic effects of hyperaccumulated methylglyoxal (MG), a highly reactive carbonyl compound. Until recently, a glutathione-dependent bi-enzymatic pathway involving glyoxalase I (GLYI) and glyoxalase II (GLYII) was considered the primary MG-detoxification system. Recently, a new glutathione-independent glyoxalase III (GLYIII) mediated direct route was also reported in plants. However, the physiological significance of this new pathway remains to be elucidated across plant species. This study identified the full complement of 22 glyoxalases in tomato. Based on their strong induction under multiple abiotic stresses, SlGLYI4, SlGLYII2 and SlGLYIII2 were selected candidates for further functional characterisation. Stress-inducible overexpression of both glutathione-dependent (SlGLYI4 + SlGLYII2) and independent (SlGLYIII2) pathways led to enhanced tolerance in both sets of transgenic plants under abiotic stresses. However, SlGLYIII2 overexpression (OE) plants outperformed the SlGLYI4 + SlGLYII2 OE counterparts for their stress tolerance under abiotic stresses. Further, knockdown of SlGLYIII2 resulted in plants with exacerbated stress responses than those silenced for both SlGLYI4 and SlGLYII2. The superior performance of SlGLYIII2 OE tomato plants for better growth and yield under salt and osmotic treatments could be attributed to better GSH/GSSG ratio, lower reactive oxygen species levels, and enhanced antioxidant potential, indicating a prominent role of GLYIII MG-detoxification pathway in abiotic stress mitigation in this species.

The Human Glyoxalase Gene Family in Health and Disease.

The glyoxalase gene family consists of six structurally and functionally diverse enzymes with broad roles in metabolism. The common feature that defines this family is based on structural motifs that coordinate divalent cations which are required for activity. These family members have been implicated in a variety of physiological processes, including amino-acid metabolism (4-hydroxyphenylpyruvate dioxygenase; HPD), primary metabolism (methylmalonyl-CoA epimerase; MCEE), and aldehyde detoxication (glyoxalase 1; GLO1) and therefore have significant associations with disease. A central function of this family is the detoxification of reactive dicarbonyls (e.g., methylglyoxal), which react with cellular nucleophiles, resulting in the modification of lipids, proteins, and DNA. These damaging modifications activate canonical stress responses such as heat shock, unfolded protein, antioxidant, and DNA damage responses. Thus, glyoxalases serve an important role in homeostasis, preventing the pathogenesis of metabolic disease states, including obesity, diabetes, cardiovascular disease, renal failure, and aging. This review presents a thorough overview of the literature surrounding this diverse enzyme class. Although extensive literature exists for some members of this family (e.g., GLO1), little is known about the physiological role of glyoxalase domain-containing protein 4 (GLOD4) and 5 (GLOD5), paving the way for exciting avenues for future research.

Genome-Wide Identification of Cassava Glyoxalase I Genes and the Potential Function of MeGLYâ -13 in Iron Toxicity Tolerance.

Glyoxalase I (GLYI) is a key enzyme in the pathway of the glyoxalase system that degrades the toxic substance methylglyoxal, which plays a crucial part in plant growth, development, and stress response. A total of 19 GLYI genes were identified from the cassava genome, which distributed randomly on 11 chromosomes. These genes were named MeGLYI-1-19 and were systematically characterized. Transcriptome data analysis showed that MeGLYIs gene expression is tissue-specific, and MeGLYI-13 is the dominant gene expressed in young tissues, while MeGLYI-19 is the dominant gene expressed in mature tissues and organs. qRT-PCR analysis showed that MeGLYI-13 is upregulated under 2 h excess iron stress, but downregulated under 6, 12, and 20 h iron stress. Overexpression of MeGLYI-13 enhanced the growth ability of transgenic yeast under iron stress. The root growth of transgenic Arabidopsis seedlings was less inhibited by iron toxicity than that of the wild type (WT). Potted transgenic Arabidopsis blossomed and podded under iron stress, but flowering of the WT was significantly delayed. The GLYI activity in transgenic Arabidopsis was improved under both non-iron stress and iron stress conditions compared to the WT. The SOD activity in transgenic plants was increased under iron stress, while the POD and CAT activity and MDA content were decreased compared to that in the WT. These results provide a basis for the selection of candidate genes for iron toxicity tolerance in cassava, and lay a theoretical foundation for further studies on the functions of these MeGLYI genes.

OsGLYI3, a glyoxalase gene expressed in rice seed, contributes to seed longevity and salt stress tolerance.

The glyoxalase pathway plays a vital role in the chemical detoxification of methylglyoxal (MG) in biological systems. Our previous study suggested that OsGLYI3 may be effective in seed natural aging. In this study, the rice OsGLYI3 gene was cloned and characterized as specifically expressed in the seed. The accelerated aging (AA) treatment results indicated significant roles of OsGLYI3 in seed longevity and vigor, as the seeds of the transgenic lines with overexpressed and knocked-out OsGLYI3 exhibited higher and lower germination, respectively. The AA treatment also increased the superoxide dismutase (SOD) activity in the overexpressed transgenic seeds compared to the wild-type seeds yet lowered the SOD activity in the CRISPR/Cas9-derived transgenic rice lines. Rice OsGLYI3 was markedly upregulated in response to NaCl induced stress conditions. Compared to wild-type plants, overexpressed transgenic rice lines exhibited increased GLYI activity, decreased MG levels and improved salt stress tolerance, while CRISPR/Cas9 knockout transgenic rice lines showed decreased glyoxalase I activity, increased MG levels, and greater sensitivity to stress treatments with NaCl. Collectively, our results confirmed for the first time that OsGLYI3 is specifically expressed in rice seeds and contributes to seed longevity and salt stress tolerance.

