Impact of genotype, body weight and sex on the prenatal muscle transcriptome of Iberian pigs

Abstract

Growth is dependent on genotype and diet, even at early developmental stages. In this study, we investigated the effects of genotype, sex, and body weight on the fetal muscle transcriptome of purebred Iberian and crossbred Iberian x Large White pigs sharing the same uterine environment. RNA sequencing was performed on 16 purebred and crossbred fetuses with high body weight (340±14g and 415±14g, respectively) and 16 with low body weight (246±14g and 311±14g, respectively), on gestational day 77. Genotype had the greatest effect on gene expression, with 645 genes identified as differentially expressed (DE) between purebred and crossbred animals. Functional analysis showed differential regulation of pathways involved in energy and lipid metabolism, muscle development, and tissue disorders. In purebred animals, fetal body weight was associated with 35 DE genes involved in development, lipid metabolism and adipogenesis. In crossbred animals, fetal body weight was associated with 60 DE genes involved in muscle development, viability, and immunity. Interestingly, the results suggested an interaction genotype*weight for some DE genes. Fetal sex had only a modest effect on gene expression. This study allowed the identification of genes, metabolic pathways, biological functions and regulators related to fetal genotype, weight and sex, in animals sharing the same uterine environment. Our findings contribute to a better understanding of the molecular events that influence prenatal muscle development and highlight the complex interactions affecting transcriptional regulation during development.

Fig 1. Venn diagram showing the total number of differentially expressed (DE) genes in all comparisons.

Fig 2

Top canonical pathways (p < 0.005) identified by functional analysis of DE genes in crossbred fetuses (orange bars) and Iberian fetuses (blue bars).

Fig 3. Enrichment of the PPAR signaling pathway in muscle from Iberian fetuses.

Fig 4. Functional network of Metabolic Disease, Cellular Movement, as well as Skeletal and Muscular System Development and function identified by analysis of DE genes in Iberian and crossbred fetuses.

The intensity of colors represents the magnitude of differential expression, with red indicating genes upregulated in crossbred fetuses, while green indicates those upregulated in Iberian fetuses. Direct interactions are represented by solid lines, while indirect interactions are represented by dashed lines. The shapes of the nodes reflect the functional class of each gene product: transcriptional regulator (horizontal ellipse), transmembrane receptor (vertical ellipse), enzyme (vertical rhombus), cytokine/growth factor (square), kinase (inverted triangle) and complex/group/other (circle).

Fig 5. Top-scoring network of regulatory effects in crossbred fetuses.

Upper tier: upstream regulators predicted to be activated (orange color). Middle tier: genes with altered expression in response to activation of upstream regulators (red = upregulation in crossbred; green = downregulation in crossbred). The shapes of the nodes reflect the functional class of each gene product: horizontal ellipse, transcriptional regulator; vertical ellipse, transmembrane receptor; vertical rhombus, enzyme; square, cytokine/growth factor; inverted triangle, kinase; and circle, complex/group/other. Direct interactions are represented by solid lines, while indirect interactions are represented by dashed lines. Orange lines lead to activation, while blue lines lead to inhibition. Lower tier: expected phenotypic consequences of gene expression changes based on data in the Ingenuity Knowledge Base (absolute z-score > 2 and p-value < 0.05). The octagonal symbol defines function.

Fig 6. Top-scoring regulator effects network activated in crossbred LBW fetuses.

Upper tier: one upstream regulator is predicted to be activated (blue). Middle tier: gene expression changes in response to activation of upstream regulators (green = upregulated in small fetuses). The shapes of the nodes reflect the functional class of each gene product: horizontal ellipse, transcriptional regulator; vertical ellipse, transmembrane receptor; and circle, complex/group/other. Direct interactions are represented by solid lines, while indirect interactions are represented by dashed lines. Orange lines lead to activation. Lower tier: expected phenotypic consequences of gene expression changes based on data in the Ingenuity Knowledge Base (absolute z-score > 2 and p < 0.05). The octagonal symbol defines function.

Fig 7. Top-scoring network of regulator effects activated in Iberian HBW fetuses.

Upper tier: one upstream regulator is predicted to be activated (orange). Middle tier: gene expression changes in response to activation of upstream regulators (red = upregulated in large fetuses). The shapes of the nodes reflect the functional class of each gene product: horizontal ellipse, transcriptional regulator; vertical rhombus, enzyme; inverted triangle, kinase; and circle, complex/group/other. Direct interactions are represented by solid lines, while indirect interactions are represented by dashed lines. Orange lines lead to activation, while blue lines lead to inhibition. Lower tier: expected phenotypic consequences of gene expression changes based on data in the Ingenuity Knowledge Base (absolute z-score > 2 and p < 0.05). The octagonal symbol defines function.