Project description:By physically linking amino acids to their codon assignments, transfer RNAs (tRNAs) are essential for protein synthesis and translation fidelity. Some natural human tRNA variants cause amino acid mis-incorporation at a codon or set of codons. Recent work showed a naturally occurring tRNASer variant decodes phenylalanine codons with serine and inhibits protein synthesis. We hypothesized that human tRNA variants that mis-read glycine (Gly) codons with alanine (Ala) will disrupt protein homeostasis. The A3G mutation occurs naturally in tRNAGly variants (tRNAGlyCCC, tRNAGlyGCC) and creates an alanyl-tRNA synthetase (AlaRS) identity element (G3:U70). Because AlaRS does not recognize the anticodon, the human tRNAAlaAGC G35C (tRNAAlaACC) variant may function similarly to mis-incorporate Ala at Gly codons. The tRNAGly and tRNAAla variants had no effect on protein synthesis in mammalian cells under normal growth conditions, however, tRNAGlyGCC A3G depressed protein synthesis in the context of proteasome inhibition. Mass spectrometry confirmed Ala mistranslation at several Gly codons caused by the tRNAGlyGCC A3G and tRNAAlaAGC G35C mutants, and in some cases, we observed multiple mistranslation events in the same peptide. The data reveal mistranslation of Ala at Gly codons and defects in protein homeostasis generated by natural human tRNA variants that are tolerated under normal conditions.
Project description:The occurrence and development of secondary injury after spinal cord injury (SCI) is a key role affecting SCI recovery, and oxidative stress plays an important role in secondary injury. α-lipoic acid (ALA) is recognized as a powerful antioxidant that mitigates secondary damage and exerts neuroprotective effects. However, the mechanism by which ALA provides antioxidant role remains unknown. The application of ALA to rats after SCI was used to explore the effect of ALA on local gene transcription and expression after SCI and its role in SCI recovery.
Project description:5-aminolevulinic acid (ALA) is one of the natural amino acids and a product of the first heme synthesis pathway in mitochondria. Recently, a supplement containing ALA is available in Japan and used for the purpose of the enhancement of ATP synthesis via mitochondrial activity. In humans, the immunomodulatory effect of ALA has attracted attention as a new function of ALA, and its application to cancer, inflammatory disease, and autoimmune diseases are being investigated. In the present study, to evaluate the effects of ALA on canine immunity, we performed in vitro studies using peripheral blood mononuclear cells (PBMC) from healthy dogs. Heme oxygenase-1 protein was expressed in Madin-Darby Canine Kidney cells and PBMCs treated with 5-ALA and ferrous sodium citrate (SFC), which showed that ALA works in dogs as well as humans. When PBMCs were stimulated with concanavalin A (ConA), the addition of ALA resulted in a significant increase in interferon-gamma (IFN-γ) produced by ConA-stimulated PBMCs. A comprehensive genetic alteration using next-generation RNA sequences (RNA-seq) was performed. From the result of RNA-seq, ALA enhanced the gene expression of T cell immunity including Th1, Th2, and Th17 subsets. In particular, the IL-17 signaling pathway was significantly upregulated. Then, we confirmed that ALA promoted the production of interleukin (IL)-17A in ConA-stimulated PBMCs. Together, these findings reveal that ALA promotes heme synthesis in mitochondria and enhances ConA-induced T cell immune responses in canine PBMCs.
Project description:Auxin amino acid conjugates are considered storage forms of auxins available as a source of active auxins on the plant demand. We treated Brassica rapa seedlings with 0.01 mM indole-3-acetyl-L-alanine (IAA-Ala), indole-3-propionyl-L-alanine (IPA-Ala), and indole- 3-butyryl-L-alanine (IBA-Ala) and examined their effects on the transcriptome. All auxin conjugate treatments caused similar patterns in transcription profiles compared to the control, but with different intensities of over- and under-expression depending on the treatment. Most auxin-related DEGs were identified after IBA-Ala treatment, followed by IPA-Ala and IAA-Ala, respectively.
Project description:Objective Improving mitochondrial function is a promising strategy for intervention in type 2 diabetes mellitus. This study investigated the preventive effects of sodium ferrous citrate (SFC) and 5-aminolevulinic acid phosphate (ALA) on several metabolic dysfunctions associated with obesity because they have been shown to alleviate abnormal glucose metabolism in humans. Methods Six-week-old male C57BL/6J mice were fed with a normal diet, a high-fat diet, or a high-fat diet supplemented with SFC and ALA for 15 weeks. Results The simultaneous supplementation of SFC + ALA to high-fat diet-fed mice prevented loss of muscle mass, improved muscle strength, and reduced obesity and insulin resistance. SFC + ALA prevented abnormalities in mitochondrial morphology and reverted the diet effect on the skeletal muscle transcriptome, including the expression of glucose uptake and mitochondrial oxidative phosphorylation-related genes. In addition, SFC + ALA prevented the decline in mitochondrial DNA copy number by enhancing mitochondrial DNA maintenance and antioxidant transcription activity, both of which are impaired in high-fat diet-fed mice during long-term fasting. Conclusions These findings suggest that SFC + ALA supplementation exerts its preventive effects in type 2 diabetes mellitus via improved skeletal muscle and mitochondrial health, further validating its application as a promising strategy for the prevention of obesity-induced metabolic disorders.
Project description:B16-BL6 mouse melanoma that had been maintained in C57BL/6J mice were used to evaluate the 5-aminolevulinic acid (ALA) and radiation dose effects on radiotherapy. Mice were divided into 6 groups after implantation of B16-BL6 cells; (1) no treatment (NT) ; (2) 5-ALA treatment (ALAT); (3) 10 session of fractionated irradiation (2Gy/day) (20XT); (4) 10 sessions of 5-ALA treatment followed by fractionated irradiation (2Gy/day) (ALA-20XT); (5) 10 session of fractionated irradiation (3Gy/day) (30XT); (6) 10 sessions of 5-ALA treatment followed by fractionated irradiation (3Gy/day) (ALA-30XT).
Project description:B16-BL6 mouse melanoma that had been maintained in C57BL/6J mice were used to evaluate the 5-aminolevulinic acid (ALA) effects on radiotherapy. Mice were divided into 4 groups after implantation of B16-BL6 cells; (1) control group; (2) ALA treatment; (3) Xray treatment; (4) ALA and Xray treatment.
Project description:5-aminolevulinic acid (ALA) is the common precursor of all biological synthezised tetrapyrroles. Inhibition of ALA synthesis results in decreased amounts of chlorophylls, heme, siroheme and phytochrome. It was previously shown that 4 out of 5 Arabidopsis mutants uncoupling nuclear gene expression from the physiological state of the chloroplast are affected in plant tetrapyrrole biosynthesis. It is common to all four mutants to show a reduced ALA formation. We investigate the impact of reduced plastidic ALA synthesis on nuclear gene expression by inhibition of ALA formation using gabaculin and compare these effects with the gun4-1 mutant.