Lipid fatty acid composition of potato plants transformed with the Δ12-desaturase gene from cyanobacterium

Potato (Solanum tuberosum L.) plants were transformed with the desA gene encoding Δ12 acyl-lipid desaturase in the cyanobacterium Synechocystis sp. PCC 6803. To evaluate the efficiency of this gene expression in the plant, its sequence was translationally fused with the sequence of the reporter gene encoding thermostable lichenase. A comparison of native and hybrid gene expression showed that lichenase retained its activity and thermostability within the hybrid protein, whereas desaturase retained its capability of inserting the double bond in fatty acid (FA) chains and, thus, to modify their composition in membrane lipids. In most transformed plants, shoots contained higher amounts of polyunsaturated FAs, linoleic and linolenic (by 39–73 and 12–41%, respectively). The total absolute content of unsaturated FAs was also higher in transformants by 20–42% as compared to wild-type plants. When transformed plants were severely cooled (to ?7°C), the rate of their membrane lipid peroxidation was not enhanced, whereas in wild-type plants, it increased substantially (by 25%) under such conditions. These results could indicate a higher tolerance of transformed plants to low temperatures and the oxidative stress induced by hypothermia.     

Hormonal regulation of tissue-specific ectopic expression of an Arabidopsis endoplasmic reticulum-type ω-3 fatty acid desaturase (FAD3) gene

A common feature of the membrane lipids of higher plants is a large content of polyunsaturated fatty acids, which typically consist of dienoic and trienoic fatty acids. Two types of omega-3 fatty acid desaturase. which are present in the plastids and in the endoplasmic reticulum (ER), respectively, are responsible for the conversion of dienoic to trienoic fatty acids. To establish a system for investigating the tissue-specific, and hor-mone-regulated expression of the ER-type desaturase gene (FAD3), transgenic plants of Arabidopsis thaliana (L.) Heynh. containing the firefly luciferase gene (LUC) fused to the FAD3 promoter (FAD3::LUC) were constructed. At different times during plant development, FAD3::LUC was actively expressed at two major sites, the vegetative shoot meristem and the floral organs. Transgenic plants with LUC fused to the promoter of FAD7 (FAD7::LUC) which encodes plastid-type desaturase, were also constructed. FAD3::LUC and FAD7::LUC were expressed in the same organs during reproductive growth, but not during vegetative growth. In plants exposed to both auxin and cytokinin, FAD3::LUC expression was ectopically induced in the root tissues. However, this induction by auxin and cytokinin was inhibited when abscisic acid was also present. FAD3::LUC expression could be induced in the roots by auxin and cytokinin if the hormones were applied during vegetative growth, but not if they were applied during germination or reproductive growth. Analysis of the fatty acid composition in the roots of Arabidopsis fad mutant and wild-type plants confirmed that the response of FAD3::LUC expression to various hormones reflected the response of endogenous FAD3 gene expression. These results suggest that the expression of ER-type desaturase is regulated through synergistic and antagonistic hormonal interactions, and that such hormonal regulation and the tissue specificity of the expression of this gene are further modified in accordance with the growth phase in plant development.     

Expression of Acyl-lipid △12-desaturase Gene in Prokaryotic and Eukaryotic Cells and Its Effect on Cold Stress Tolerance of Potato

We report the expression profile of acyl-lipid Δ12-desaturase (desA) gene from Synechocystis sp. PCC6803 and its effect on cell membrane lipid composition and cold tolerance in prokaryotic (Escherichia coli) and eukaryotic (Solanum tuberosum) cells. For this purpose, a hybrid of desA and reporter gene encoding thermostable lichenase (licBM3) was constructed and used to transform these cells. The expression of this hybrid gene was measured using qualitative (Petri dish test, electrophoregram and zymogram) and quantitative methods (spectrometry and gas liquid chromatography assays). The maximum level of linoleic acid in the bacterial cells containing hybrid gene was 1.9% of total fatty acids. Cold stress tolerance assays using plant damage index and growth parameters showed that cold tolerance was enhanced in primary transgenic lines because of increased unsaturated fatty acid concentration in their lipids. The greatest content of 18:2 and 18:3 fatty acids in primary transgenic plants was observed for lines 2 (73%) and 3 (41%). Finally, our results showed that desaturase could enhance tolerance to cold stress in potato, and desaturase and lichenase retain their functionality in the structure of the hybrid protein where the enzymatic activity of target gene product was higher than in the case of reporter lichenase gene absence in the construction.     

The phylogenetic relationship of possible progenitors of the cultivated peanut

The cultivated peanut (Arachis hypogaea L.) is an allotetraploid composed of A and B genomes. The phylogenetic relationship among the cultivated peanut, wild diploid, and tetraploid species in the section Arachis was studied based on sequence comparison of stearoyl-ACP desaturase and oleoyl-PC desaturase. The topology of the trees for both fatty acid desaturases displayed two clusters; one cluster with A genome diploid species and the other with B genome diploid species. The two homeologous genes obtained for each of the two fatty acid desaturases from the tetraploid species A. hypogaea and A. monticola were separated into the A and B genome clusters, respectively. The gene phylogenetic trees showed that A. hypogaea is more closely related to the diploid species A. duranensis and A. ipaensis than to the wild tetraploid species A. monticola, suggesting that A. monticola is not a progenitor of the cultivated peanut. In addition, for the stearoyl-ACP desaturase, the A. duranensis sequence was identical with one of the sequences of A. hypogaea and the A. ipaensis sequence was identical with the other. These results support the hypothesis that A. duranensis and A. ipaensis are the most likely diploid progenitors of the cultivated tetraploid A. hypogaea.     

脂肪酸脱氢酶基因在兔体内表达的生物学效应研究

目的研究脂肪酸脱氢酶基因在兔子体内的生物效应,探索不饱和脂肪酸与兔子免疫的相互作用。方法用壳聚糖(CS)和壳聚糖的聚乙烯亚胺(PEI)、聚乙二醇(PEG)修饰产物(mPEG-O-CS-PEI),对含有脂肪酸脱氢酶基因的真核表达质粒VRMFat-1进行包裹,肌肉注射新西兰白兔,每两周采取抗凝血,检测血液中免疫细胞数量的变化情况,利用气相色谱仪检测动物组织中脂肪酸的含量变化,并通过荧光定量PCR检测免疫细胞内免疫相关基因的表达情况。结果两组实验兔组织中的n-3不饱和脂肪酸含量显著高于对照组(P<0.05),其TLR4、CD3、IL-4和IL-6基因表达水平显著升高(P<0.05)。结论转入的脂肪酸去饱和脱氢酶基因能够在实验兔体内正常表达,初步证明不饱和脂肪酸对兔子的固有免疫和获得性免疫机能均有明显的增强作用。     

Changes in membrane fatty acids and delta-9 desaturase in senescence accelerated (SAMP8) mouse hippocampus with aging

Senescence accelerated mice (SAMP8) exhibit age induced impairments such as loss of memory and learning disabilities by the age of 8-10 months. Analysis of hippocampus of SAMP8 mice revealed that delta 9-desaturase (delta9desaturase) activity reduced up to 44-50% with age. Correspondingly, levels of unsaturated fatty acids are also lowered in the aged animals approximately to the same levels. RNase protection assay showed that delta9specific message decreased similarly with age. As such a decrease is known to cause alterations in membrane fluidity and affect cellular signaling pathways, these results suggest that lowering of delta9gene expression may be partly involved in age induced impairments.