COUGHLIN, D.J.; DONATO, M. E. ; ULERICH, A. D.; SCHIAVI, J. M.; WEAVER, F. E.; Widener University, Chester, PA: Developmental Transitions in Myosin Light Chain 2 Expression in Trout Muscle
Rainbow trout (Oncorhynchus mykiss) display developmental shifts in the contractile properties of their swimming musculature. For instance, the red muscle of younger, smaller parr has faster contractile properties than that of older, larger smolts. Further, this shift in physiological properties is correlated with a change in myosin heavy chain expression. The red muscle of parr is more likely than that of smolts to express both slow and fast isoforms of MHC. We hypothesized that a similar variation in expression of myosin light chain 2 (MLC2) will be observed in the red and white muscle of the same juvenile stages. MLC2 (or regulatory light chain) is part of the myosin hexamer, the principle protein of muscle, and is thought to modulate muscle contractile properties. The objective of this project is to employ quantitative PCR (qPCR) to examine MLC2 expression levels in the red and white muscle of trout parr and smolt. Initially, two full length MLC2 cDNAs were cloned using RT-PCR and extended by 5� RACE. The cDNAs were sequenced and identified as slow and fast isoforms of MLC2 (sMLC2 and fMLC2) based on the muscle of origin. MLC2 isoform-specific qPCR primers were designed based on the sequence information. To examine expression levels, muscle samples were taken from three body positions (anterior, middle, and posterior) of red and white muscle of five parr and five smolt rainbow trout. Total RNA was extracted from the muscle samples, and single strand cDNA was obtained. QPCR with the isoform-specific primers permitted the quantification of mRNA expression levels of each isoform of MLC2 in red and muscle from the different body positions of each age of fish. A statistically significant down regulation in fMLC2 expression was observed in the transition from parr to smolt, in agreement with the earlier observations of MHC expression.
