Seasonal liver protein differences in a hibernator revealed by quantitative proteomics using whole animal isotopic labeling.
Seasonal liver protein differences in a hibernator revealed by quantitative proteomics using whole animal isotopic labeling.
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DOI：
10.1016/j.cbd.2011.02.003
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发表时间：
2011-06
影响因子：
3
通讯作者：
Martin, Sandra L.
中科院分区：
文献类型：
--
作者：
Rose, J. Cameron;Epperson, L. Elaine;Carey, Hannah V.;Martin, Sandra L.
冬眠是多种哺乳动物在冬季生存的一种节能策略。其特征是在具有低体温（Tb）的多日麻木期和短时间快速自发复温之间的周期。在休眠和复温过程中保持细胞完整性和功能的能力是冬眠的一个关键属性。来自冬季冬眠动物的肝脏能够抵抗由冷储存随后热再灌注引起的细胞损伤。识别夏季敏感和冬季保护表型状态之间的差异蛋白是一种有用的方法，可以阐明这种保护的分子机制。在这里，我们采用了一种新的定量蛋白质组学筛选策略，即一个新断奶的13线地松鼠代谢标记的重同位素取代（15 N）螺旋藻。该动物的肝脏蛋白提取物通过将其添加到夏季活动期（SA）或冬季进入期（Ent）冬眠地松鼠的合并样本提取物中，为定量评估蛋白质差异提供了通用参考。我们在两组之间鉴定了61种显著不同的蛋白质，并将其与先前使用2D凝胶在相同样品中鉴定的蛋白质进行了比较。在两个数据集共有的20种蛋白质中，18种蛋白质的差异方向和幅度完全一致，这表明两组改变的蛋白质反映了两种生理状态之间的真正差异。此外，在这项研究中回收的41种新蛋白质包括许多以前鉴定的途径中的新酶：具体地说，属于尿素循环、氨基酸和碳水化合物降解以及脂质生物合成途径的其它酶减少，而参与酮体合成、脂肪酸利用与夏季活动动物相比，来自入口冬眠动物的样品中的蛋白质合成和胚胎发生增加，为这些途径在冬眠表型中的重要性提供了额外的具体证据。
Hibernation is an energy-saving strategy used by diverse species of mammals to survive winter. It is characterized by cycles between multi-day periods of torpor with low body temperature (Tb), and short periods of rapid, spontaneous rewarming. The ability to retain cellular integrity and function throughout torpor and rewarming is a key attribute of hibernation. Livers from winter hibernators are resistant to cellular damage induced by cold storage followed by warm reperfusion. Identifying proteins that differ between the summer-sensitive and winter-protected phenotypic states is one useful approach that may elucidate the molecular mechanisms that underlie this protection. Here we employ a novel quantitative proteomics screening strategy whereby a newly-weaned 13-lined ground squirrel was metabolically labeled by ingesting heavy-isotope substituted (15N) Spirulina. The liver protein extract from this animal provided a common reference for quantitative evaluation of protein differences by its addition to extracts from pooled samples of summer active (SA) or winter entrance (Ent) phase hibernating ground squirrels. We identified 61 significantly different proteins between the two groups and compared them to proteins identified previously in the same samples using 2D gels. Of the 20 proteins common to the two datasets, the direction and magnitude of their differences were perfectly concordant for 18, providing confidence that both sets of altered proteins reflect bona fide differences between the two physiological states. Furthermore, the 41 novel proteins recovered in this study included many new enzymes in pathways identified previously: specifically, additional enzymes belonging to the urea cycle, amino acid and carbohydrate degradation, and lipid biosynthetic pathways were decreased, whereas enzymes involved in ketone body synthesis, fatty acid utilization, protein synthesis and gluconeogenesis were increased in the samples from entrance hibernators compared to summer active animals, providing additional specific evidence for the importance of these pathways in the hibernating phenotype.
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