Identification and characterization of a plastidial ω-3 fatty acid desaturase <i>EgFAD8</i> from oil palm (<i>Elaeis guineensis</i> Jacq.) and its promoter response to light and low temperature

<div><p>In higher plants, ω-3 fatty acid desaturases are the key enzymes in the biosynthesis of alpha-linolenic acid (18:3), which plays key roles in plant metabolism as a structural component of both storage and membrane lipids. Here, the first ω-3 fatty acid desaturase gene was identified and characterized from oil palm. The bioinformatic analysis indicated it encodes a temperature-sensitive chloroplast ω-3 fatty acid desaturase, designated as <i>EgFAD8</i>. The expression analysis revealed that <i>EgFAD8</i> is highly expressed in the oil palm leaves, when compared with the expression in the mesocarp. The heterologous expression of <i>EgFAD8</i> in yeast resulted in the production of a novel fatty acid 18:3 (about 0.27%), when fed with 18:2 in the induction culture. Furthermore, to detect whether <i>EgFAD8</i> could be induced by the environment stress, we detected the expression efficiency of the <i>EgFAD8</i> promoter in transgenic Arabidopsis treated with low temperature and darkness, respectively. The results indicated that the promoter of <i>EgFAD8</i> gene could be significantly induced by low temperature and slightly induced by darkness. These results reveal the function of EgFAD8 and the feature of its promoter from oil palm fruits, which will be useful for understanding the fuction and regulation of plastidial ω-3 fatty acid desaturases in higher plants.</p></div

Expression analysis of <i>EgFAD8</i> in oil palm leaves and mesocarp at five different developmental stages.

<p>P1: mesocarp at 30–60 DAP; P2: mesocarp at 60–100 DAP; P3: mesocarp at 100–120 DAP; P4: mesocarp at 120–140 DAP; P5: mesocarp at 140–160 DAP. DAP: days after pollination.</p

Phylogenetic analysis of EgFAD8 and fatty acid desaturases from <i>Arabidopsis thaliana</i>.

<p>A neighbor-joining tree was generated by MEGA5 with bootstrap analysis based on 1000 replications. Bootstrap values are shown at branch points. Scale bar indicates the number of substitutions per site.</p

Total fatty acid composition of transformed yeast with <i>EgFAD8</i> or pYES2.

<p>All experiments were carried out in triplicate. Asterisks indicate statistically significant differences compared with the control (Student’s <i>t</i> test: **, p < 0.01). 16:0, palmitic acid; 16:1, palmitoleic acid; 18:0, stearic acid; 18:1, oleic acid; 18:2, linoleic acid; 18:3,alpha linoleic acid.</p

Polymorphism of DNA Methyltransferase 3b and Association with Development and Prognosis in Gastric Cancer

<div><p>Objective</p><p>DNA methyltransferase 3b (DNMT3b) plays an important role in abnormal methylation during tumorigenesis. Polymorphism of the <i>DNMT3b</i> gene may influence <i>DNMT3b</i> activity and be associated with cancer risk. This study aimed to investigate the association between single nucleotide polymorphisms (SNPs) of the <i>DNMT3b</i> gene and susceptibility and prognosis of gastric cancer.</p><p>Methods</p><p>Four hundred and forty-seven histologically-confirmed gastric cancer cases, 111 gastric atrophy cases and 961 tumor-free controls were enrolled into the study. Five tag SNPs (rs6119954, rs1569686, rs4911107, rs4911259 and rs8118663) of the <i>DNMT3b</i> gene were genotyped by TaqMan assay. DNMT3b expression was evaluated in 104 cancer tissues by immunohistochemistry method.</p><p>Results</p><p>The median follow-up time for 422 gastric patients with prognosis information was 55.1 (51.8–58.5) month. We found that individuals with the rs1569686 variant genotype (TG/GG) were significantly associated with poor prognosis in gastric cancer compared to those carrying the TT genotype (HR = 1.43, 95%CI: 1.02–1.99). This trend was more evident in the long-term survival of gastric cancer. Similar results were observed for the G allele carriers of rs4911107 and T allele carriers of rs4911259 as these two sites were in complete linkage disequilibrium with rs1569686. The rs8118663 GG carriers tended to live shorter than AA/AG genotype (HR = 2.72, 95%CI: 1.45–5.12) in patients living longer than 2.0 years. None of the five SNPs was associated with the risks of gastric cancer or gastric atrophy. And no relationship was found between each of the five SNPs and DNMT3b expression.</p><p>Conclusions</p><p>This study provides evidence that <i>DNMT3b</i> polymorphisms may predict long-term survival of gastric cancer. However, further studies are needed to reveal the underlying biological roles of <i>DNMT3b</i> polymorphism.</p></div

Quantification of GUS activity in leaves of transgenic Arabidopsis transformed with <i>ProEgFAD8</i>::<i>GUS</i> under the treatment of darkness or low temperature (15°C) for 24h and 48h.

<p>Quantification of GUS activity in leaves of transgenic Arabidopsis transformed with <i>ProEgFAD8</i>::<i>GUS</i> under the treatment of darkness or low temperature (15°C) for 24h and 48h.</p

List of primers used in the study.

<p>List of primers used in the study.</p

Influence of DNMT3b polymorphisms on expression of DNMT3b.

<p>Influence of DNMT3b polymorphisms on expression of DNMT3b.</p

Characteristics of the subjects.

<p>TNM: Tumor, Lymph Node, Metastasis.</p><p>^aFOLFOX-4 (5-fluorouracil, leucovorin and oxaliplatin).</p><p>^bXELOX (capecitabine and oxaliplatin).</p><p>^cOther chemotherapies included: 5-fluorouracil; xeloda alone; paclitaxel plus leucovorin and tegafurum; LV5-FU2 (leucovorin plus 5-fluorouracil); FOLFIRI (irinotecan, 5-fluorouracil and leucovorin).</p><p>Characteristics of the subjects.</p

Survival plots of gastric cancer patients.

<p>(A) plot for rs1569686 using the dominant model (TG/GG <i>vs</i>.TT). Patients carrying rs1569686 TG/GG genotype tended to live shorter than those carrying TT genotype. This trend was more evident in patients who lived longer than 2.0 years with a hazard ratio (HR) 2.46 (95% CI 1.29–4.69) after adjusting for age, sex TNM stage and chemotherapy type. (B) plot for rs8118663 using the recessive model (GG vs. AA/AG). Similar to rs1569686, rs8118663 GG carriers tended to have shorter survival time.</p