The Hsp70 co-chaperone Ydj1/HDJ2 regulates ribonucleotide reductase activity.
The Hsp70 co-chaperone Ydj1/HDJ2 regulates ribonucleotide reductase activity.
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DOI：
10.1371/journal.pgen.1007462
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发表时间：
2018-11
期刊：
影响因子：
4.5
通讯作者：
Truman AW
中科院分区：
文献类型：
--
作者：
Sluder IT;Nitika;Knighton LE;Truman AW
Hsp70是一种保守的分子伴侣，参与许多“客户”蛋白的折叠、稳定和最终降解。Hsp70受一系列协同伴侣分子调控，这些分子协助Hsp70-client相互作用并刺激Hsp70的内在atp酶活性。虽然先前的研究表明抗癌靶标核糖核苷酸还原酶（RNR）是Hsp70的客户端，但所涉及的调节共伴侣仍有待确定。为了鉴定参与RNR活性的共伴侣蛋白，筛选了28个酵母共伴侣蛋白敲除突变体对RNR干扰剂羟基脲的敏感性。Ydj1是一种重要的细胞质Hsp70共伴侣，被鉴定为在HU上生长所必需的。Ydj1结合RNR亚基Rnr2，缺乏Ydj1的细胞显示出不稳定的RNR复合物。HDJ2结合R2B并调节人类细胞中RNR的稳定性，表明从酵母到人类都具有广泛的保守性。通过突变或通过小分子116-9e干扰Ssa1-Ydj1相互作用或Hsp70-HDJ2破坏RNR功能，表明这种新作用依赖于伴侣。缺乏HDJ2的哺乳动物细胞对羟脲、吉西他滨和曲平等抑制RNR的药物更为敏感。综上所述，这项工作提出了一种新的抗癌策略-通过靶向Hsp70共伴侣功能抑制RNR。核糖核苷酸还原酶（RNR）是DNA合成的关键酶，抑制RNR可导致细胞对辐射的敏感性。因此，RNR是包括癌症在内的多种疾病的有效治疗靶点。抗rnr药物是有效的，但与患者的一系列副作用有关。我们之前的工作已经确定了Hsp90和Hsp70分子伴侣蛋白调控RNR。Hsp70和Hsp90的特异性和活性受“co-chaperone”蛋白调控。我们在缺乏单个共伴侣的细胞中检测了RNR活性，发现Ydj1/HDJ2蛋白是酵母和人类细胞中RNR的一种新的调节因子。重要的是，我们证明抑制HDJ2使细胞对目前使用的抗癌药物敏感。
Hsp70 is a well-conserved molecular chaperone involved in the folding, stabilization, and eventual degradation of many “client” proteins. Hsp70 is regulated by a suite of co-chaperone molecules that assist in Hsp70-client interaction and stimulate the intrinsic ATPase activity of Hsp70. While previous studies have shown the anticancer target ribonucleotide reductase (RNR) is a client of Hsp70, the regulatory co-chaperones involved remain to be determined. To identify co-chaperone(s) involved in RNR activity, 28 yeast co-chaperone knockout mutants were screened for sensitivity to the RNR-perturbing agent Hydroxyurea. Ydj1, an important cytoplasmic Hsp70 co-chaperone was identified to be required for growth on HU. Ydj1 bound the RNR subunit Rnr2 and cells lacking Ydj1 showed a destabilized RNR complex. Suggesting broad conservation from yeast to human, HDJ2 binds R2B and regulates RNR stability in human cells. Perturbation of the Ssa1-Ydj1 interaction through mutation or Hsp70-HDJ2 via the small molecule 116-9e compromised RNR function, suggesting chaperone dependence of this novel role. Mammalian cells lacking HDJ2 were significantly more sensitive to RNR inhibiting drugs such as hydroxyurea, gemcitabine and triapine. Taken together, this work suggests a novel anticancer strategy-inhibition of RNR by targeting Hsp70 co-chaperone function. Ribonucleotide reductase (RNR) is a key enzyme in the synthesis of DNA and inhibition of RNR leads to cellular sensitivity to radiation. As such, RNR is a well-validated therapeutic target for a variety of diseases including cancer. Anti-RNR drugs are effective but are associated with a range of side effects in patients. Our previous work had identified that the Hsp90 and Hsp70 molecular chaperone proteins regulate RNR. The specificity and activity of Hsp70 and Hsp90 are regulated by “co-chaperone” proteins. We examined RNR activity in cells lacking individual co-chaperones and identified the Ydj1/HDJ2 protein as a novel regulator of RNR in yeast and human cells. Importantly, we demonstrate that inhibiting HDJ2 sensitizes cells to currently used anticancer drugs.