Ectopic expression of a bacterial thiamin monophosphate kinase enhances vitamin B1 biosynthesis in plants.

Plants and bacteria have distinct pathways to synthesize the bioactive vitamin B1 thiamin diphosphate (TDP). In plants, thiamin monophosphate (TMP) synthesized in the TDP biosynthetic pathway is first converted to thiamin by a phosphatase, which is then pyrophosphorylated to TDP. In contrast, bacteria use a TMP kinase encoded by ThiL to phosphorylate TMP to TDP directly. The Arabidopsis THIAMIN REQUIRING2 (TH2)-encoded phosphatase is involved in TDP biosynthesis. The chlorotic th2 mutants have high TMP and low thiamin and TDP. Ectopic expression of Escherichia coli ThiL and ThiL-GFP rescued the th2-3 mutant, suggesting that the bacterial TMP kinase could directly convert TMP into TDP in Arabidopsis. These results provide direct evidence that the chlorotic phenotype of th2-3 is caused by TDP rather than thiamin deficiency. Transgenic Arabidopsis harboring engineered ThiL-GFP targeting to the cytosol, chloroplast, mitochondrion, or nucleus accumulated higher TDP than the wild type (WT). Ectopic expression of E. coli ThiL driven by the UBIQUITIN (UBI) promoter or an endosperm-specific GLUTELIN1 (GT1) promoter also enhanced TDP biosynthesis in rice. The pUBI:ThiL transgenic rice accumulated more TDP and total vitamin B1 in the leaves, and the pGT1:ThiL transgenic lines had higher TDP and total vitamin B1 in the seeds than the WT. Total vitamin B1 only increased by approximately 25-30% in the polished and unpolished seeds of the pGT1:ThiL transgenic rice compared to the WT. Nevertheless, these results suggest that genetic engineering of a bacterial vitamin B1 biosynthetic gene downstream of TMP can enhance vitamin B1 production in rice.

Bacterial lipopolysaccharide inhibits free thiamin uptake along the intestinal tract via interference with membrane expression of thiamin transporters 1 and 2.

This study examined the effect of exposure of small and large intestinal epithelial cells to the bacterial lipopolysaccharide (LPS) on uptake of free form of vitamin B1, i.e., thiamin. The intestinal tract encounters two sources of thiamin: diet and the gut microbiota. Absorption of thiamin in both the small and large intestine occurs via a carrier-mediated process that involves thiamin transporters 1 and 2 (THTR-1 and -2). Complementary in vitro (human duodenal epithelial HuTu-80 cells and human colonic epithelial NCM460 cells), in vivo (mice), and ex vivo (human primary differentiated enteroid and colonoid monolayers) models were used. The results showed that exposure to LPS causes a significant inhibition in carrier-mediated [3H]-thiamin uptake by small and large intestinal epithelia, with no change in the levels of expression of THTR-1 and -2 mRNAs and their total cellular proteins. However, a significant decrease in the fractions of the THTR-1 and -2 proteins that are expressed at the cell membranes of these epithelial cells was observed. These effects of LPS appeared to involve a protein kinase A (PKA) signaling pathway as activating this pathway caused a reversal in the inhibition of thiamin uptake and level of expression of its transporters at the cell membrane. These findings demonstrate that exposure of gut epithelia to LPS (a situation that occurs under different pathological conditions) leads to inhibition in thiamin uptake due to a decrease in level of expression of its transporters at the cell membrane that is likely mediated via a PKA signaling pathway. NEW & NOTEWORTHY This study shows that the exposure of gut epithelial cells to bacterial LPS negatively impact the uptake process of the free form of vitamin B1 (i.e., thiamin). This appears to be mediated via suppression in the level of thiamin transporters 1 and 2 (THTR-1 and -2) expression at the cell membrane and involves a protein kinase A (PKA) signaling pathway.

IQGAP-2: A novel interacting partner for the Human Colonic Thiamin Pyrophosphate Transporter (hcTPPT).

