Submesoscale characteristics and transcription of a fatty acid elongase gene from a freshwater green microalgae, Myrmecia incisa Reisigl

Myrmecia incisa is a green coccoid freshwater microalgae, which is rich in arachidonic acid （ArA, C20： 4ω-6, △5, 8, 11, 14）, a long chain polyunsaturated fatty acid （PUFA）, especially under nitrogen starvation stress. A cDNA library of M. incisa was constructed with λ. phage vectors and a 545 nt expressed sequence tag （EST） was screened from this library as a putative elongase gene due to its 56% and 49% identity to Marchantia polymorpha L. and Ostreococcus tauri Courties et Chretiennot-Dinet, respectively. Based upon this EST sequence, an elongase gene designated MiFAE was isolated from M. incisa via 5＇/3＇ rapid amplification of cDNA ends （RACE）. The cDNA sequence was 1 331 bp long and included a 33 bp 5＇-untranslated region （UTR） and a 431 bp 3＇-UTR with a typical poly-A tail. The 867 bp ORF encoded a predicted protein of 288 amino acids. This protein was characterized by a conserved histidine-rich box and a MYxYY motif that was present in other members of the elongase family. The genomic DNA sequence of MiFAE was found to be interrupted by three introns with splicing sites of Introns I （81 bp）, II （81 bp）, and III （67 bp） that conformed to the GT-AG rule. Quantitative real-time PCR showed that the transcription level of MiFAE in this microalga under nitrogen starvation was higher than that under normal condition. Prior to the ArA content accumulation, the transcription of MiFAE was enhanced, suggesting that it was possibly responsible for the ArA accumulation in this microalga cultured under nitrogen starvation conditions.

Molecular evidence for blocking erucic acid synthesis in rapeseed(Brassica napus L.) by a two-base-pair deletion in FAE1(fatty acid elongase 1)

DNA sequences of fatty acid elongase 1 genes FAE1.1 （EA） and FAE1.2 （Ec） were isolated and characterized for 30 com- mercialized low erucic acid rapeseed （LEAR） cultivars in China. Four types of independent mutation leading to low erucic acid trait were found, i.e., a single-base transition （eAl）, a two-base deletion （ec2） and four-base deletion （eC4） as well as single-base transition with a four-base deletion （eA.）. Three genotypes, i.e., eA1eA1eC2eC2, eA1eA1eC4eC4 and eA.eA.ec4ec4 were responsible for LEA content in storage Iipids of different rapeseed cultivars. Most of the LEAR cultivars had a genotype of eA1eA1ec2ec2, which were descended from the first LEAR cultivar, Oro. Yeast expression analysis revealed that two-base-pair （AA） deletion （ec2） at the base sites of 1 422-1 423 in the C genome FAE1 gene resulted in the absence of the condensing enzyme and led to the failure to produce erucic acid. Coexpression of FAE1 and ketoacyI-CoA reductase （KCR） or enoyI-CoA reductase （ECR） was found in high erucic acid rapeseed （HEAR） but not in LEAR （eA1eA1ec2ec2oreA1eA1ec4ec4）. Moreover, KCR and ECR were still coordinately regulated in eA1eA1ec2ec2 or eA1eA1ec4ec4 genotypes, suggesting that the expression of two genes was tightly linked. In addition, specific detection methods were developed by high-resolution melting curve analysis in order to detect eA1 and ec4.

Cloning of fatty acid elongase1 gene and molecular identification of A and C genome in Brassica species

The fatty acid elongase 1 (FAE1) genes of Brassic napus were cloned from two cultivars, i.e. Zhong- shuan No. 9 with low erucic acid content, and Zhongyou 821 with high erucic acid content, using the degenerate PCR primers. The sequence analysis showed that there was no intron within the FAE1 genes. The FAE1 genes from Zhongyou 821 contained a coding sequence of 1521 nucleotides, and those cloned from Zhongshuan No. 9 contained a 1517 bp coding sequence. Alignment of the FAE1 sequences from Brassica rapa, B. oleracea and B. napus detected 31 single nucleotide polymorphic sites (2.03%), which resulted in 7 amino-acid substitutions. Further analysis indicated that 19 SNPs were genome-specific, of which, 95% were synonymous mutations. The nucleotide substitution at po- sition 1217 in the FAE1 genes led to a specific site of restricted cleavage. An AvrII cleavage site was present only in the C genome genes and absent in the A genome FAE1 genes. Digestion profile of the FAE1 sequences from B. rapa, B. oleracea and B. napus produced with AvrII confirmed that the FAE1 genes of B. oleracea origin was recognized and digested, while that of B. rapa origin could not. The results indicated that by AvrII cleavage it was possible to distinguish B. rapa from B. oleracea and be- tween the A and C genome of B. napus. In addition, the FAE1 genes could be used as marker genes to detect the pollen flow of B. napus, thus providing an alternative method for risk assessment of gene flow.

Roles of vitamin A in the regulation of fatty acid synthesis

Dietary macronutrients and micronutrients play important roles in human health.On the other hand,the excessive energy derived from food is stored in the form of triacylglycerol.A variety of dietary and hormonal factors affect this process through the regulation of the activities and expression levels of those key player enzymes involved in fatty acid biosynthesis such as acetyl-CoA carboxylase,fatty acid synthase,fatty acid elongases,and desaturases.As a micronutrient,vitamin A is essential for the health of humans.Recently,vitamin A has been shown to play a role in the regulation of glucose and lipid metabolism.This review summarizes recent research progresses about the roles of vitamin A in fatty acid synthesis.It focuses on the effects of vitamin A on the activities and expression levels of mRNA and proteins of key enzymes for fatty acid synthesis in vitro and in vivo.It appears that vitamin A status and its signaling pathway regulate the expression levels of enzymes involved in fatty acid synthesis.Future research directions are also discussed.