Comparison of intra-organellar chaperone capacity for dealing with stress-induced protein unfolding
Comparison of intra-organellar chaperone capacity for dealing with stress-induced protein unfolding
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DOI：
10.1074/jbc.m703876200
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发表时间：
2007-11-23
影响因子：
4.8
通讯作者：
Kampinga, Harm H.
中科院分区：
文献类型：
--
作者：
Hageman, Jurre;Vos, Michel J.;Kampinga, Harm H.
分子伴侣对于细胞防止部分未折叠的蛋白质形成无功能的有毒聚集体是必不可少的。当细胞经历蛋白质解折叠应激时，这种需求增加，并且这可能影响真核细胞中的所有隔室。是否所有的细胞器都配备了可比的伴侣能力在很大程度上是未知的，主要是由于缺乏合适的记者，允许这样的比较。在这里，我们描述了荧光荧光素酶报告的发展，被分类到各种细胞位置（细胞核，细胞质，内质网，和过氧化物酶体），并在其内在的热稳定性的差异最小。当加热活细胞时，核靶向荧光素酶的失活速率最快，表明细胞核是对热诱导的变性应激最敏感的细胞器。然而，热后再活化，发生在相同的动力学，无论荧光素酶的本地化。此外，诱导耐热性的引发热处理，协调上调所有的热诱导分子伴侣，导致在一个短暂的耐热性的荧光素酶在所有的细胞器中以可比的方式。过表达的主要热诱导Hsp 70家族成员，HspA 1A，保护只有胞质和核，但没有其他酶。总之，我们的数据表明，在每个隔室的调查，包括过氧化物酶体中，到目前为止没有伴侣可以检测到，伴侣机器存在，并可以诱导与那些存在于胞质/核隔室的活动类似。
Molecular chaperones are essential for cells to prevent that partially unfolded proteins form non-functional, toxic aggregates. This requirement is increased when cells experience protein unfolding stresses and such could affect all compartments in the eukaryotic cell. Whether all organelles are equipped with comparable chaperone capacities is largely unknown, mainly due to the lack of suitable reporters that allow such a comparison. Here we describe the development of fluorescent luciferase reporters that are sorted to various cellular locations (nucleus, cytoplasm, endoplasmic reticulum, and peroxisomes) and that differ minimally in their intrinsic thermal stability properties. When heating living cells, the rate of inactivation was most rapid for the nuclear-targeted luciferase, indicating that the nucleus is the most sensitive organelle toward heat-induced denaturing stress. Post-heat re-activation, however, occurred at equal kinetics irrespective of luciferase localization. Also, induction of thermotolerance by a priming heat treatment, that coordinately up-regulates all heat-inducible chaperones, resulted in a transient heat resistance of the luciferase in all organelles in a comparable manner. Overexpression of the main heat-inducible Hsp70 family member, HspA1A, protected only the cytosolic and nuclear, but not the other luciferases. Together, our data suggest that in each compartment investigated, including the peroxisome in which so far no chaperones could be detected, chaperone machines are present and can be induced with activities similar to those present in the cytosolic/nuclear compartment.