Functional analysis of the fiber type-specific gene regulation in skeletal muscle: Modulation of NFAT function by post-translational modifications

骨骼肌的主要特征之一是它的可塑性。肌肉纤维可以根据生理需求而改变，例如耐力活动，但也可以不活动、年龄相关性萎缩或肌肉营养不良。这涉及到协调基因程序的改变，导致纤维类型的不同收缩和代谢特征。运动蛋白肌球蛋白重链(MyHC)的慢型和快型是每种纤维的主要特征。从快纤维到慢纤维的转变(快到慢的转变)是由钙调神经磷酸酶/NFATc(激活的T细胞核因子)信号通路介导的，并可以通过添加钙离子载体在培养细胞中诱导。这一更新建议研究了蛋白质1-4的另一条调控途径，包括通过翻译后修饰(PTM)在钙调神经磷酸酶下游的C1的C末端变体，并专注于通过骨骼肌中蛋白质的乙酰化或SUMO化来微调NFATc信号。在其他细胞模型中，例如T细胞的免疫反应，转录因子的乙酰化或SUMO化修饰可以诱导或抑制其靶基因的转录激活，这取决于启动子的上下文。在这里，我们将研究一种可能的调节机制，即通过蛋白质的乙酰化或SUMO化导致慢纤维中慢纤维类型基因程序的激活和抑制。这包括鉴定蛋白质1-4和C1亚型中的功能性乙酰化/SUMO化受体位点，以及以启动子依赖的方式研究修饰。初步数据表明，乙酰化和SUMO化NFATc分别与基因激活和抑制有关。此外，我们将寻找与NFATc反应元件相邻的结合位点和转录辅助因子。细胞培养的初步实验将通过分析人体模型系统中的快速到缓慢的转化来完成。目标是更好地理解骨骼肌纤维的高效调节机制，以及它们对纤维类型特定基因程序的交替激活和抑制。蛋白质的其他调控机制可以进一步阐明具有临床意义的纤维类型特异性基因调控，例如神经肌肉疾病。此外，它还可能对横纹心肌中NFATc信号通路的调节产生更广泛的影响。

One of the major characteristics of the skeletal muscle is its plasticity. Muscle fibers can be altered in response to physiological demands e.g. endurance activity, but also inactivity, age-dependent atrophy or muscle dystrophies. This involves the alteration of concerted gene programs resulting in the different contractile and metabolic features of fibre types. The slow and fast isoforms of the motor protein myosin heavy chain (MyHC) are a main characteristic of each fiber type. A shift from fast to slow fibers (fast-to-slow transformation) is mediated by the calcineurin/NFATc (nuclear factor of activated T cells)-signaling pathway and can be induced in cultured cells by addition of Ca2+-Ionophore. This renewal proposal investigates an additional regulatory pathway of proteins 1-4, including the C-terminal variants of c1, downstream of calcineurin by post-translational modifications (PTMs) and focus on the fine-tuning of NFATc signaling by acetylation or SUMOylation of proteins in skeletal muscle. In other cell models, e.g. immune response of T-cells, a modification by acetylation or SUMOylation of transcription factors can induce or repress transcriptional activation of their target genes depending on the promoter context. Here, a possible regulatory mechanism by acetylation or SUMOylation of proteins leading to the activation of the slow and inhibition of the fast fiber type gene program in slow fibers will be investigated. This includes the identification of functional acetylation/SUMOylation acceptor sites in the proteins 1-4 and c1 Isoforms and the investigation of a modification in a promoter-dependant manner. Preliminary data indicate a correlation of gene activation and repression by acetylated and SUMOylated NFATc, respectively. Furthermore, we will search for binding sites adjacent to NFATc response elements and for transcriptional cofactors. The initial experiments in cell culture will be completed by the analysis of a fast-to-slow transformation in a human model system.The objectives are a better understanding of the highly efficient regulatory mechanisms in skeletal muscle fibers with their alternating activation and inhibition of fiber type-specific gene programs. Additional regulatory mechanisms of proteins could further elucidate fiber type-specific gene regulations with clinical implications, in respect to e.g. neuromuscular diseases. In addition, it might also have a broader impact on the regulation of NFATc signaling pathway in the cross-striated cardiac muscle.