The human colonic thiamin pyrophosphate transporter (hcTPPT) mediates the uptake of the microbiota-generated and phosphorylated form of vitamin B1 (i. e., thiamin pyrophosphate) in the large intestine. Expression of hcTPPT along the absorptive tract is restricted to the large intestine and the transporter is exclusively localized at the apical membrane domain of the polarized epithelial cells/colonocytes. Previous studies have characterized different physiological/pathophysiological aspects of the hcTPPT system, but nothing is currently known on whether the transporter has interacting partner(s) that affects its physiology/biology. We addressed this issue using a Y2H to screen a human colonic cDNA library, and have identified 3 putative interactors, namely IQGAP-2, SNX-6 and DMXL-1. Focusing on IQGAP-2 (whose expression in human colonocytes is the highest), we found (using fluorescent microscopy imaging and co-immunoprecipitation approaches) the putative interactor to co-localize with hcTPPT, and to directly interact with the transporter. Also, over-expressing IQGAP-2 in NCM460 cells and in human primary differentiated colonoid monolayers was found to lead to significant (P < 0.01) induction in TPP uptake, while it's knocking down (using gene-specific siRNAs) caused significant (P < 0.01 & < 0.05) decrease in uptake. Furthermore, over-expressing IQGAP-2 in NCM460 cells was found to lead to a significant enhancement in hcTPPT protein stability. Finally, we found the expression of IQGAP-2 to be markedly suppressed in conditions/factors that negatively impact colonic TPP uptake. These results identify the IQGAP-2 as an interacting partner with the hcTPPT in human colonocytes and show that this interaction has physiological and biological consequences.

Intramolecular Cyclization and a Retro-Ene Reaction Enable the Rapid Fragmentation of a Vitamin B1-Derived Breslow Intermediate.

In solution, analogues of the Breslow intermediate formed during catalysis by benzoylformate decarboxylase (BFDC) undergo rapid, irreversible fragmentation. The ability of BFDC to prevent this reaction and preserve its cofactor is a striking example of an enzyme 'steering' a reactive intermediate towards a productive pathway. To understand how BFDC suppresses the off-pathway reactivity of this Breslow intermediate, a clear mechanistic understanding of the fragmentation reaction is required. Here, DFT calculations reveal an unexpected mechanism for the solution-phase fragmentation that involves an intramolecular cyclization and a subsequent retro-ene reaction to release the final products. Free energy profiles demonstrate that this pathway is significantly more facile than the previously proposed mechanism that invoked Breslow intermediate enolates as intermediates. Additional computations have been performed to understand why related Breslow intermediates do not undergo analogous fragmentation reactions. Calculations performed with two closely related Breslow intermediates suggest that subtle differences in the relative values of ΔG≠ for protonation and fragmentation dictate whether a given intermediate will fragment or not. These differences and the fragmentation mechanism unveiled in this work may have ramifications for the mechanism of BFDC and other thiamin-dependent enzymes and could provide general lessons related to the control of reactive intermediates by enzymes.

Green synthesis of self-oriented flower-like Ag@Ag2O nanostructures functionalized with L-Tryptophan for colorimetric simultaneous determination of ultra-trace level of thiamin and riboflavin.

The study focuses on the green synthesis of Ag@Ag2O nanostructures using Padina algae extract and functionalizing them with L-tryptophan to enhance their properties as a colorimetric sensor for simultaneous detection of ultra-trace levels of thiamin and riboflavin. The nanostructures are characterized using techniques like XRD, FESEM, FTIR, TEM, AFM, and DLS to understand their morphology, structure, and interactions with target molecules. FESEM analysis revealed the hierarchical flower-like Ag@Ag2O nanostructures. The TEM image shows the formation of core-shell nanostructures. Also, DLS analysis and surface zeta potential spectra illustrated the aggregated nature of fabricated nanocomposites in the presence of vitamins. The study is the first to report simultaneous determination of thiamin and riboflavin using a colorimetric sensor based on Ag@Ag2O-L-Try nanocomposites using partial leas square (PLS). The dynamic range of thiamin and riboflavin was achieved in 0.1 mol L-1 acetate buffer pH 4 and the ratio Ag@Ag2O: L-try 1:1. The Ag@Ag2O-L-Try sensor exhibited two linear ranges of 0.1- 1.0 and 3-350 µMol L- 1 for riboflavin and a linear range 3.0-60 µMol L- 1 for thiamin. Also, low detection limit of 1.92 µMol L- 1 and 0.048 µMol L- 1 was obtained for riboflavin and thiamin, respectively. The results indicated that the success of the method depends on the selective and sensitive colorimetric assay of the sensor along with the simultaneous determination by the PLS algorithm. Hence, the proposed technique can be used for the accurate and precise determination of vitamins in different pharmaceutical syrup and tablet samples.

Thiamin dynamics during the adult life cycle of Atlantic salmon (Salmo salar).

