Folate Compounds Research Articles

Progressive Lower Extremity Paralysis Caused by Intrathecal MTX-Induced Myelopathy Mimicking Guillain-Barre Syndrome: A Case Report.

Methotrexate (MTX) is commonly used in intrathecal chemotherapy for patients with acute lymphocytic leukemia (ALL) to prevent central nervous system (CNS) involvement. However, the use of MTX-based chemotherapy can lead to rare yet severe complications, such as MTX-induced myelopathy. Here, we report the case of MTX-induced myelopathy initially misdiagnosed as Guillain-Barre syndrome, leading to a delay in diagnosis and treatment. We present a case of a 39-year-old male with a history of B-cell acute lymphoblastic leukemia (B-ALL) who experienced bilateral foot paresthesia and progressive lower extremity weakness after intrathecal methotrexate (MTX) treatment. Initially, the patient was suspected as having Guillain-Barre syndrome (GBS) due to similar clinical features and nerve conduction studies. The patient received intravenous immunoglobulin (IVIG) treatment, but his condition worsened. T2-weighted images of the thoracic spinal cord revealed high signal intensity in both lateral and posterior columns, typically associated with subacute combined degeneration. However, elevated vitamin B12 levels ruled out SCD in this case. Based on the aforementioned findings, intrathecal methotrexate-induced myelopathy was diagnosed. This case highlights the diagnostic challenge posed by the similarity in clinical presentation between MTX-induced myelopathy and GBS. Differentiating between these conditions is critical for appropriate management. Prompt recognition and treatment with folate metabolism compounds may mitigate neurological sequelae.

Folate and Its Significance in Depressive Disorders and Suicidality: A Comprehensive Narrative Review.

Depressive disorders pose significant challenges to global public health, necessitating effective prevention and management strategies. Notably, the occurrence of suicide frequently coincides with depressive episodes. Suicide is as a paramount global health concern that demands efficacious preventive strategies. Current psychiatric approaches heavily rely on pharmacological interventions but have had limited success in addressing the global burden of mental health issues. Suboptimal nutrition, with its impact on the neuroendocrine system, has been implicated in the underlying pathology of depressive disorders. Folate, a group of water-soluble compounds, plays a crucial role in various central nervous system functions. Depressed individuals often exhibit low levels of serum and red blood cell folate. Multiple studies and systematic reviews have investigated the efficacy of folic acid and its derivative, L-methylfolate, which can cross the blood-brain barrier, as stand-alone or adjunct therapies for depression. Although findings have been mixed, the available evidence generally supports the use of these compounds in depressed individuals. Recent studies have established links between the one-carbon cycle, folate-homocysteine balance, immune system function, glutamate excitation via NMDA (N-methyl-D-aspartate) receptors, and gut microbiome eubiosis in mood regulation. These findings provide insights into the complex neurobiological mechanisms underlying the effects of folate and related compounds in depression. Through a comprehensive review of the existing literature, this study aims to advance our understanding of the therapeutic potential of folic acid and related compounds in depression treatment. It also seeks to explore their role in addressing suicidal tendencies and shed light on the neurobiological mechanisms involved, leveraging the latest discoveries in depression research.

Determination of Vitamin B12 and Folate Compounds in Commercially Available Edible Seaweed Products.

Information on the contents of both vitamin B12 and folate in edible seaweeds is limited, of which deficiencies disrupt methionine biosynthesis to accumulate homocysteine as a risk factor of cardiovascular diseases. Both vitamins were determined in commercially available edible seaweed products using high-performance liquid chromatography. Dried purple laver (Neopyropia yezoensis) products contain higher levels of vitamin B12 (approximately 30-60 μg/100 g dry weight) and folate compounds (approximately 880-1300 μg/100 g dry weight) than other seaweed products, such as kombu (Saccharina japonica), hijiki (Sargassum fusiformis), and wakame (Undaria pinnatifida). 5-methyltetrahydrofolate was the major folate compound in purple laver products. 5-formyltetrahydrofolate was found at a moderate level, whereas tetrahydrofolate, 5,10-metenyltetrahydrofolate, 10-formyltetrahydrofolate, and folic acid were found to be minor folate compounds. These findings suggest that dried purple laver (nori) products are suitable sources of vitamin B12 and folate compounds for humans, especially vegetarians.

