The Longissimus and Semimembranosus muscles display marked differences in their gene expression profiles in pig

Abstract

Background: Meat quality depends on skeletal muscle structure and metabolic properties. While most studies carried on pigs focus on the Longissimus muscle (LM) for fresh meat consumption, Semimembranosus (SM) is also of interest because of its importance for cooked ham production. Even if both muscles are classified as glycolytic muscles, they exhibit dissimilar myofiber composition and metabolic characteristics. The comparison of LM and SM transcriptome profiles undertaken in this study may thus clarify the biological events underlying their phenotypic differences which might influence several meat quality traits.

Methodology/principal findings: Muscular transcriptome analyses were performed using a custom pig muscle microarray: the 15 K Genmascqchip. A total of 3823 genes were differentially expressed between the two muscles (Benjamini-Hochberg adjusted P value ≤0.05), out of which 1690 and 2133 were overrepresented in LM and SM respectively. The microarray data were validated using the expression level of seven differentially expressed genes quantified by real-time RT-PCR. A set of 1047 differentially expressed genes with a muscle fold change ratio above 1.5 was used for functional characterization. Functional annotation emphasized five main clusters associated to transcriptome muscle differences. These five clusters were related to energy metabolism, cell cycle, gene expression, anatomical structure development and signal transduction/immune response.

Conclusions/significance: This study revealed strong transcriptome differences between LM and SM. These results suggest that skeletal muscle discrepancies might arise essentially from different post-natal myogenic activities.

Figure 1. Gene expression ratio between muscles.

Muscle fold change ratio is expressed as the expression ratio of Longissimus (LM) to Semimembranosus (SM) samples when genes are highly expressed in Longissimus and as the expression ratio of SM to LM samples when genes are highly expressed in Semimembranosus.

Figure 2. Validation of seven microarray differentially expressed genes between Longissimus (LM) and Semimembranosus (SM) muscles by quantitative RT-PCR.

mRNA level is expressed using arbitrary units. Quantitative RT-PCR expression levels (LM = 8, SM = 8) were normalized to the expression of beta 2 microglobulin (B2M), TATAbox binding protein (TBP) and 18S using geNorm algorithm. Microarray adjusted means for LM and SM (LM = 90; SM = 90) were calculated using least square means for the muscle effect. Data are expressed as means±s.d. Statistical significances are reported below the plot as Benjamini and Hochberg adjusted P-value for microarray data and as Student t-test P value for q RT-PCR. Fold change ratio is expressed as the expression ratio of LM to SM when genes are overrepresented in LM and as the expression ratio of SM to LM when genes are overrepresented in SM.

Figure 3. Hierarchical clustering of differentially expressed genes according to their GO BP terms semantic similarity.

Annotated differentially expressed genes with a muscle fold change above 1.5 were clustered based on their functional annotation (GO BP) semantic similarity. Hierarchical clustering was performed using “1-semantic similarity” as distance between two genes (similar genes have a distance close to zero) to identify clusters of genes sharing BP terms. Five clusters were identified. Cluster 1 comprised 98 genes highly expressed in SM and 44 in LM. Cluster 2 included 73 highly expressed genes in SM and 102 in LM. Cluster 3 contained 43 highly expressed genes in SM and 84 in LM. Cluster 4 comprised 288 genes overexpressed in SM and 192 in LM. Cluster 5 involved 90 overexpressed genes in SM and 33 overexpressed genes in LM.