Thiamin is an essential water-soluble B vitamin known for its wide range of metabolic functions and antioxidant properties. Over the past decades, reproductive failures induced by thiamin deficiency have been observed in several salmonid species worldwide, but it is unclear why this micronutrient deficiency arises. Few studies have compared thiamin concentrations in systems of salmonid populations with or without documented thiamin deficiency. Moreover, it is not well known whether and how thiamin concentration changes during the marine feeding phase and the spawning migration. Therefore, samples of Atlantic salmon (Salmo salar) were collected when actively feeding in the open Baltic Sea, after the sea migration to natal rivers, after river migration, and during the spawning period. To compare populations of Baltic salmon with systems without documented thiamin deficiency, a population of landlocked salmon located in Lake Vänern (Sweden) was sampled as well as salmon from Norwegian rivers draining into the North Atlantic Ocean. Results showed the highest mean thiamin concentrations in Lake Vänern salmon, followed by North Atlantic, and the lowest in Baltic populations. Therefore, salmon in the Baltic Sea seem to be consistently more constrained by thiamin than those in other systems. Condition factor and body length had little to no effect on thiamin concentrations in all systems, suggesting that there is no relation between the body condition of salmon and thiamin deficiency. In our large spatiotemporal comparison of salmon populations, thiamin concentrations declined toward spawning in all studied systems, suggesting that the reduction in thiamin concentration arises as a natural consequence of starvation rather than to be related to thiamin deficiency in the system. These results suggest that factors affecting accumulation during the marine feeding phase are key for understanding the thiamin deficiency in salmonids.

[Wernicke encephalopathy : a neuro-ophthalmological side effect of bariatric surgery]. / L'encéphalopathie de Gayet-Wernicke : une complication neuro-ophtalmologique de la chirurgie bariatrique.

Due to an increase in the worldwide prevalence of obesity and the efficiency of bariatric surgery, this procedure is more often performed. Besides its benefits, it has also disadvantages and may be the cause of nutritional deficiencies. Thiamin deficiency is particularly important to diagnose and to treat early as it can lead to major sequelae and even to death. Wernicke's encephalopathy is the most frequent presentation associating confusion, ataxia, ophtalmoplegia and nystagmus. The full triad is not usually observed, which may lead to sub-diagnosis of this affection. The diagnosis is clinical, biological and radiologic thanks to the brain MRI. Intravenous thiamin supplementation therapy must be administered as fast as possible in order to avoid long-term damages. In the ophthalmological field, the potential sequelae are ophthalmoplegia, nystagmus and optic neuropathy. Therapeutics for nystagmus are pharmacological, surgical and/or optical. We illustrate this condition with a case report of an 18-year-old man developing Wernicke's encephalopathy as early as six weeks after a sleeve gastrectomy.

Suite à une augmentation de la prévalence de l'obésité dans le monde et à l'efficacité de la chirurgie bariatrique, cette technique est pratiquée de plus en plus fréquemment. Malgré ses avantages, elle n'est pas sans risque et peut être responsable de déficits nutritionnels multiples. Le déficit en vitamine B1 ou thiamine est particulièrement important à connaître et, à rapidement diagnostiquer en raison des nombreuses séquelles invalidantes, voire le décès du patient, dont il peut être responsable. Le tableau classique est l'encéphalopathie de Gayet-Wernicke associant confusion, ataxie et troubles oculomoteurs. Néanmoins, il n'est pas toujours complet, ce qui participe au sous-diagnostic de cette pathologie. Le diagnostic est clinique, biologique et/ou radiologique grâce à l'IRM cérébrale. La supplémentation vitaminique intraveineuse doit être instaurée le plus rapidement possible afin d'éviter des séquelles à long terme. D'un point de vue ophtalmologique, les séquelles potentielles sont les ophtalmoplégies, les nystagmus et les neuropathies optiques. Les thérapies envisageables du nystagmus, outre la supplémentation en thiamine en aigu, sont pharmacologiques, chirurgicales et/ou optiques. Nous illustrons cette pathologie par un cas clinique d'encéphalopathie de Gayet-Wernicke dès la 6ème semaine post-opératoire d'une chirurgie bariatrique de type «sleeve¼ chez un patient de 18 ans.

Bacterial lipopolysaccharide inhibits colonic carrier-mediated uptake of thiamin pyrophosphate: roles for TLR4 receptor and NF-κB/P38/JNK signaling pathway.

