Research ArticleORGANISMAL BIOLOGY

Genes for de novo biosynthesis of omega-3 polyunsaturated fatty acids are widespread in animals

See allHide authors and affiliations

Science Advances  02 May 2018:
Vol. 4, no. 5, eaar6849
DOI: 10.1126/sciadv.aar6849
  • Fig. 1 Distribution of ωx desaturases.

    (A) A condensed phylogenetic tree depicting relationships among ωx desaturases from Cyanobacteria (8) and Eukaryota (279). Metazoan phyla (represented with 121 sequences) are named in the tree and assigned with colors for the following taxa: Nematoda (green), Cnidaria (blue), Arthropoda (purple), and Lophotrochozoa (orange). Nonmetazoan clades were collapsed and named according to their general taxonomic composition. “Miscellaneous Clade A” contains representatives of Amoeboza, Apusozoa, Haptophyceae, Heterolobosea, Nucleariidae, SAR, Chlorophyta, Rhodophyta, Cryptophyta, and the insect L. migratoria. Representatives of Excavata, Amoebozoa, Fungi, SAR, and the hexapod S. viridis compose “Miscellaneous Clade B.” The phylogenetic tree presented corresponds to the 80% rule consensus tree estimated in MrBayes. Colored dots on the nodes represent supporting values of 80% or above for MrBayes and PhyloBayes analyses (blue) or for all three (MrBayes, PhyloBayes, and RAxML) analyses (red). (B) Summary of taxonomic groups highlighting metazoans in which ωx desaturases have been identified in this study. Number of species is indicated in brackets. A list of all genomes and transcriptomes searched, including those in which no ωx desaturase genes were found, can be accessed as described in the Data and materials availability section.

  • Fig. 2 HGT accounts for the presence of ωx desaturase in some animal genomes.

    (A) Multiple copies of ωx desaturase–like sequences scattered among several loci of the B. tabaci genome. The ωx desaturase–like sequences are found in five different scaffolds and are flanked by animal genes. * indicates sequences that did not pass the filter (probability between 0.50 and 0.70). (B) Intron-exon structures in the ωx desaturase genes from selected metazoan species. The arrows indicate aligned coding sequences (CDS) extracted from the genomic assembly of each species. The positions of introns are indicated as solid lines within the arrows. The dotted line indicates shared intron-exon boundaries. All cnidarian genes and several genes from Annelida and Mollusca have an “intronless” organization. The CDS of the three ωx desaturase genes from A. vaga (Rotifera) share the same overall structure consisting of eight exons. In addition, all genes from three Heterobranchia species have the same structure with six exons in the CDS. Variable gene structures were found in Nematoda, Annelida, and Arthropoda. The arrow colors are consistent with those used to represent clades in Fig. 1.

  • Fig. 3 Functional characterization of metazoan ωx desaturases.

    (A) The de novo production of ω3 PUFA requires both ω6 (Δ12) (blue arrow) and ω3 (Δ15) desaturases (red arrow). Both LA (18:2ω6) and ALA (18:3ω3) can be subsequently modified through the ω6 and ω3 long-chain PUFA biosynthesis pathways that proceed separately, at least in vertebrates (9), or can be interconnected by ω3 desaturases with Δ15, Δ17, or Δ19 activities (green arrows). (B) Chromatograms of FA methyl esters (FAME) from yeast expressing ωx desaturases from A. vaga (MF448339) (top) and A. millepora (KY658237) (middle) show Δ12/Δ15 and Δ12 activities, respectively. Further ωx desaturases with Δ12 activity were characterized from P. vulgata, P. dumerilii, and L. salmonis (Table 1). (C) Chromatograms of FAME from yeast expressing the ωx desaturase from P. vulgata (KY658241) and grown with supplemented ω6 substrates including 18:2ω6 (left), 20:4ω6 (middle), and 22:4ω6 (right) show the resulting ω3 products 18:3ω3, 20:5ω3, and 22:5ω3, respectively. Similar functions obtained for ωx desaturases from A. millepora, A. vaga, P. dumerilii, and L. salmonis are shown in Table 2. Molecular structures and diagnostic mass ions of 4,4-dimethyloxazoline derivatives from desaturation products are shown in figs. S3 and S4.

  • Table 1 Characterization of Δ12/Δ15 activities of metazoan ωx desaturases.

    FA profiles of yeast transformed with empty pYES2 (control) were compared with those of yeast transformed with the corresponding ωx desaturase [A. millepora (KY658237), A. vaga (ωx1, MF448337; ωx2, MF448339), P. vulgata (KY658242), P. dumerilii (KY658238), and L. salmonis (KY658240)]. FA compositions (% of total FA) are given as means ± SEM of replicate samples (n = 6 for control and n = 4 for transgenic yeast). An asterisk indicates significant differences between control and yeast expressing the corresponding desaturase (Dunnett’s test; *P < 0.05). nd, not detected.

