Absence of γ-sarcoglycan alters the response of p70S6 kinase to mechanical perturbation in murine skeletal muscle.
Absence of γ-sarcoglycan alters the response of p70S6 kinase to mechanical perturbation in murine skeletal muscle.
复制标题
DOI：
10.1186/2044-5040-4-13
复制
发表时间：
2014
期刊：
影响因子：
4.9
通讯作者：
Barton ER
中科院分区：
文献类型：
--
作者：
Moorwood C;Philippou A;Spinazzola J;Keyser B;Macarak EJ;Barton ER
肌营养不良蛋白糖蛋白复合物（DGC）位于肌纤维的肌膜处，提供结构完整性。DGC蛋白的突变和缺失导致一系列肌营养不良症。当仅肌聚糖亚复合物不存在时，肌肉显示严重的肌纤维变性，但对收缩损伤几乎不敏感，这表明疾病不是通过结构缺陷而是通过异常信号传导发生的，即通过肌聚糖复合物的正常机械转导信号传导的丧失。我们扩展了我们以前对机械敏感性、γ-肌聚糖依赖性ERK 1/2磷酸化的研究，以确定γ-肌聚糖的丢失是否会改变其他途径。我们使用C2 C12肌管、原代培养物以及来自C57 Bl/6（C57）和γ-肌聚糖缺失（γ-SG-/-）小鼠的分离肌肉检查了在存在和不存在γ-肌聚糖的情况下的机械转导。所有人都受到周期性被动拉伸。信号蛋白磷酸化通过来自拉伸和非拉伸样品的裂解物的免疫印迹来测定。通过将肌肉保持在无钙或补充丁卡因的林格氏溶液中来评估钙依赖性。通过在存在或不存在雷帕霉素的情况下拉伸分离的肌肉来确定对mTOR的依赖性。C2 C12肌管牵拉引起P-p70 S6 K的强烈增加，但P-FAK和P-ERK 2降低。Akt和ERK 1对被动牵张均无反应。在C57原代培养物中观察到响应拉伸的相似但不显著的趋势，并且γ-SG-/-培养物未显示p70 S6 K响应。相反，在离体肌肉中，p70 S6 K是机械反应性的。在γ-SG-/-小鼠的肌肉中，基础p70 S6 K活化以钙非依赖性方式升高。在C57和γ-SG-/-分离肌肉中，p70 S6 K激活随着牵拉而增加，并且在γ-SG-/-肌肉中持续存在，这与C57肌肉中的瞬时反应不同。雷帕霉素处理阻断了拉伸的C57肌肉中的所有p70 S6 K活化，并减少下游S6 RP磷酸化。然而，即使雷帕霉素处理降低了拉伸的γ-SG-/-肌肉中的p70 S6 K活化，S6 RP磷酸化仍然升高。p70 S6 K是骨骼肌中γ-肌聚糖依赖性力学信号转导的重要组成部分。我们的研究结果表明，γ-肌聚糖的损失解偶联p70 S6 K的反应，拉伸，并暗示，γ-肌聚糖是重要的失活这一途径。总的来说，我们断言，改变负荷感应机制存在于肌营养不良症的肌聚糖是缺席。
The dystrophin glycoprotein complex (DGC) is located at the sarcolemma of muscle fibers, providing structural integrity. Mutations in and loss of DGC proteins cause a spectrum of muscular dystrophies. When only the sarcoglycan subcomplex is absent, muscles display severe myofiber degeneration, but little susceptibility to contractile damage, suggesting that disease occurs not by structural deficits but through aberrant signaling, namely, loss of normal mechanotransduction signaling through the sarcoglycan complex. We extended our previous studies on mechanosensitive, γ-sarcoglycan-dependent ERK1/2 phosphorylation, to determine whether additional pathways are altered with the loss of γ-sarcoglycan. We examined mechanotransduction in the presence and absence of γ-sarcoglycan, using C2C12 myotubes, and primary cultures and isolated muscles from C57Bl/6 (C57) and γ-sarcoglycan-null (γ-SG-/-) mice. All were subjected to cyclic passive stretch. Signaling protein phosphorylation was determined by immunoblotting of lysates from stretched and non-stretched samples. Calcium dependence was assessed by maintaining muscles in calcium-free or tetracaine-supplemented Ringer’s solution. Dependence on mTOR was determined by stretching isolated muscles in the presence or absence of rapamycin. C2C12 myotube stretch caused a robust increase in P-p70S6K, but decreased P-FAK and P-ERK2. Neither Akt nor ERK1 were responsive to passive stretch. Similar but non-significant trends were observed in C57 primary cultures in response to stretch, and γ-SG-/- cultures displayed no p70S6K response. In contrast, in isolated muscles, p70S6K was mechanically responsive. Basal p70S6K activation was elevated in muscles of γ-SG-/- mice, in a calcium-independent manner. p70S6K activation increased with stretch in both C57 and γ-SG-/- isolated muscles, and was sustained in γ-SG-/- muscles, unlike the transient response in C57 muscles. Rapamycin treatment blocked all of p70S6K activation in stretched C57 muscles, and reduced downstream S6RP phosphorylation. However, even though rapamycin treatment decreased p70S6K activation in stretched γ-SG-/- muscles, S6RP phosphorylation remained elevated. p70S6K is an important component of γ-sarcoglycan-dependent mechanotransduction in skeletal muscle. Our results suggest that loss of γ-sarcoglycan uncouples the response of p70S6K to stretch and implies that γ-sarcoglycan is important for inactivation of this pathway. Overall, we assert that altered load-sensing mechanisms exist in muscular dystrophies where the sarcoglycans are absent.
登录
查看更多内容
DOI：
10.1083/jcb.142.5.1279
发表时间：
1998-09-07
期刊：
The Journal of cell biology
影响因子：
--
作者：
Hack AA;Ly CT;Jiang F;Clendenin CJ;Sigrist KS;Wollmann RL;McNally EM
通讯作者：
McNally EM
DOI：
10.1073/pnas.96.19.10723
发表时间：
1999-09-14
影响因子：
11.1
作者：
Hack, AA;Cordier, L;McNally, EM
通讯作者：
McNally, EM
影响因子：
5.5
作者：
Cho, Du-Hyong;Choi, Yoon Jung;Jo, Inho
通讯作者：
Jo, Inho
影响因子：
4.8
作者：
Carriere, Audrey;Romeo, Yves;Roux, Philippe P.
通讯作者：
Roux, Philippe P.
影响因子：
4.8
作者：
Axelrod MJ;Gordon V;Mendez RE;Leimgruber SS;Conaway MR;Sharlow ER;Jameson MJ;Gioeli DG;Weber MJ
通讯作者：
Weber MJ