This study investigated the effect of the bacterial endotoxin lipopolysaccharide (LPS) on colonic uptake of thiamin pyrophosphate (TPP), the biologically active form of vitamin B1 that is generated by gut microbiota. We used three complementary models in our study: in vitro (human-derived colonic epithelial NCM460), ex vivo (human differentiated colonoid monolayers), and in vivo (mouse colonic tissue). The results showed that exposure of NCM460 cells to LPS leads to a significant inhibition of carrier-mediated TPP uptake as well as in decreased expression of the colonic TPP transporter (cTPPT) protein, mRNA, and heterologous nuclear RNA (hnRNA) compared with untreated controls. Similarly, exposure of human differentiated colonoid monolayers and mice to LPS caused significant inhibition in colonic carrier-mediated TPP uptake and in cTPPT protein, mRNA, and hnRNA expression. The effect of LPS on colonic TPP uptake and cTTPT expression was also found to be associated with a significant reduction in activity of the SLC44A4 promoter as well as in decreased expression of the nuclear factor Elf-3 (E74-like ETS transcription factor 3), which is needed for promoter activity. Finally, we found that knocking down the Toll-like receptor 4 (TLR4) and blocking the nuclear factor kappa B (NF-κB), JNK, and p38 signaling pathways with the use of pharmacological inhibitors lead to significant abrogation in the degree of LPS-mediated inhibition in TPP uptake and cTPPT expression. These results demonstrated that exposure of colonic epithelia to LPS inhibits colonic TPP uptake via transcriptional mechanism(s) and that the effect is mediated via TLR4 receptor and NF-κB/p38/JNK signaling pathways.NEW & NOTEWORTHY This study examined the effect of the bacterial lipopolysaccharide (LPS) on the colonic uptake of thiamin pyrophosphate (TPP), the biologically active form of vitamin B1. Three complementary models were used: in vitro (human NCM460 cells), ex vivo (human colonoids), and in vivo (mice). The results showed LPS to significantly suppress TPP uptake and the expression of its transporter, and that these effects are mediated via the membrane TLR4 receptor, and involve the NF-κB/p38/JNK signaling pathways.

A Possible Aquatic Origin of the Thiaminase TenA of the Human Gut Symbiont Bacteroides thetaiotaomicron.

TenA thiamin-degrading enzymes are commonly found in prokaryotes, plants, fungi and algae and are involved in the thiamin salvage pathway. The gut symbiont Bacteroides thetaiotaomicron (Bt) produces a TenA protein (BtTenA) which is packaged into its extracellular vesicles. An alignment of BtTenA protein sequence with proteins from different databases using the basic local alignment search tool (BLAST) and the generation of a phylogenetic tree revealed that BtTenA is related to TenA-like proteins not only found in a small number of intestinal bacterial species but also in some aquatic bacteria, aquatic invertebrates, and freshwater fish. This is, to our knowledge, the first report describing the presence of TenA-encoding genes in the genome of members of the animal kingdom. By searching metagenomic databases of diverse host-associated microbial communities, we found that BtTenA homologues were mostly represented in biofilms present on the surface of macroalgae found in Australian coral reefs. We also confirmed the ability of a recombinant BtTenA to degrade thiamin. Our study shows that BttenA-like genes which encode a novel sub-class of TenA proteins are sparingly distributed across two kingdoms of life, a feature of accessory genes known for their ability to spread between species through horizontal gene transfer.

Exchange of Vitamin B1 and Its Biosynthesis Intermediates Shapes the Composition of Synthetic Microbial Cocultures and Reveals Complexities of Nutrient Sharing.

