Department of Nutrition, Exercise and Sports, Section of Integrated Physiology, University of Copenhagen, August Krogh Building, Universitetsparken 13, 2. sal, 2100 Copenhagen, Denmark.
J Physiol. 2013 Mar 15;591(6):1523-33. doi: 10.1113/jphysiol.2012.247312. Epub 2013 Jan 28.
The aim of the present study was to examine fibre type-specific Na(+)-K(+) pump subunit expression and exercise-induced alterations in phospholemman (FXYD1) phosphorylation in humans. Segments of human skeletal muscle fibres were dissected and fibre typed, and protein expression was determined by Western blotting. The protein expression of the Na(+)-K(+) pump α2 isoform was lower in type I than in type II fibres (0.63 ± 0.04 a.u. vs. 1.00 ± 0.07 a.u., P < 0.001), while protein expression of the Na(+)-K(+) pump α1 and β1 isoforms was not different. Protein expression of the ATP-dependent potassium channel Kir6.2 was higher in type I compared with type II fibres. In both type I (P < 0.01) and type II fibres (P < 0.001) the AB_FXYD1 signal was lower after exercise compared with rest, indicating an increase in unspecific FXYD1 phosphorylation. The FXYD1 serine 68 phosphorylation was higher (P < 0.001) after exercise compared with rest in type II fibres (1.90 ± 0.25 vs. 1.00 ± 0.08) and not changed in type I fibres. Total FXYD1 was not expressed in a fibre type-specific manner. Expression of phosphofructokinase was lower (P < 0.001) in type I than in type II fibres, whereas citrate synthase and 3-hydroxyacyl-CoA dehydrogenase were more abundant (P < 0.001) in type I fibres. In conclusion, FXYD1 phosphorylation at serine 68 increased after an acute bout of intense exercise in human type II fibres, while AB_FXYD1 signal intensity was lower in both type I and type II fibres, indicating fibre type-specific differences in FXYD1 phosphorylation on serine 63, serine 68 and threonine 69. This, together with the observation of a higher abundance of the Na(+)-K(+) pump α2 isoform protein in type II fibres, is likely to have importance for the exercise-induced human Na(+)-K(+) pump activity in the different fibre types.
本研究旨在研究纤维类型特异性 Na(+)-K(+)泵亚基表达和运动诱导的人类磷酸烯醇式丙酮酸（FXYD1）磷酸化改变。分离并纤维类型化人类骨骼肌纤维段，并通过 Western 印迹法确定蛋白质表达。Na(+)-K(+)泵α2 同工型的蛋白表达在 I 型纤维中低于 II 型纤维（0.63 ± 0.04 a.u. 与 1.00 ± 0.07 a.u.，P < 0.001），而 Na(+)-K(+)泵α1 和β1 同工型的蛋白表达无差异。ATP 依赖性钾通道 Kir6.2 的蛋白表达在 I 型纤维中高于 II 型纤维。在 I 型（P < 0.01）和 II 型纤维（P < 0.001）中，与休息相比，运动后 AB_FXYD1 信号降低，表明非特异性 FXYD1 磷酸化增加。与休息相比，运动后 II 型纤维中的 FXYD1 丝氨酸 68 磷酸化更高（P < 0.001）（1.90 ± 0.25 与 1.00 ± 0.08），而 I 型纤维中无变化。FXYD1 总表达没有纤维类型特异性。I 型纤维中磷酸果糖激酶的表达低于 II 型纤维（P < 0.001），而柠檬酸合酶和 3-羟酰基辅酶 A 脱氢酶在 I 型纤维中更为丰富（P < 0.001）。总之，在人类 II 型纤维中，急性剧烈运动后 FXYD1 丝氨酸 68 的磷酸化增加，而 I 型和 II 型纤维中的 AB_FXYD1 信号强度降低，表明 FXYD1 在丝氨酸 63、丝氨酸 68 和苏氨酸 69 上的磷酸化存在纤维类型特异性差异。这与 II 型纤维中 Na(+)-K(+)泵α2 同工型蛋白表达增加一起，可能对不同纤维类型中运动诱导的人类 Na(+)-K(+)泵活性具有重要意义。
