Characterization of E3Histone, a novel testis ubiquitin protein ligase which ubiquitinates histones
Characterization of E3Histone, a novel testis ubiquitin protein ligase which ubiquitinates histones
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DOI：
10.1128/mcb.25.7.2819-2831.2005
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发表时间：
2005-04-01
影响因子：
5.3
通讯作者：
Wing, SS
中科院分区：
文献类型：
--
作者：
Liu, ZQ;Oughtred, R;Wing, SS
在精子发生过程中，随着精子细胞向其细长的成熟形式进化，大部分细胞蛋白质被降解。特别是，组蛋白必须在早期伸长精子细胞中降解，以允许染色质凝聚。我们的实验室先前证明了精子发生过程中泛素结合的激活。这种激活依赖于泛素结合酶（E2）UBC 4，并且当组蛋白降解时诱导睾丸特异性同种型UBC 4-睾丸。因此，我们测试了是否有UBC 4依赖的泛素蛋白连接酶（E3），可以泛素化组蛋白。事实上，发现了一种新的酶E3（Histone），其可以在体外将泛素缀合到组蛋白H1、H2 A、H2 B、H3和H4，只有UBC 4/UBC 5家族的E2支持E3（Histone）依赖的组蛋白H2 A的泛素化，并且在该家族中，UBC 4 -1和UBC 4-testis是优选的E2。我们将这种连接酶活性纯化了3，600倍至接近同质。最终材料的质谱分析显示存在一种482 kDa的含HECT结构域的蛋白质，该蛋白质先前被命名为LASU 1。抗LASU 1抗体免疫耗竭E3（组蛋白）活性。通过凝胶过滤和甘油梯度离心的质谱和大小分析表明E3（组蛋白）是LASU 1的单体。我们的分析还表明，这种酶是主要的UBC 4 -1依赖组蛋白泛素化E3。因此，E3（组蛋白）是一个HECT结构域E3，可能在精子细胞成熟过程中发生的染色质凝聚中起重要作用。
During spermatogenesis, a large fraction of cellular proteins is degraded as the spermatids evolve to their elongated mature forms. In particular, histones must be degraded in early elongating spermatids to permit chromatin condensation. Our laboratory previously demonstrated the activation of ubiquitin conjugation during spermatogenesis. This activation is dependent on the ubiquitin-conjugating enzyme (E2) UBC4, and a testis-particular isoform, UBC4-testis, is induced when histones are degraded. Therefore, we tested whether there are UBC4-dependent ubiquitin protein ligases (E3s) that can ubiquitinate histones. Indeed, a novel enzyme, E3(Histone), which could conjugate ubiquitin to histones H1, H2A, H2B, H3, and H4 in vitro, was found. Only the UBC4/UBC5 family of E2s supported E3(Histone)-dependent ubiquitination of histone H2A, and of this family, UBC4-1 and UBC4-testis are the preferred E2s. We purified this ligase activity 3,600-fold to near homogeneity. Mass spectrometry of the final material revealed the presence of a 482-kDa HECT domain-containing protein, which was previously named LASU1. Anti-LASU1 antibodies immunodepleted E3(Histone) activity. Mass spectrometry and size analysis by gel filtration and glycerol gradient centrifugation suggested that E3(Histone) is a monomer of LASU1. Our assays also show that this enzyme is the major UBC4-1-dependent histone-ubiquitinating E3. E3(Histone) is therefore a HECT domain E3 that likely plays an important role in the chromatin condensation that occurs during spermatid maturation.