Microbial communities occupy diverse niches in nature, and community members routinely exchange a variety of nutrients among themselves. While large-scale metagenomic and metabolomic studies shed some light on these exchanges, the contribution of individual species and the molecular details of specific interactions are difficult to track. In this study, we follow the exchange of vitamin B1 (thiamin) and its intermediates between microbes within synthetic cocultures of Escherichia coli and Vibrio anguillarum. Thiamin contains two moieties, 4-amino-5-hydroxymethyl-2-methylpyrimidine (HMP) and 4-methyl-5-(2-hydroxyethyl)thiazole (THZ), which are synthesized by distinct pathways using enzymes ThiC and ThiG, respectively, and then coupled by ThiE to form thiamin. Even though E. coli ΔthiC, ΔthiE, and ΔthiG mutants are thiamin auxotrophs, we observed that cocultures of ΔthiC-ΔthiE and ΔthiC-ΔthiG mutants are able to grow in a thiamin-deficient medium, whereas the ΔthiE-ΔthiG coculture does not. Further, the exchange of thiamin and its intermediates in V. anguillarum cocultures and in mixed cocultures of V. anguillarum and E. coli revealed that there exist specific patterns for thiamin metabolism and exchange among these microbes. Our findings show that HMP is shared more frequently than THZ, concurrent with previous observations that free HMP and HMP auxotrophy is commonly found in various environments. Furthermore, we observe that the availability of exogenous thiamin in the media affects whether these strains interact with each other or grow independently. These findings collectively underscore the importance of the exchange of essential metabolites as a defining factor in building and modulating synthetic or natural microbial communities. IMPORTANCE Vitamin B1 (thiamin) is an essential nutrient for cellular metabolism. Microorganisms that are unable to synthesize thiamin either fully or in part exogenously obtain it from their environment or via exchanges with other microbial members in their community. In this study, we created synthetic microbial cocultures that rely on sharing thiamin and its biosynthesis intermediates and observed that some of them are preferentially exchanged. We also observed that the coculture composition is dictated by the production and/or availability of thiamin and its intermediates. Our studies with synthetic cocultures provide the molecular basis for understanding thiamin sharing among microorganisms and lay out broad guidelines for setting up synthetic microbial cocultures by using the exchange of an essential metabolite as their foundation.

Tumor necrosis factor α impedes colonic thiamin pyrophosphate and free thiamin uptake: involvement of JNK/ERK1/2-mediated pathways.

The aim of this study was to examine the effect of TNFα (i.e., a predominant proinflammatory cytokine produced during chronic gut inflammation) on colonic uptake of thiamin pyrophosphate (TPP) and free thiamin, forms of vitamin B1 that are produced by the gut microbiota and are absorbed via distinct carrier-mediated systems. We utilized human-derived colonic epithelial CCD841 and NCM460 cells, human differentiated colonoid monolayers, and mouse intact colonic tissue preparations together with an array of cellular/molecular approaches in our investigation. The results showed that exposure of colonic epithelial cells to TNFα leads to a significant inhibition in TPP and free thiamin uptake. This inhibition was associated with: 1) a significant suppression in the level of expression of the colonic TPP transporter (cTPPT; encoded by SLC44A4), as well as thiamin transporters-1 & 2 (THTR-1 & -2; encoded by SLC19A2 & SLC19A3, respectively); 2) marked inhibition in activity of the SLC44A4, SLC19A2, and SLC19A3 promoters; and 3) significant suppression in level of expression of nuclear factors that are needed for activity of these promoters (i.e., CREB-1, Elf-3, NF-1A, SP-1). Furthermore, the inhibitory effects were found to be mediated via JNK and ERK1/2 signaling pathways. We also examined the level of expression of cTPPT and THTR-1 & -2 in colonic tissues of patients with active ulcerative colitis and found the levels to be significantly lower than in healthy controls. These findings demonstrate that exposure of colonocytes to TNFα suppresses TPP and free thiamin uptake at the transcriptional level via JNK- and Erk1/2-mediated pathways.

Alzheimer's disease is associated with disruption in thiamin transport physiology: A potential role for neuroinflammation.

Alzheimer's disease (AD) is a neurodegenerative disease characterized by Amyloid-ß peptide (Aß) containing plaques and cognitive deficits. The pathophysiology of AD also involves neuroinflammation. Vitamin B1 (thiamin) is indispensable for normal cellular energy metabolism. Thiamin homeostasis is altered in AD, and its deficiency is known to aggravate AD pathology. Little, however, is known about possible alterations in level of expression of thiamin transporters-1 and -2 (THTR-1 and -2) in the brain of AD, and whether pro-inflammatory cytokines affect thiamin uptake by brain cells. We addressed these issues using brain tissue samples [prefrontal cortex (PFC) and hippocampus (HIP)] from AD patients and from 5XFAD mouse model of AD, together with cultured human neuroblastoma SH-SY5Y cells as model. Our results revealed a significantly lower expression of both THTR-1 and THTR-2 in the PFC and HIP of AD patients and 5XFAD mouse model of AD compared to appropriate normal controls. Further, we found that exposure of the SH-SY5Y cells to pro-inflammatory cytokines (IL-1ß, IL-6, and TNF-α) led to a significant inhibition in thiamin uptake. Focusing on IL-1ß, we found the inhibition in thiamin uptake to be time-dependent and reversible; it was also associated with a substantial reduction in expression of THTR-1 (but not THTR-2) protein and mRNA as well as a decrease in promoter activity of the SLC19A2 gene (which encodes THTR-1). Finally, using transcriptomic analysis, we found that thiamin availability in SH-SY5Y cells caused changes in the expression of genes relevant to AD pathways. These studies demonstrate, for the first time, that thiamin transport physiology/molecular biology parameters are negatively impacted in AD brain and that pro-inflammatory cytokines inhibit thiamin uptake by neuroblastoma cells. The results also support a possible role for thiamin in the pathophysiology of AD.