Folate shapes plant root architecture by affecting auxin distribution.

Folate (vitamin B9) is important for plant root development, but the mechanism is largely unknown. Here we characterized a root defective mutant, folb2, in Arabidopsis, which has severe developmental defects in the primary root. The root apical meristem of the folb2 mutant is impaired, and adventitious roots are frequently found at the root-hypocotyl junction. Positional cloning revealed that a 61-bp deletion is present in the predicted junction region of the promoter and the 5' untranslated region of AtFolB2, a gene encoding a dihydroneopterin aldolase that functions in folate biosynthesis. This mutation leads to a significant reduction in the transcript level of AtFolB2. Liquid chromatography-mass spectrometry analysis showed that the contents of the selected folate compounds were decreased in folb2. Arabidopsis AtFolB2 knockdown lines phenocopy the folb2 mutant. On the other hand, the application of exogenous 5-formyltetrahydrofolic acid could rescue the root phenotype of folb2, indicating that the root phenotype is indeed related to the folate level. Further analysis revealed that folate could promote rootward auxin transport through auxin transporters and that folate may affect particular auxin/indole-3-acetic acid proteins and auxin response factors. Our findings provide new insights into the important role of folic acid in shaping root structure.

Host-dependent resistance of Group A Streptococcus to sulfamethoxazole mediated by a horizontally-acquired reduced folate transporter

Described antimicrobial resistance mechanisms enable bacteria to avoid the direct effects of antibiotics and can be monitored by in vitro susceptibility testing and genetic methods. Here we describe a mechanism of sulfamethoxazole resistance that requires a host metabolite for activity. Using a combination of in vitro evolution and metabolic rescue experiments, we identify an energy-coupling factor (ECF) transporter S component gene (thfT) that enables Group A Streptococcus to acquire extracellular reduced folate compounds. ThfT likely expands the substrate specificity of an endogenous ECF transporter to acquire reduced folate compounds directly from the host, thereby bypassing the inhibition of folate biosynthesis by sulfamethoxazole. As such, ThfT is a functional equivalent of eukaryotic folate uptake pathways that confers very high levels of resistance to sulfamethoxazole, yet remains undetectable when Group A Streptococcus is grown in the absence of reduced folates. Our study highlights the need to understand how antibiotic susceptibility of pathogens might function during infections to identify additional mechanisms of resistance and reduce ineffective antibiotic use and treatment failures, which in turn further contribute to the spread of antimicrobial resistance genes amongst bacterial pathogens.

Evaluation of Silver Nanoparticles Caffeic folate (NSFC) Complex Compound as New Potential Therapeutic Agent against Cancer Xenograft in Mice: Thesis Abstract

Thesis Abstract In the last few decades the properties of silver compounds have been of interest as potential cancer treatments. The present work was designed to study the effect of new conjugated caffeic and folic acid with silver nanoparticle with definite molecular size applied with and without gamma radiation exposure, as an antitumor agent against experimentally induced Ehrlich tumor and attempted to identify their potential molecular mechanisms of action throughout determination of anti-tumor activities using MTT cytotoxic assay against two human carcinoma cell lines in vitro, Apoptosis analysis by flow cytometry through caspase-8, caspase-3 and TNF determine in vivo . Adult female albino mice were used and divided into five groups, Animals were scarified and the following parameters were estimated, glutathione, glutathione peroxidase, superoxide dismutase in blood and caspase8, caspase 3 and TNF from scarified tumor tissue. The tumor specimens were processed for histopathological examination. Nano-silver folate caffeic (NSFC) complex compound treatment resulted in a growth inhibition in Hep-G2 and MCF-7 cells (IC50 7.70 µM and 14.50 µM, respectively). Flow cytometric analysis revealed that (NSFC) with radiation exposed has apoptotic effect at caspases 8, 3 and TNF more than any compound of them alone. That disturbance was found to be associated with a kinetic induction of apoptosis and showed modulation of antioxidant system beside its high percentage of necrotic cells by histopathological studies. the novel synthetic Nano-silver folate Caffeic complex compound may potentially present a new hope for the development of breast cancer therapeutics, which should attract further scientific and pharmaceutical interest.