    FAFA composition (% of total FA)
    ControlA. milleporaA. vagaP. vulgataP. dumeriliiL. salmonis
    ωx1ωx2
    16:029.0 ± 1.031.7 ± 1.025.7 ± 0.624.4 ± 0.7*24.8 ± 0.4*28.5 ± 1.122.6 ± 1.3*
    16:1ω735.0 ± 1.223.3 ± 2.4*22.4 ± 0.3*19.5 ± 0.5*36.4 ± 0.634.5 ± 1.037.6 ± 0.8
    16:2ω4nd2.8 ± 0.3*6.9 ± 0.3*10.4 ± 0.3*0.4 ± 0.10.2 ± 0.00.2 ± 0.0
    16:3ω1ndnd0.5 ± 0.0*1.5 ± 0.1*ndndnd
    18:010.1 ± 0.915.8 ± 0.6*15.8 ± 1.2*17.0 ± 1.0*10.6 ± 0.311.1 ± 0.411.7 ± 0.5
    18:1ω925.6 ± 0.613.9 ± 2.2*13.0 ± 1.4*8.6 ± 0.4*24.1 ± 1.021.2 ± 0.424.6 ± 0.5
    18:2ω60.3 ± 0.112.5 ± 0.7*13.8 ± 0.4*15.1 ± 0.5*3.8 ± 0.6*4.3 ± 0.2*3.3 ± 0.5*
    18:3ω3ndnd1.8 ± 0.1*3.4 ± 0.2*ndndnd

    †Mostly 16:1ω7, although it also includes some 16:1ω9.

    • Table 2 Functional characterization of metazoan ω3 desaturases.

      Transgenic yeast expressing the corresponding ω3 desaturases [A. millepora (KY658236), A. vaga (ωx1, MF448337; ωx2, MF448339), P. vulgata (KY658241), P. dumerilii (KY707886), and L. salmonis (KY658239)] were grown in the presence of exogenously added substrates 18:2ω6, 18:3ω6, 20:2ω6, 20:3ω6, 20:4ω6, and 22:4ω6. Conversions are expressed as a percentage of total FA substrate converted to desaturated product, with the corresponding activity (Δ) detected also shown. nd, not detected.

      SubstrateProductConversion (%)Activity
      A. milleporaA. vagaP. vulgataP. dumeriliiL. salmonis
      ωx1ωx2
      18:2ω618:3ω370.510.611.734.79.733.6Δ15
      18:3ω618:4ω364.814.314.318.86.539.6Δ15
      20:2ω620:3ω315.11.12.211.39.510.4Δ17
      20:3ω620:4ω325.74.24.516.115.814.2Δ17
      20:4ω620:5ω332.01.02.738.013.330.4Δ17
      22:4ω622:5ω32.8ndnd3.412.06.1Δ19

    Supplementary Materials

    • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/4/5/eaar6849/DC1

      fig. S1. Complete view of the phylogenetic tree presented in Fig. 1A, with major metazoan taxonomic clades colored as in Fig. 1.

      fig. S2. Summary tree (cladogram) showing relationships between major well-supported clades together with sequence logo representation of conserved histidine box motifs in the same clades.

      fig. S3. Mass spectra of DMOX derivatives obtained from desaturation products as shown in Table 1.

      fig. S4. Mass spectra of DMOX derivatives obtained from desaturation products as shown in Table 2.

      table S1. List of databases and query sequences for retrieval of functionally characterized genes.

      table S2. PCR conditions for the RACE and full-length ORF amplifications.

      table S3. List of primers used for the cloning process and amplification of full-length ORF of ωx desaturases.

    • Supplementary Materials

      This PDF file includes:

      • fig. S1. Complete view of the phylogenetic tree presented in Fig. 1A, with major metazoan taxonomic clades colored as in Fig. 1.
      • fig. S2. Summary tree (cladogram) showing relationships between major well-supported clades together with sequence logo representation of conserved histidine box motifs in the same clades.
      • fig. S3. Mass spectra of DMOX derivatives obtained from desaturation products as shown in Table 1.
      • fig. S4. Mass spectra of DMOX derivatives obtained from desaturation products as shown in Table 2.
      • table S1. List of databases and query sequences for retrieval of functionally characterized genes.
      • table S2. PCR conditions for the RACE and full-length ORF amplifications.
      • table S3. List of primers used for the cloning process and amplification of full-length ORF of ωx desaturases.

      Download PDF

      Files in this Data Supplement:

    Navigate This Article