Regulation of stress-induced intracellular sorting and chaperone function of Hsp27 (HspB1) in mammalian cells
Regulation of stress-induced intracellular sorting and chaperone function of Hsp27 (HspB1) in mammalian cells
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DOI：
10.1042/bj20070195
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发表时间：
2007-11-01
影响因子：
4.1
通讯作者：
Kampinga, Harm H.
中科院分区：
文献类型：
--
作者：
Bryantsev, Anton L.;Kurchashova, Svetlana Yu.;Kampinga, Harm H.
在体外，小 Hsp（热休克蛋白）已被证明具有伴侣功能，能够保持未折叠蛋白处于适合 Hsp70 依赖性重折叠的形式。然而，这一点从未在活哺乳动物细胞中得到证实。在本研究中，我们发现 Hsp27 (HspB1) 在热激后易位到细胞核中，在细胞核中形成与 IGC（染色质间颗粒簇）共定位的颗粒。尽管热诱导的 Hsp27 寡聚状态变化与其磷酸化和核转位相关，但仅 Hsp27 磷酸化不足以实现 HspB1 的有效核转位。使用萤火虫荧光素酶作为热敏报告蛋白，我们证明 HspB1 缺陷的成纤维细胞中 HspB 1 的表达增强了热休克后的蛋白质重折叠。 HspB 1 对重折叠的积极作用被 Bag-1（Bcl-2 相关的 athanogene）（Hsp70 的负调节因子）的过表达完全减弱，这与 HspB 1 作为 Hsp70 底物持有者的想法一致。尽管热激后HspB1和荧光素酶均在核颗粒中积累，但我们的结果表明这与HspB1的重折叠活性无关。相反，颗粒积累可能反映了重新折叠失败的情况，其中基质被存储以供后续降解。一致地，我们发现热激后 20S 蛋白酶体集中在 HspB1 的核颗粒中。我们得出的结论是，HspB1 通过结合未折叠蛋白来增强细胞的伴侣能力，这些蛋白因此保持能够被 Hsp70 重折叠，或者如果重折叠失败则被分选到核颗粒。
In vitro, small Hsps (heat-shock proteins) have been shown to have chaperone function capable of keeping unfolded proteins in a form competent for Hsp70-dependent refolding. However, this has never been confirmed in living mammalian cells. In the present study, we show that Hsp27 (HspB1) translocates into the nucleus upon heat shock, where it forms granules that co-localize with IGCs (interchromatin granule clusters). Although heat-induced changes in the oligomerization status of Hsp27 correlate with its phosphorylation and nuclear translocation, Hsp27 phosphorylation alone is not sufficient for effective nuclear translocation of HspB1. Using firefly luciferase as a heat-sensitive reporter protein, we demonstrate that HspB 1 expression in HspB1-deficient fibroblasts enhances protein refolding after heat shock. The positive effect of HspB 1 on refolding is completely diminished by overexpression of Bag-1 (Bcl-2-associated athanogene), the negative regulator of Hsp70, consistent with the idea of HspB 1 being the substrate holder for Hsp70. Although HspB1 and luciferase both accumulate in nuclear granules after heat shock, our results suggest that this is not related to the refolding activity of HspB1. Rather, granular accumulation may reflect a situation of failed refolding where the substrate is stored for subsequent degradation. Consistently, we found 20S proteasomes concentrated in nuclear granules of HspB1 after heat shock. We conclude that HspB1 contributes to an increased chaperone capacity of cells by binding unfolded proteins that are hereby kept competent for refolding by Hsp70 or that are sorted to nuclear granules if such refolding fails.