Improvement and Characterization of Naturally Occurring Folate Compounds in Fruiting Bodies of Culinary-Medicinal Winter Mushroom, Flammulina velutipes (Agaricomycetes).

Edible mushrooms are among food sources containing natural folate compounds. However, little is known about how the content of folates in edible mushrooms may be improved. This study aimed to enrich Flammulina velutipes with higher levels of folates and to characterize patterns of the bioconversion of folates in the fruiting bodies. A convenient method was developed to treat a lignocellulosic substrate with synthetic folic acid. Folate compounds in the fruiting bodies grown in folic acid-treated substrates were evaluated relative to those in untreated substrates. Reverse phase high-performance liquid chromatography revealed different patterns of changes in levels of unsubstituted and substituted folates. While there was an approximately 15-fold and 8-fold increase in 5-formyl tetrahydrofolate and 5-methyl tetrahydrofolate, respectively, the most inconsistency was seen in the tetrahydrofolate content. There were also differences in the level of folate derivatives between commercial and native F. velutipes mushrooms. Since F. velutipes mushrooms can be consumed raw, the enriched mushrooms can be used as a dietary source to meet adult requirements for the daily uptake of natural folates.

Serine, glycine and one‑carbon metabolism in cancer (Review).

Serine/glycine biosynthesis and one-carbon metabolism are crucial in sustaining cancer cell survival and rapid proliferation, and of high clinical relevance. Excessive activation of serine/glycine biosynthesis drives tumorigenesis and provides a single carbon unit for one-carbon metabolism. One-carbon metabolism, which is a complex cyclic metabolic network based on the chemical reaction of folate compounds, provides the necessary proteins, nucleic acids, lipids and other biological macromolecules to support tumor growth. Moreover, one-carbon metabolism also maintains the redox homeostasis of the tumor microenvironment and provides substrates for the methylation reaction. The present study reviews the role of key enzymes with tumor-promoting functions and important intermediates that are physiologically relevant to tumorigenesis in serine/glycine/one-carbon metabolism pathways. The related regulatory mechanisms of action of the key enzymes and important intermediates in tumors are also discussed. It is hoped that investigations into these pathways will provide new translational opportunities for human cancer drug development, dietary interventions, and biomarker identification.