Sustainable bio-based antimicrobials derived from fatty acids: Synthesis, safety, and efficacy.

Some conventional sanitizers and antibiotics used in food industry may be of concerns due to generation of toxic byproducts, impact on the environment, and the emergence of antibiotic resistance bacteria. Bio-based antimicrobials can be an alternative to conventional sanitizers since they are produced from renewable resources, and the bacterial resistance to these compounds is of less concern than those of currently used antibiotics. Among the bio-based antimicrobial compounds, those produced via either fermentation or chemical synthesis by covalently or electrovalently attaching specific moieties to the fatty acid have drawn attention in recent years. Disaccharide, arginine, vitamin B1, and phenolics are linked to fatty acids resulting in the production of sophorolipid, lauric arginate ethyl ester, thiamin dilauryl sulfate, and phenolic branched-chain fatty acid, respectively, all of which are reported to exhibit antimicrobial activity by targeting the cell membrane of the bacteria. Also, studies that applied these compounds as food preservatives by combining them with other compounds or treatments have been reviewed regarding extending the shelf life and inactivating foodborne pathogens of foods and food products. In addition, the phenolic branched-chain fatty acids, which are relatively new compounds compared to the others, are highlighted in this review.

Structure and function of aerotolerant, multiple-turnover THI4 thiazole synthases.

Plant and fungal THI4 thiazole synthases produce the thiamin thiazole moiety in aerobic conditions via a single-turnover suicide reaction that uses an active-site Cys residue as sulfur donor. Multiple-turnover (i.e. catalytic) THI4s lacking an active-site Cys (non-Cys THI4s) that use sulfide as sulfur donor have been biochemically characterized -- but only from archaeal methanogens that are anaerobic, O2-sensitive hyperthermophiles from sulfide-rich habitats. These THI4s prefer iron as cofactor. A survey of prokaryote genomes uncovered non-Cys THI4s in aerobic mesophiles from sulfide-poor habitats, suggesting that multiple-turnover THI4 operation is possible in aerobic, mild, low-sulfide conditions. This was confirmed by testing 23 representative non-Cys THI4s for complementation of an Escherichia coli ΔthiG thiazole auxotroph in aerobic conditions. Sixteen were clearly active, and more so when intracellular sulfide level was raised by supplying Cys, demonstrating catalytic function in the presence of O2 at mild temperatures and indicating use of sulfide or a sulfide metabolite as sulfur donor. Comparative genomic evidence linked non-Cys THI4s with proteins from families that bind, transport, or metabolize cobalt or other heavy metals. The crystal structure of the aerotolerant bacterial Thermovibrio ammonificans THI4 was determined to probe the molecular basis of aerotolerance. The structure suggested no large deviations compared with the structures of THI4s from O2-sensitive methanogens, but is consistent with an alternative catalytic metal. Together with complementation data, use of cobalt rather than iron was supported. We conclude that catalytic THI4s can indeed operate aerobically and that the metal cofactor inserted is a likely natural determinant of aerotolerance.

A case of dry beriberi from alcohol use disorder and disordered eating.

Background: Thiamin is an essential vitamin that is involved in every organ system in the body. Thiamin deficiency can present as beriberi or Wernicke's encephalopathy. We seek to educate practitioners in developed countries to include beriberi on the differential diagnosis when a patient with alcohol use disorder, poor diet and/or disordered eating presents with ascending paralysis without albuminocytologic dissociation. Case: In this case report, a 20-year-old female with no past medical history presented with three weeks of ascending paralysis. At presentation, she could not grasp objects, walk, or rise from a seated position. She reported consuming excessive alcohol and an otherwise limited diet due to picky eating. The patient was ultimately diagnosed with acute inflammatory demyelinating polyneuropathy secondary to dry beriberi from severe protein-calorie malnutrition and alcohol use disorder. She received an aggressive thiamin replacement regimen as well as physical and occupational therapy. She was discharged to home 24 days after her initial presentation. Discussion: This patient case offers a unique presentation of ascending paralysis without albuminocytologic dissociation due to severe dry beriberi from a diet of unenriched carbohydrates and excessive alcohol in an otherwise young, healthy adult in the United States. Our goal is that in reviewing the unusual details of this case, providers will be better equipped for the timely diagnosis and treatment of similar cases in the future.

