Structure/ function analyses of sarco(endo)plasmic reticulum CA2+ - ATPases : effects of oxidative stress , redox signaling and protein-protein interactions

我们身体中的肌肉细胞经常暴露于各种应激条件下，如热应激或剧烈运动或与各种疾病有关。当肌肉细胞暴露于压力时，细胞内的钙水平增加，并且据信细胞损伤甚至细胞死亡将由于细胞钙的增加而发生。压力下钙增加的一个主要原因是负责维持肌肉中正常钙水平的酶，即Ca2+泵，在压力下受损。因此，可以预期，如果钙泵能够以某种方式得到保护，那么应激后细胞损伤将减少，存活率将增加。我们最近发现，一种名为热休克蛋白70（Hsp70）的蛋白质可以在热应激期间保护肌肉中的钙泵，但我们还没有弄清楚这是如何工作的，或者肌肉中的其他蛋白质是否也可以保护钙泵。该项目的目标是专门研究Hsp70和另外两种称为受磷蛋白和肌磷脂的蛋白质如何在广泛的压力条件下保护Ca2+泵。我们将使用重组DNA技术操纵活细胞中Ca2+泵、Hsp70、受磷蛋白和肌磷脂的水平，然后将细胞暴露于各种类型的应激（即热、活性氧等）。然后评估对Ca2+泵的结构和功能损害。我们研究的另一个目标是了解是什么控制了在休息和运动条件下通常在肌肉中发现的Hsp70的数量。我们将招募年轻（<30岁）和老年（>70岁）男性和女性进行实验，以研究人类骨骼肌中Hsp70对运动反应的潜在性别和年龄差异。这项工作可能具有重要的意义，可以扩展到许多领域（即癌症和心脏病研究），因为保护钙泵可以防止钙的增加，并提高细胞在压力下的存活率。

Muscle cells in our body are often exposed to a variety of stressful conditions such as heat stress or intense exercise or in association with a variety of diseases. When muscle cells are exposed to stress the calcium level inside the cells increases and it is believed that cell damage and even cell death will occur as a result of the increased cellular calcium. One major reason for the increase in calcium with stress is that the enzyme responsible for maintaining normal calcium levels in muscle, namely the Ca2+ pump, becomes damaged with stress. Therefore, it would be expected that cell damage would be reduced and survival would be increased after stress if the calcium pump could somehow be protected. We have recently shown that a protein called heat shock protein 70 (Hsp70) can protect the calcium pump in muscle during heat stress but we haven't figured out exactly how this works yet or whether other proteins in muscle might also protect the calcium pump. The goal of this project is to examine specifically how Hsp70 and two other proteins called phospholamban and sarcolipin, protect the Ca2+ pump under a wide range of stress conditions. We will manipulate the levels of the Ca2+ pump, Hsp70, phospholamban and sarcolipin in living cells using recombinant DNA technology and then expose the cells to various types of stress (i.e. heat, reactive oxygen species, etc.) followed by assessment of structural and functional damage to the Ca2+ pump. Another goal of our research is to understand what controls the amount of Hsp70 that is normally found in muscle under resting and exercise conditions. We will recruit young (<30 years) and old (>70 years) men and women for our experiments to investigate potential sex and age differences in the Hsp70 response to exercise in human skeletal muscle. This work could have important implications that extend into a number of fields (i.e. cancer and heart disease research) because protecting the Ca2+ pump could prevent increases in calcium and improve cell survival following stress.