Artificial light impacts the physical and nutritional quality of lettuce plants

Leafy vegetables, including lettuce (Lactuca sativa L.), are considered to be healthy due to their high content of fiber, folate, carotenoids, phenolic and antioxidant compounds, minerals, and vitamins A, C, and K. Recently, LEDs are being used extensively as a supplementary light source in indoor agriculture due to the economical and physiological advantages that this artificial illumination offers compared to traditional fluorescence illumination. In this work, two commercially important lettuce varieties, Batavia Lettony (green leaves) and Batavia Diablotin (red leaves), were used to study the impact of LEDs (white and red–blue lights) and fluorescent illumination on their quality and health properties. Changes in the photosynthetic photon flux density from 250 to 400 µmol m−2 s−1 of fluorescent light increased growth parameters (leaf number, fresh and dry weight, and percentage of dry matter) of B. Lettony plants. We observed a positive impact of red–blue LED illumination on growth parameters analyzed in B. Diablotin plants compared to plants grown under fluorescent light at 250 µmol m−2 s−1. Leaf texture significantly increased in B. Lettony plants grown under 400 µmol m−2 s−1 fluorescent and LED illumination compared to that of plants grown under 250 µmol m−2 s−1 fluorescent light. This variable was only increased under red–blue LED illumination in B. Diablotin plants. Accumulation of bioactive compounds, such as anthocyanins and vitamin C, was higher in B. Diablotin plants grown under 250 µmol m−2 s−1 fluorescent light. Nutrient content in the foliar part was not modified under the light conditions used, except the Ca2+ content of B. Lettony plants grown under PPFD 400 µmol m−2 s−1 fluorescent light. Catalase (CAT) and peroxidase (POX) activities were differentially modified by light conditions in B. Lettony plants. However, POX activity was only modified in response to light conditions in B. Diablotin plants. Thus, this study demonstrates that LEDs could be used as an alternative to produce food under sustainable conditions. In this sense, although several horticultural studies have been conducted to establish the effectiveness of LEDs in lettuce growth, additional investigations are necessary to determine the optimal conditions for the use of LEDs to promote lettuce production and the accumulation of beneficial components, such as vitamins, minerals, fiber, and antioxidant compounds.

Growth Response of Lactobacillus rhamnosus Chloramphenicol-Resistant Strain ATCC 27773 to Pteroyl-Mono- and -Di-Glutamates.

This study investigated the effect of various concentrations of pteroyl-mono-γ-glutamate (PteGlu1) and pteroyl-di-γ-glutamate (PteGlu2) on the growth of Lactobacillus rhamnosus strains ATCC 7469 (wild-type strain) and ATCC 27773 (chloramphenicol-resistant strain) used for folate microbiological assays. At concentrations of 0.025-0.20 nmol/L, the growth of the chloramphenicol-resistant strain was stimulated to a greater extent by PteGlu1 than by PteGlu2, but the wild-type strain did not show such phenomena. L. rhamnosus ATCC 27773 bioassays were used to determine the total folate content of various foods treated with a chicken pancreas folate conjugase. This showed a significantly lower value when PteGlu1 was used as a calibrator than with PteGlu2. These results indicated that PteGlu2 should be the standard folate compound when chicken pancreas folate conjugase is used in preparing samples for L. rhamnosus ATCC 27773 bioassay.

An Immunohistopathologic Study to Profile the Folate Receptor Beta Macrophage and Vascular Immune Microenvironment in Giant Cell Arteritis.

Giant cell arteritis (GCA) is a chronic immune-mediated disease of medium-to-large sized arteries that affects older adults. GCA manifests with arthritis and occlusive symptoms of headaches, stroke or vision loss. Macrophages and T-helper lymphocytes infiltrate the vascular wall and produce a pro-inflammatory response that lead to vessel damage and ischemia. To date, there is no GCA biomarker that can monitor disease activity and guide therapeutic response. Folate receptor beta (FRB) is a glycosylphosphatidylinositol protein that is anchored on cell membranes and normally expressed in the myelomonocytic lineage and in the majority of myeloid leukemia cells as well as in tumor and rheumatoid synovial macrophages, where its expression correlates with disease severity. The ability of FRB to bind folate compounds, folic acid-conjugates and antifolate drugs has made it a druggable target in cancer and inflammatory disease research. This report describes the histopathologic and immunohistochemical methods used to assess expression and distribution of FRB in relation to GCAimmunopathology. Formalin-fixed and paraffin-embedded temporal artery biopsies from GCA and normal controls were stained with Hematoxylin and Eosin to review tissue histology and identify pathognomonic features.Immunohistochemistry was used to detect FRB, CD68 and CD3 expression. A microscopic analysis was performed to quantify the number of positively stained cells on 10 selected high-power-field sections and their respective locations in the arterial wall. Lymphohistiocytic (LH) inflammation accompanied by intimal hyperplasia and disrupted elastic lamina was seen in GCA with none found in controls. The LH infiltrate was composed of approximately 60% lymphocytes and 40% macrophages. FRB expression was restricted to macrophages, comprising 31% of the total CD68+ macrophage population and localized to the media and adventitia. No FRB was seen in controls. This protocol demonstrated a distinct numerical and spatial pattern of the FRB macrophage relative to the vascular immune microenvironment in GCA.