Vitamin B1 THIAMIN REQUIRING1 synthase mediates the maintenance of chloroplast function by regulating sugar and fatty acid metabolism in rice.

Vitamin B1 (VB1), including thiamin, thiamin monophosphate (TMP), and thiamin pyrophosphate (TPP), is an essential micronutrient for all living organisms. Nevertheless, the precise function of VB1 in rice remains unclear. Here, we described a VB1 auxotrophic mutant, chlorotic lethal seedling (cles) from the mutation of OsTH1, which displayed collapsed chloroplast membrane system and decreased pigment content. OsTH1 encoded a phosphomethylpyrimidine kinase/thiamin-phosphate pyrophosphorylase, and was expressed in various tissues, especially in seedlings, leaves, and young panicles. The VB1 content in cles was markedly reduced, despite an increase in the expression of VB1 synthesis genes. The decreased TPP content affected the tricarboxylic acid cycle, pentose phosphate pathway, and de novo fatty acid synthesis, leading to a reduction in fatty acids (C16:0 and C18:0) and sugars (sucrose and glucose) of cles. Additionally, irregular expression of chloroplast membrane synthesis genes led to membrane collapse. We also found that alternative splicing and translation allowed OsTH1 to be localized to both chloroplast and cytosol. Our study revealed that OsTH1 was an essential enzyme in VB1 biosynthesis and played crucial roles in seedling growth and development by participating in fatty acid and sugar metabolism, providing new perspectives on VB1 function in rice.

Dietary thiamin requirement of fingerling major carp Catla catla (Hamilton).

Dietary thiamin requirement of fingerling Catla catla (3.5 ± 0.15 g) was evaluated by feeding casein-gelatin-based iso-nitrogenous (350 g/kg crude protein) and iso-caloric (16.72 kJ/g GE) diets containing six graded levels of thiamin (0, 0.2, 0.4, 0.8, 1.6 and 3.2 mg/kg dry diet) for 12 weeks. Significantly (p < 0.05) higher weight gain (AWG), best feed conversion ratio (FCR), protein retention efficiency (PRE), RNA/DNA ratio and haematological indices were recorded in fish fed diet containing 0.8 mg/kg thiamin. Dietary thiamin supplementation improved transketolase activity (TKA) and maximum value was recorded in fish fed 0.8 mg/kg thiamin beyond which stagnation in TKA activity was evident. Liver thiamin concentration was found to be maximum in fish fed diet containing 1.6 mg/kg thiamin. A significant (p < 0.05) consistent reduction in the hepatic thiobarbituric acid reactive substances (TBARS) activity was displayed with incremental concentration of thiamin up to 0.8 mg/kg, beyond which a reverse trend was evident. However, a significant (p < 0.05) improvement was noted in superoxide dismutase (SOD) and catalase (CAT) activity with the increasing level of dietary thiamin from 0 to 0.8 mg/kg. Broken-line regression analysis of AWG, FCR, PRE and TKA estimated the requirement in the range of 0.74-0.79 mg/kg dry diet.

Developmental maturation of the colonic uptake process of the microbiota-generated thiamin pyrophosphate.

The water-soluble vitamin B1 is essential for normal human health and physiology. In its main biologically active form, i.e., thiamin pyrophosphate (TPP), the vitamin plays many critical roles in cell metabolism; thus, its deficiency leads to a variety of adverse effects. Humans/mammals obtain vitamin B1 from two exogenous sources: diet and gut microbiota. Considerable amount of the microbiota-generated vitamin B1 exists in the form of TPP, and colonocytes can efficiently absorb this TPP via a high-affinity and specific carrier-mediated mechanism that involves the recently cloned colonic TPP transporter (cTPPT; product of SLC44A4 gene). There is nothing currently known about colonic uptake of TPP during early stages of life and whether the process undergoes developmental regulation. We addressed this issue using the mouse as animal model. Our results showed that colonic uptake of TPP undergoes developmental upregulation as the animal moves from the suckling period to weanling and adulthood. This upregulation in uptake was found to be associated with a parallel induction in level of expression of the cTPPT protein, mRNA, and heterogeneous nuclear RNA, suggesting possible involvement of transcriptional mechanism(s). We also found a parallel upregulation in the level of expression of the two nuclear factors that drive activity of the SLC44A4 promoter (i.e., CREB-1 and Elf-3) with maturation. These results demonstrate, for the first time, to our knowledge, that colonic TPP uptake process and cTPPT expression are developmentally upregulated and that this upregulation is likely driven via transcriptional mechanism(s).NEW & NOTEWORTHY The colonic carrier-mediated uptake process of the microbiota-generated and phosphorylated form of vitamin B1, i.e., thiamin pyrophosphate, undergoes ontogenic changes that parallel the development of the gut microbiota (and their ability to generate vitamins) during early stages of life.

