Molecular cloning and characterization of porcine calcineurin-alpha subunit expression in skeletal muscle.

The calmodulin/Ca2+-dependent serine/threonine phophatase, calcineurin (CaN), has been implicated in controlling muscle fiber phenotype. However, little information is available concerning the expression of CaN in porcine skeletal muscle. Therefore, the porcine CaN alpha (CaN-A) was cloned by reverse transcription-PCR and its expression characterized in selected porcine skeletal muscles. We successfully cloned porcine CaN gene using semitendinosus muscle (GenBank accession number AF193515). Sequence analysis showed both the full length and a 30-bp deletion splice variant in coding region of the gene reported in other species. The deduced AA sequence showed 99.4% homology with the rat CaN-A delta isoform gene. Real-time PCR analysis showed CaN is present in all tissues. However, using primers targeting the region containing the 30-bp deletion, the full length sequence is only found in skeletal muscle and brain tissues. Using a CaN-A monoclonal antibody, we localized CaN-A in porcine LM and soleus muscle and the red and white portions of the semitendinosus muscle. The CaN-A protein was abundant in fast fibers and primarily localized in the cytoplasm, whereas slow fibers expressed reduced abundance of CaN-A. Further studies are required to understand the functions of CaN-A isoform in skeletal muscle.

Paylean alters myosin heavy chain isoform content in pig muscle.

Feeding beta-adrenergic agonists promotes muscle growth. Early histological techniques failed to show precisely how feeding ractopamine-HCl (Paylean) alters muscle growth in pigs. To understand these effects, an indirect enzyme-linked immunosorbent assay (ELISA) was used to determine the abundance of each adult skeletal muscle myosin heavy chain isoform, one means of assigning muscle fiber type, in fast and slow muscles of pigs fed Paylean. Sixty growing pigs (-85 kg) were randomly assigned to three Paylean doses (0, 20, or 60 ppm). At 3, 7, 14, 28, and 42 d of treatment, four pigs per dose were harvested and white (WST) and red (RST) semitendinosus and longissimus (LM) muscles were removed and processed, and myosin heavy chain was quantified by ELISA. Feeding Paylean enhanced (P < 0.05) pigs' average daily gain. Muscle myosin heavy chain (slow, 2A, 2AX, and 2B) composition differed (P < 0.05) across muscles. Compared with LM, RST contained approximately five times more (P < 0.0001) slow and type 2A myosin heavy chain and three times more 2AX myosin heavy chain but nearly undetectable amounts of 2B myosin heavy chain. Myosin heavy chain composition of the WST closely resembled that of the LM (i.e., greater 2AX and 2B and less slow and 2A). After 42d of 60 ppm Paylean, the amount of slow, 2A, and 2AX myosin heavy chain decreased (P < 0.05) across the three muscles whereas the amount of 2B myosin heavy chain increased (P < 0.05). In contrast, relative amounts of 2A and 2AX myosin heavy chain increased (P < 0.05) in muscle of control pigs at 42d. Changes associated with the 20-ppm dose were intermediate to and different from (P < 0.05) control and 60 ppm treatments. Correlations (P < 0.05) among various myosin heavy chain within muscles suggest that slow, type 2A, and 2X decrease with increases in 2B myosin heavy chain. These data show that administration of Paylean affects myosin heavy chain isoform composition in a time- and dose-dependent manner and provides a mechanism of action for Paylean altering animal growth.

Myosin heavy chain isoforms account for variation in pork quality.

The objective of this study was to determine the impact of myosin heavy chain (MyHC) isoforms (I, IIB, IIA and IIX) on pork quality traits of halothane (HAL)-negative (NN) and halothane-carrier (Nn) pigs. Gilts (n=32) were assigned to a 2×2 factorial of genetic population (GP) and slaughter weight (WT; 120 and 135 kg). Classical meat quality characteristics were collected and MyHC content was determined on muscle samples. Regression equations for pork quality and carcass composition traits were determined. Only I/IIB accounted for variation in drip loss of NN gilts (R(2)=0.18), while GP related to drip loss in Nn gilts (R(2)=0.70). Type I MyHC content explained variation in ultimate (24 h) muscle pH of NN gilts (R(2)=0.09), while I/IIB, I/IIX and IIB/IIX were significant for Nn gilts (R(2)=0.99). I/IIA, I/IIX, IIB/IIX and GP accounted for variation in Hunter Color a (redness) values of NN gilts (R(2)=0.69), while IIB, IIA, IIB/IIA and GP were significant for Nn gilts (R(2)=0.97). Overall, fiber type composition accounts for a larger proportion of variation in the quality traits of Nn compared to NN gilts.