Changes in folate characteristics and its identification in broccoli (Brassica oleracea Italica) extract fermented by Lactic Acid Bacteria Mixed Culture (LAB)

Broccoli (Brassica oleracea Italica) was fermented by cultures of lactic acid bacteria (LAB) as a potential source of natural folic acid. This study aimed to evalte characteristic changes and to identify folate compounds from broccoli extract, fermented by mixed LAB cultures (L. bulgaricus, S. thermophulus, L.acidophilus, Bd. bifidum). The formulation of broccoli extract was fermented with variation of LAB starter culture with concentrations of 10 and 20%(v/v), and the change of characteristic of folic acid compound during fermentation (0 to 48 hours) with an interval of 8 hours was evaluated. The results showed that the fermentation of broccoli extract with different concentration of LAB culture had an effect on the concentration of folic acid produced, as well as the change of concentration of folic acid during the fermentation time interval. The optimum condition was obtained based on the highest folic acid concentration of 6.74%, at culture concentration of 20% during 24 hour fermentation with the value of folic acid concentration of 72.11 μg/mL, pH value of 4.29, total sugars of 34.61%, total acids of 0, 97%, dissolved protein of 14.64 mg/mL and total LAB of log 13.02 + 0.05 cfu / ml.

Nature's hydrides: rapid reduction of halocarbons by folate model compounds.

Halocarbons R-X are reduced to hydrocarbons R-H by folate model compounds under biomimetic conditions. The reactions correspond to a halide-hydride exchange with the methylenetetrahydrofolate (MTHF) models acting as hydride donors. The MTHF models are also functional equivalents of dehalohydrogenases but, unlike these enzymes, do not require a metal cofactor. The reactions suggest that halocarbons have the potential to act as endocrinological disruptors of biochemical pathways involving MTHF. As a case in point, we observe the rapid reaction of the MTHF models with the inhalation anaesthetic halothane. The ready synthetic accessibility of the MTHF models as well as their dehalogenation activity in the presence of air and moisture allow for the remediation of toxic, halogenated hydrocarbons.

Folates as adjuvants to anticancer agents: Chemical rationale and mechanism of action.

Folates have been used with cytotoxic agents for decades and today they are used in hundreds of thousands of patients annually. Folate metabolism is complex. In the treatment of cancer with 5-fluorouracil, the administration of folates mechanistically leads to the formation of [6R]-5,10-methylene-tetrahydrofolate, and the increased concentration of this molecule leads to stabilization of the ternary complex comprising thymidylate synthase, 2'-deoxy-uridine-5'-monophosphate, and [6R]-5,10-methylene-tetrahydrofolate. The latter is the only natural folate that can bind directly in the ternary complex, with other folates requiring metabolic activation. Modulation of thymidylate synthase activity became central in the study of folate/cytotoxic combinations and, despite wide use, research into the folate component was neglected, leaving important questions unanswered. This article revisits the mechanisms of action of folates and evaluates commercially available folate derivatives in the light of current research. Better genomic insight and availability of new analytical techniques and stable folate compounds may open new avenues of research and therapy, ultimately bringing increased clinical benefit to patients.

Bacterial Folates Provide an Exogenous Signal for C. elegans Germline Stem Cell Proliferation

Here we describe an invitro primary culture system for Caenorhabditis elegans germline stem cells. This culture system was used to identify a bacterial folate as a positive regulator of germ cell proliferation. Folates are a family of B-complex vitamins that function in one-carbon metabolism to allow the de novo synthesis of amino acids and nucleosides. We show that germ cell proliferation is stimulated by the folate 10-formyl-tetrahydrofolate-Glun both invitro and in animals. Other folates that can act as vitamins to rescue folate deficiency lack this germ cell stimulatory activity. The bacterial folate precursor dihydropteroate also promotes germ cell proliferation invitro and invivo, despite its inability to promote one-carbon metabolism. The folate receptor homolog FOLR-1 is required for the stimulation of germ cells by 10-formyl-tetrahydrofolate-Glun and dihydropteroate. This work defines a folate and folate-related compound as exogenous signals to modulate germ cell proliferation.