Methylglyoxal and glyoxalase 1-a metabolic stress pathway-linking hyperglycemia to the unfolded protein response and vascular complications of diabetes.

The study of the glyoxalase system by Thornalley and co-workers in clinical diabetes mellitus and correlation with diabetic complications revealed increased exposure of patients with diabetes to the reactive, dicarbonyl metabolite methylglyoxal (MG). Twenty-eight years later, extended and built on by Thornalley and co-workers and others, the glyoxalase system is an important pathway contributing to the development of insulin resistance and vascular complications of diabetes. Other related advances have been: characterization of a new kind of metabolic stress-'dicarbonyl stress'; identification of the major physiological advanced glycation endproduct (AGE), MG-H1; physiological substrates of the unfolded protein response (UPR); new therapeutic agents-'glyoxalase 1 (Glo1) inducers'; and a refined mechanism underlying the link of dysglycemia to the development of insulin resistance and vascular complications of diabetes.

Glyoxalase 1 knockdown induces age-related ß-cell dysfunction and glucose intolerance in mice.

Tight control of glycemia is a major treatment goal for type 2 diabetes mellitus (T2DM). Clinical studies indicated that factors other than poor glycemic control may be important in fostering T2DM progression. Increased levels of methylglyoxal (MGO) associate with complications development, but its role in the early steps of T2DM pathogenesis has not been defined. Here, we show that MGO accumulation induces an age-dependent impairment of glucose tolerance and glucose-stimulated insulin secretion in mice knockdown for glyoxalase 1 (Glo1KD). This metabolic alteration associates with the presence of insular inflammatory infiltration (F4/80-positive staining), the islet expression of senescence markers, and higher levels of cytokines (MCP-1 and TNF-α), part of the senescence-activated secretory profile, in the pancreas from 10-month-old Glo1KD mice, compared with their WT littermates. In vitro exposure of INS832/13 ß-cells to MGO confirms its casual role on ß-cell dysfunction, which can be reverted by senolytic treatment. These data indicate that MGO is capable to induce early phenotypes typical of T2D progression, paving the way for novel prevention approaches to T2DM.

Glyoxalase III enhances salinity tolerance through reactive oxygen species scavenging and reduced glycation.

Methylglyoxal (MG) is a metabolically generated highly cytotoxic compound that accumulates in all living organisms, from Escherichia coli to humans, under stress conditions. To detoxify MG, nature has evolved reduced glutathione (GSH)-dependent glyoxalase and NADPH-dependent aldo-keto reductase systems. But both GSH and NADPH have been reported to be limiting in plants under stress conditions, and thus detoxification might not be performed efficiently. Recently, glyoxalase III (GLY III)-like enzyme activity has been reported from various species, which can detoxify MG without any cofactor. In the present study, we have tested whether an E. coli gene, hchA, encoding a functional GLY III, could provide abiotic stress tolerance to living systems. Overexpression of this gene showed improved tolerance in E. coli and Saccharomyces cerevisiae cells against salinity, dicarbonyl, and oxidative stresses. Ectopic expression of the E. coli GLY III gene (EcGLY-III) in transgenic tobacco plants confers tolerance against salinity at both seedling and reproductive stages as indicated by their height, weight, membrane stability index, and total yield potential. Transgenic plants showed significantly increased glyoxalase and antioxidant enzyme activity that resisted the accumulation of excess MG and reactive oxygen species (ROS) during stress. Moreover, transgenic plants showed more anti-glycation activity to inhibit the formation of advanced glycation end product (AGE) that might prevent transgenic plants from stress-induced senescence. Taken together, all these observations indicate that overexpression of EcGLYIII confers salinity stress tolerance in plants and should be explored further for the generation of stress-tolerant plants.

ABI3- and PIF1-mediated regulation of GIG1 enhances seed germination by detoxification of methylglyoxal in Arabidopsis.

Methylglyoxal (MG) is a toxic by-product of the glycolysis pathway in most living organisms and was previously shown to inhibit seed germination. MG is detoxified by glyoxalase I and II family proteins in plants. MG is abundantly produced during early embryogenesis in Arabidopsis seeds. However, the mechanism that alleviates the toxic effect of MG in maturing seeds is poorly understood. In this study, by T-DNA mutant population screening, we found that mutations in a glyoxalase I gene (named GERMINATION-IMPAIRED GLYOXALASE 1, GIG1) led to significantly impaired germination compared with wild-type seeds. Transformation of full-length GIG1 cDNA under the constitutively active cauliflower mosaic virus 35S promoter in the gig1 background completely recovered the seed germination phenotype. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) analyses revealed that GIG1 is uniquely expressed in seeds and is upregulated by abscisic acid (ABA) and downregulated by gibberellic acid (GA) during seed germination. An ABA signaling component, ABI3, directly activated GIG1 in maturing seeds. In addition, PHYTOCHROME INTERACTING FACTOR 1 (PIF1) also plays cooperatively with ABI3 in the regulation of GIG1 expression in the early stage of imbibed seeds. Furthermore, GIG1 expression is stably silenced by epigenetic repressors such as polycomb repressor complexes. Altogether, our results indicate that light and ABA signaling cooperate to enhance seed germination by the upregulation of GIG1 to detoxify MG in maturing seeds.