Phylogenetic analysis and syntenic location of elovl8. (A) Phylogenetic analysis of Elovl1, Elovl7, Elovl4, and Elovl8 sequences; values at node correspond to posterior probabilities provided by aBayes. Tree was rooted at midpoint. (B) Syntenic location of the Elovl8 genes in several species; Elovl8 gene is represented by black box; dotted black box in human represents a pseudogene; color code of the remaining boxes is conserved corresponding to the same gene identified in several species. Genes identified in a limited number of species with limited or no cross species conservation indicated in grey.

The mRNA expression levels of elovl8a and elovl8b in different fatty acids treatment zebrafish liver (ZFL) cell. (A,B) The expression levels of elovl8a in ZFL cells supplemented with SFAs (A) or PUFAs (B). (C,D) The expression levels of elovl8b in SFAs (C) or PUFAs (D) treatment ZFL cell. The statistical analyses were conducted by t test. Data were expressed as mean ± SD (standard deviation) of three biological replicates. The asterisks labeled above the error bars indicated significant differences (* p < 0.05). SFAs, saturated fatty acids; MUFAs, monounsaturated fatty acids; PUFAs, polyunsaturated fatty acids; elovl, elongation of very long-chain fatty acid protein.

Effects of elovl8a and elovl8b knockdown on liver fatty acid composition. (A) The expression level of elovl8a in si:elovl8a treated ZFL cells. (B) The expression level of elovl8b in si:elovl8b treated ZFL cells. (C) PUFA composition of control and si:elovl8a treated ZFL cells. (D,E) SFA (D) and MUFA (E) composition of control and si:elovl8b treated ZFL cells. The statistical analyses were conducted by t test. Data were expressed as mean ± SD (standard deviation) of three biological replicates. The asterisks labeled above the error bars indicate significant differences (* p < 0.05). NC, negative control; SFAs, saturated fatty acids; MUFAs, monounsaturated fatty acids; PUFAs, polyunsaturated fatty acids; elovl, elongation of very long-chain fatty acid protein.

elovl8a and elovl8b gene deletion in zebrafish and effects of elovl8a and elovl8b deletion on liver fatty acid compositions. (A,B) The targeting site for elovl8a (A) and elovl8b (B) gene knockout. The gray boxes were the 5′-untranslated region and 3′-untranslated region and black boxes were the exons. The red box indicated the targeting sequences. (C) The PUFA composition in liver of wild-type (WT) and elovl8a^−/−. (D) The ratio of C20:2n-6/C18:2n-6, C20:3n-3/C18:3n-3, and C22:5n-3/C20:5n-3 in the liver of WT and elovl8a^−/−. (E) The SFA and MUFA composition in the liver of WT and elovl8b^−/−. (F) The ratio of C20:0/C18:0 in the liver of WT and elovl8b^−/−. The statistical analyses were conducted by the t test. Data were expressed as mean ± SD (standard deviation) of four biological replicates. Asterisks above the error bars indicate significant differences (* p < 0.05. aa, amino acid; elovl, elongation of very long-chain fatty acid protein; SFAs, saturated fatty acids; MUFAs, monounsaturated fatty acids; PUFAs, polyunsaturated fatty acids.

The expression levels of elovl8a and elovl8b in liver of diet-treatment zebrafish and other elongase knockout zebrafish. (A) The expression levels of elovl8a in the liver of C18:3n-3 diet-treatment zebrafish. (B,C) The expression levels of elovl8b in the liver of C18:0 and C20:0 diet-treatment zebrafish. (D) The expression levels of elovl2, elovl4s, and elovl5 in the liver of elovl8a knockout zebrafish (elovl8a^−/−). (E) The expression levels of elovl1s, elovl3s, and elovl7s in the liver of elovl8b knockout zebrafish (elovl8b^−/−). (F) The expression levels of elovl8a in the liver of elovl5 knockout zebrafish (elovl5^−/−). (G–I) The expression levels of elovl8b in the liver of elovl1a knockout zebrafish (elovl1a^−/−) (G), elovl1b knockout zebrafish (elovl1b^−/−) (H), and elovl3b knockout zebrafish (elovl3b^−/−) (I). The statistical analyses were conducted by the t test. Data were expressed as mean ± SD (standard deviation) of three biological replicates. Asterisks above the error bars indicate significant differences (* p < 0.05, ** p < 0.01). WT, wild type zebrafish; elovl, elongation of very long-chain fatty acid protein.

The schematics of biosynthesis pathways of fatty acid synthesis in teleosts. SFAs, saturated fatty acids; MUFAs, monounsaturated fatty acids; PUFAs, polyunsaturated fatty acids; elovl, elongation of very long-chain fatty acid protein.