Effect of Biscuit Baking Conditions on the Stability of Microencapsulated 5-Methyltetrahydrofolic Acid and Their Physical Properties

Among the folate compounds, 5-methyltetrahydrofolic acid (5-CH3THF) is regarded as one of the most bioactive forms of folate. It is regarded as the better source of folate to humans as compared to folic acid, a synthetic form of folate, which is used for fortifying foods to prevent the incidence of neural tube defects in the new born babies. The use of 5-CH3THF as an alternative fortificant, in place of folic acid, has been explored by various researchers. However, fortification of 5-CH3THF is problematic due to its lower stability. This study investigated the stability of microencapsulated 5-CH3THF in biscuits baked at various temperatures and times as well as changes in their physical properties. Microcapsule with pectin and alginate ratio of 80:20, prepared by spray drying, gave the highest retention (68.6%) of the 5-CH3THF, therefore, chosen for fortification. The encapsulated and unencapsulated 5-CH3THF were mixed separately with flour and biscuit ingredients and baked at 180℃, 200 and 220℃, each for 5, 9 and 12 min. The inclusion of encapsulated and unencapsulated 5-CH3THF in the biscuit formulation and subsequent baking at various temperatures and times resulted in retention of 5-CH3THF from 19.1% to 1.7%. Microencapsulation of 5-CH3THF slightly improved the retention of 5-CH3THF over unencapsuated biscuits at 180℃ for 5 min, but almost no such effect was achieved under baking temperatures of 200℃ and 220℃. Physical analysis showed darker colour, harder texture and lower moisture content for biscuits baked at higher test temperatures. It seems intense heating condition that caused “over baking” of the biscuit likely to be responsible for the loss of the vitamin as well as less desirable physical properties of the biscuits.

Clinical signs and treatment of cerebral folate transport deficiency, a novel inherited disorder of folate metabolism

Background: Folates are essential cofactors for important metabolic pathways such as synthesis of amino acids, DNA and lipophilic substances. The main folate compound 5-methyltetrahydofolate (5MTHF) is actively transported across the blood- cerebrospinal fluid (CSF)-barrier to supply the brain. Cellular uptake of folates is mediated by the proton-coupled intestinal transporter, the reduced folate carrier, and by two GPI-anchored receptors, folate receptor α (FRa) and β (FRa). At least five distinct inherited disorders of folate transport and metabolism are presently. An inherited disorder of folate transport into brain has not been described yet.

Characterization of folate-dependent enzymes and indices of folate status in laying hens supplemented with folic acid or 5-methyltetrahydrofolate