Proinflammatory cytokines inhibit thiamin uptake by human and mouse pancreatic acinar cells: involvement of transcriptional mechanism(s).

Thiamin (vitamin B1) plays critical roles in normal metabolism and function of all mammalian cells. Pancreatic acinar cells (PACs) import thiamin from circulation via specific carrier-mediated uptake that involves thiamin transporter-1 and -2 (THTR-1 and -2; products of SLC19A2 and SLC19A3, respectively). Our aim in this study was to investigate the effect(s) of proinflammatory cytokines on thiamin uptake by PACs. We used human primary (h)PACs, PAC 266-6 cells, and mice in vivo as models in the investigations. First, we examined the level of expression of THTR-1 and -2 mRNA in pancreatic tissues of patients with chronic pancreatitis and observed severe reduction in their expression compared with normal control subjects. Exposing hPACs and PAC 266-6 to proinflammatory cytokines (hyper IL-6, TNF-α, and IL-1ß) was found to lead to a significant inhibition in thiamin uptake. Focusing on hyper-IL-6 (which also inhibited thiamin uptake by primary mouse PACs), the inhibition in thiamin uptake was found to be associated with significant reduction in THTR-1 and -2 proteins and mRNA expression as well as in activity of the SLC19A2 and SLC19A3 promoters; it was also associated with reduction in level of expression of the transcription factor Sp1 (which is required for activity of these promoters). Finally, blocking the intracellular Stat3 signaling pathway was found to lead to a significant reversal in the inhibitory effect of hyper IL-6 on thiamin uptake by PAC 266-6. These results show that exposure of PACs to proinflammatory cytokines negatively impacts thiamin uptake via (at least in part) transcriptional mechanism(s).NEW & NOTEWORTHY Findings of the current study demonstrate, for the first time, that exposure of pancreatic acinar cells to proinflammatory cytokines (including hyper IL-6) cause significant inhibition in vitamin B1 (thiamin; a micronutrient that is essential for normal cellular energy metabolism) and that this effect is mediated at the level of transcription of the thiamin transporter genes SLC19A2 and SLC19A3.

Bioinformatic and experimental evidence for suicidal and catalytic plant THI4s.

Like fungi and some prokaryotes, plants use a thiazole synthase (THI4) to make the thiazole precursor of thiamin. Fungal THI4s are suicide enzymes that destroy an essential active-site Cys residue to obtain the sulfur atom needed for thiazole formation. In contrast, certain prokaryotic THI4s have no active-site Cys, use sulfide as sulfur donor, and are truly catalytic. The presence of a conserved active-site Cys in plant THI4s and other indirect evidence implies that they are suicidal. To confirm this, we complemented the Arabidopsistz-1 mutant, which lacks THI4 activity, with a His-tagged Arabidopsis THI4 construct. LC-MS analysis of tryptic peptides of the THI4 extracted from leaves showed that the active-site Cys was predominantly in desulfurated form, consistent with THI4 having a suicide mechanism in planta. Unexpectedly, transcriptome data mining and deep proteome profiling showed that barley, wheat, and oat have both a widely expressed canonical THI4 with an active-site Cys, and a THI4-like paralog (non-Cys THI4) that has no active-site Cys and is the major type of THI4 in developing grains. Transcriptomic evidence also indicated that barley, wheat, and oat grains synthesize thiamin de novo, implying that their non-Cys THI4s synthesize thiazole. Structure modeling supported this inference, as did demonstration that non-Cys THI4s have significant capacity to complement thiazole auxotrophy in Escherichia coli. There is thus a prima facie case that non-Cys cereal THI4s, like their prokaryotic counterparts, are catalytic thiazole synthases. Bioenergetic calculations show that, relative to suicide THI4s, such enzymes could save substantial energy during the grain-filling period.