Project description:Messenger RNA (mRNA) translation can lead to higher rates of mRNA decay, suggesting a role for the ribosome in mRNA destruction. Furthermore, features of an mRNA, such as codon identities, that are directly probed by the ribosome also correlate with mRNA decay rates. Specifically, many amino acids are encoded by synonymous codons, and some synonymous codons are decoded by more abundant tRNAs leading to more optimal translation and increased mRNA stability. In addition to different translation rates, the presence of individual codons can lead to higher or lower rates of amino acid misincorporation which could potentially lead to protein misfolding if an individual amino acid makes many critical contacts in a structure. Here, we directly test whether amino acid misincorporation affects mRNA stability, taking advantage of an aminoglycoside antibiotic (G418) which promotes higher error rates in the ribosome. We observe that G418 decreases firefly luciferase mRNA stability in an in vitro system, and we similarly observe that G418 reduces mRNA stability in mouse embryonic stem cells (mESCs). G418-sensitive mRNAs are enriched for suboptimal hydrophobic amino acid codons as well as other codons that are known to result in higher rates of amino acid misincorporation. Since protein folding is highly sensitive to the identity of hydrophobic amino acids, these results strongly suggest that defects in protein folding are linked to mRNA decay.
Project description:Two-week old bread wheat seedlings hydroponically grown Hoagland solution were transferred to potassium (K+)-free conditions for 8 d, their root and leaf proteome profiles were assessed using iTRAQ proteome method, and NCBInr database combined with the recently published bread wheat genome information were used to analyze the identified protein species. Over 4,000 unique proteins were identified, 818 K+-responsive protein species showed significant abundance regulation. The most majority of the identified K+-responsive protein species had exact gene loci, and showed no global but tissue- and chromosome- dependent genome distributions. The identified protein species were associated with diverse functions and exhibited organ-specific differences. Most of identified protein species associated with hormone synthesis were the enzymes involved in the synthesis of jasmonic acid (JA). Allene oxide synthase (AOS), a key JA synthesis-related enzyme, was significantly induced in both root and leaf organs of K+-deficient wheat seedlings, and its overexpression in rice enhanced the tolerance to low K+ or K+ deficiency, increased contents of K+ and JA and transcription levels of some K+-responsive genes. However, rice AOS T-DNA inserted mutant (osaos) exhibited more sensitivity to K+ deficiency. K+ deficiency significantly increased abundance of a high affinity K+ transporter (TaAHAK1), TaAHAK1 transgenic rice seedlings markedly alleviated sensitivity to K+ deficiency, and K+ deficiency also upregulated expression of homologous OsHAK1 gene in TaAOS transgenic rice plants. These results suggested an essential role of JA in K+ deficiency and gave molecular insight into the responses of plant to K+ deficiency.
Project description:Fusarium head blight (FHB), caused by Fusarium graminearum is a devastating disease that affects global wheat production. F. graminearum encodes many effector proteins; however, its virulence mechanisms are poorly understood. In this study, we identified a secretory effector candidate protein (FgEC10) that is essential for the virulence of F. graminearum and suppresses the basal immune response. FgEC10 interacted strongly with wheat fumarylacetoacetate hydrolase (TaFAH) and accelerated its degradation via the 26S proteasome pathway. In addition, we showed that TaFAH interacted with TaPSMD, an important component of the 26S proteasome and that FgEC10 enhanced the interaction between TaFAH and TaPSMD. RNA silencing or overexpression of TaFAH in wheat plants showed that TaFAH positively regulated wheat FHB resistance, heading period, and grain yield. Transcriptomic analysis revealed that TaFAH promoted the expression of genes associated with disease resistance and the heading period. Metabolomic analysis revealed that overexpression of TaFAH increased the levels of several amino acids that inhibited F. graminearum growth and enhanced wheat resistance to FHB. Collectively, our study reveals a novel pathogenic mechanism and provides a valuable genetic resource for improving FHB resistance and grain yield in wheat.
Project description:This is report of GWAS and GS of amino acid contents in cultivated soybean seeds. The results are anticipated to facilitate marker assisted selection and GS of nutritional traits in soybean, gene identification and cloning.