The conversion of folic acid (FA) to the biologically active 5-methyltetrahydrofolate (5-MTHF) is necessary for the deposition of folate in the egg. A study was conducted to compare egg folate concentrations, indices of folate status, and activities of folate-dependent enzymes in response to equimolar intake of either FA or 5-MTHF in laying hens. Forty-eight laying hens, 24 wk of age, from 2 different strains (Shaver White and Shaver Brown) were randomly assigned to receive 1 of 3 (n = 8 per strain) dietary treatments: 1) basal diet with no supplemental folate, 2) basal diet + 10 mg/kg of FA, or 3) basal diet + 11.3 mg/kg of 5-MTHF for 3 wk. A completely randomized design with 3 dietary treatments and 2 laying hen strains in a 3 x 2 factorial arrangement was used. Data were subjected to ANOVA, using the PROC GLM procedure of SAS. Plasma homocysteine, serum, and egg folate concentrations; hepatic serine hydroxymethyltransferase; and methionine synthase activity were affected by dietary folate supplementation but not by its form (FA and 5-MTHF). Relative to control hens, plasma homocysteine was decreased (P < 0.05) by 14.2%, whereas serum and egg folate were increased (P < 0.05) by 78.3 and 61.8%, respectively, in hens consuming either folate compound. Hepatic serine hydroxymethyltransferase and methionine synthase activity were increased and decreased (P < 0.05), respectively, in folate-fed birds compared with control-fed birds. Hepatic dihydrofolate reductase was influenced by both the addition and form of dietary folate, being higher (P < 0.05) in FA-fed birds than in 5-MTHF and control-fed birds. Feed efficiency was improved (P < 0.05) in 5-MTHF-fed birds relative to FA-fed birds. Strain of hen influenced serum folate and plasma homocysteine concentrations but not other indices of folate metabolism. Overall, FA and 5-MTHF have equivalent effects in enhancing egg folate concentrations and improving folate status in laying hens. Also, supplementation and form of folate may modulate the activity of folate-dependent enzymes.

Folate Content in Tomato (Lycopersicon esculentum). Influence of Cultivar, Ripeness, Year of Harvest, and Pasteurization and Storage Temperatures

The effects of cultivar, on-vine ripening, and year of harvest on the folate content of raw tomatoes were studied. Folate content in hot-break tomato puree (HTP) subjected to pasteurization at different temperatures and its evolution during the shelf life of tomato juice were also investigated. 5-Methyltetrahydrofolate (5-CH(3)-H(4)-folate) was the only folate compound identified in raw tomatoes and HTP, but tetrahydrofolate (H(4)-folate) was 10% of the folate detected in tomato juice. The content of folates in raw tomatoes ranged from 4.1 to 35.3 microg/100 g of fresh weight and was highly influenced by all of the factors studied. No clear trend of folate content with ripening stage was observed. The extractability of 5-CH(3)-H(4)-folate from HTP increased significantly after pasteurization at 98 degrees C for 40 s, but higher temperatures decreased its content. Tomato juice showed folate losses during storage independent of the storage temperature. Folate losses were higher when tomato juice was packed in glass bottles than in Tetra Pak.

The Role of the Mitochondrial Glycine Cleavage Complex in the Metabolism and Virulence of the Protozoan Parasite Leishmania major

For the human pathogen Leishmania major, a key metabolic function is the synthesis of thymidylate, which requires 5,10-methylenetetrahydrofolate (5,10-CH(2)-THF). 5,10-CH(2)-THF can be synthesized from glycine by the mitochondrial glycine cleavage complex (GCC). Bioinformatic analysis revealed the four subunits of the GCC in the L. major genome, and the role of the GCC in parasite metabolism and virulence was assessed through studies of the P subunit (glycine decarboxylase (GCVP)). First, a tagged GCVP protein was expressed and localized to the parasite mitochondrion. Second, a gcvP(-) mutant was generated and shown to lack significant GCC activity using an indirect in vivo assay after incorporation of label from [2-(14)C]glycine into DNA. The gcvP(-) mutant grew poorly in the presence of excess glycine or minimal serine; these studies also established that L. major promastigotes require serine for optimal growth. Although gcvP(-) promastigotes and amastigotes showed normal virulence in macrophage infections in vitro, both forms of the parasite showed substantially delayed replication and lesion pathology in infections of both genetically susceptible or resistant mice. These data suggest that, as the physiology of the infection site changes during the course of infection, so do the metabolic constraints on parasite replication. This conclusion has great significance to the interpretation of metabolic requirements for virulence. Last, these studies call attention in trypanosomatid protozoa to the key metabolic intermediate 5,10-CH(2)-THF, situated at the junction of serine, glycine, and thymidylate metabolism. Notably, genome-based predictions suggest the related parasite Trypanosoma brucei is totally dependent on the GCC for 5,10-CH(2)-THF synthesis.