Exon-intron split analysis reveals posttranscriptional regulatory signals induced by high and low n-6/n-3 polyunsaturated fatty acid ratio diets in piglets

Polyunsaturated fatty acids (PUFA), such as omega-6 (n-6) and omega-3 (n-3), play a vital role in nutrient metabolism, inflammatory response, and gene regulation. microRNAs (miRNA), which can potentially degrade targeted messenger RNAs (mRNA) and/or inhibit their translation, might play a relevant role in PUFA-related changes in gene expression. Although differential expression analyses can provide a comprehensive picture of gene expression variation, they are unable to disentangle when in the mRNA life cycle the regulation of expression is taking place, including any putative functional miRNA-driven repression. To capture this, we used an exon-intron split analysis (EISA) approach to account for posttranscriptional changes in response to extreme values of n-6/n-3 PUFA ratio. Longissimus dorsi muscle samples of male and female piglets from sows fed with n-6/n-3 PUFA ratio of 13:1 (SOY) or 4:1 (LIN), were analyzed in a bidirectional contrast (LIN vs. SOY, SOY vs. LIN). Our results allowed the identification of genes showing strong posttranscriptional downregulation signals putatively targeted by significantly upregulated miRNA. Moreover, we identified genes primarily involved in the regulation of lipid-related metabolism and immune response, which may be associated with the pro- and anti-inflammatory functions of the n-6 and n-3 PUFA, respectively. EISA allowed us to uncover regulatory networks complementing canonical differential expression analyses, thus providing a more comprehensive view of muscle metabolic changes in response to PUFA concentration. Exon-intron split analysis reveals posttranscriptional regulatory signals and genes involved in lipid metabolism and immune signaling in response to high and low n-6/n-3 polyunsaturated fatty acid ratio diets in piglets. The relationship between dietary lipids, such as omega-6 and omega-3 polyunsaturated fatty acids (PUFA), and gene expression regulation was explored in piglet muscle. While these PUFA can influence nutrient metabolism and inflammatory response, small regulatory molecules called microRNAs (miRNA) can also influence the activity of genes. In this experiment, we used a computational approach dubbed exon-intron split analysis (EISA) to fully understand the role of miRNA in this context and how the genes and miRNA respond to changes in PUFA levels. Our findings demonstrated that some genes involved in lipid metabolism and immune response were affected by different PUFA concentrations and that EISA provides a more comprehensive view of how genes are regulated throughout their life cycle.