The mRNA expression of myogenic regulatory factors including myoD1 gene paralogues, and their regulation by amino acids and insulin-like growth factors was investigated in primary cell cultures isolated from fast myotomal muscle of Atlantic salmon (Salmo salar). The cell cycle and S-phase was determined as 28.1 h and 13.3 h respectively at 18°C. MyoD1b and myoD1c expression peaked at 8 d of culture in the initial proliferation phase then declined >6-fold as cells differentiated and was correlated with PCNA expression (R=0.88 P<0.0001, R=0.70 P<0.0001). In contrast, myoD1a transcripts increased from 2d to 8d and remained at elevated levels as myotubes were formed. MyoD1c mRNA levels were on average 3.1 and 5.7-fold higher than myoD1a and myoD1b respectively. Depriving cells of amino acids and serum led to a rapid increase in pax7 and a decrease in myoD1c and PCNA expression, indicating a transition to a quiescent state. In contrast, amino acid replacement in starved cells produced significant increases in myoD1c (at 6h), PCNA (at 12h) and myoD1b (at 24h), and decreased pax7 expression as cells entered the cell cycle. Our results are consistent with temporally distinct patterns of myod1c and myoD1b expression at the G1 and S/G2 phases of the cell cycle. Treatment of starved cells with IGF-I or IGF-II did not alter expression of the myoD paralogues. It was concluded that in vitro, amino acids alone are sufficient to stimulate expression of genes regulating myogenesis in myoblasts involving autocrine/paracrine pathways. The differential responses of myoD paralogues during myotube maturation and amino acid treatments suggest myoD1b and myoD1c are primarily expressed in proliferating cells and myoD1a in differentiating cells, providing evidence for their sub-functionalisation following whole genome and local duplications in the Atlantic salmon lineage.
- cell cycle
- amino acid stimulation
- Copyright © 2010, American Journal of Physiology - Regulatory, Integrative and Comparative Physiology