Project description:Overexpression of asip1 in transgenic zebrafish disrupts dorsoventral pigment pattern in addition to increasing food intake levels and linear growth. A higher feed intake is unnecessary in transgenic fish to enable larger and heavier growth. A plausible explanation may rely on the enhanced feeding efficiency mediated by improved nutrient absorption in transgenic animals. To test this hypothesis, wide scope transcriptomic techniques were used to pipe the potential pathways involved in the enhanced nutrient absorption and intestinal epithelium permeability/integrity, in addition, the electrogenic capacity for amino acid transport was analysed. Transcriptomic results suggested that amino acid, monocarboxylates, ionic and vitamin transmembrane transport was substantially modified. Enrichment analysis also revealed an inhibition of intestinal lipid metabolism and down-regulation of KEGG pathways related to membrane integrity suggesting an augmented intestinal laxity that may result in enhanced paracellular transport. Electrophysiological experiments carried out in Ussing chambers show that asip1 overexpression decrease membrane tissue resistance (Rt), indicating a modification of the intestinal barrier function in ASIP1 transgenic animals. Similarly, paracellular permeability was higher in transgenic zebrafish. Both the decrease in Rt and the increase in permeability point to an ASIP1-dependent decrease in the tissue barrier function. Electrogenic amino acid transport was also enhanced in transgenic animals providing a strong indication that ASIP1 fish can extract more amino acids from their diet at similar feeding levels. Both transcriptomic and electrophysiological results suggest that asip1-overexpressing zebrafish display improved nutrient absorption and by extension a higher feed efficiency which explains enhanced growth in the absence of augmented food intake. The enhanced growth of ASIP1 zebrafish potentially mediated by improved nutrient uptake and feed efficiency suggests that the melanocortin system, specifically asip1 overexpression, is a potential target for the development of genetically engineered fish displaying improved performance and no differential lipid accumulation.
Project description:External application of acetic acid has been recently reported to enhance the survival to drought in plants such as Arabidopsis, rapeseed, maize, rice and wheat, but the effects of acetic acid application on increased drought tolerance in woody plants such as a tropical crop “cassava” remain elusive. A molecular understanding of acetic acid-induced drought avoidance in cassava will contribute to the development of technology that can be used to enhance drought tolerance without resorting to transgenic technology or advancements in cassava cultivation. In the present study, morphological, physiological and molecular responses to drought were analyzed in cassava after the treatment with acetic acid. Results indicated that the acetic acid-treated cassava plants had a higher level of drought avoidance than water-treated, control plants. Specifically, higher leaf relative water content, and chlorophyll and carotenoid levels were observed as soils dried out during the drought treatment. Leaf temperatures in acetic acid-treated cassava plants were higher relative to leaves on plants pretreated with water and the increase of ABA content was observed in leaves of acetic acid-treated plants, suggesting that stomatal conductance and the transpiration rate in leaves of acetic acid-treated plants decreased to maintain relative water contents and avoid drought. Transcriptome analysis revealed that the acetic acid treatment increased the expression of ABA signaling-related genes, such as OPEN STOMATA 1 (OST1) and protein phosphatase 2C; as well as drought response and tolerance-related genes, such as outer membrane tryptophan-rich sensory protein (TSPO), and heat shock proteins. Collectively, the external application of acetic acid enhances drought avoidance in cassava through the upregulation of ABA signaling pathway genes and several stress response- and tolerance-related genes. These data support the idea that adjustments of the acetic acid application to plants is useful to enhance drought tolerance in order to minimize the growth inhibition in the agricultural field.
Project description:Recent observations about how cells sense amino acids have argued for preeminent roles of mTOR and the stress kinase GCN2 in allowing cells to estimate their amino acid needs. Here we used models of programmed immune microenvironments where helper T cells have to sense how much amino acids are available to engage in antigen-fueled proliferation. Contrary to current models, T cells activate mTOR in the competency phase of the cell cycle regardless of amino acid amounts, GCN2 or surface TCR. Instead, we found T cells use an amino acid sensing system to target IL-2-induced STAT5 phosphorylation at the restriction point of cell cycle commitment. mTOR activity is subsequently reduced and specifically connected to SREBP activation. T cells can be pushed into cycle by increasing IL-2 even when no amino acids are available. Collectively, our studies reveal helper T cells use sequential and distinct pathways to measure local amino acid concentrations.