Sock E, Enderich J, Rosenfeld M G, Wegner M
Zentrum für Molekulare Neurobiologie, Universität Hamburg, Martinistrasse 52, D-20246 Hamburg, Federal Republic of Germany.
J Biol Chem. 1996 Jul 19;271(29):17512-8. doi: 10.1074/jbc.271.29.17512.
POU domain proteins are important regulators of development and terminal differentiation based upon their transcriptional activity in the nucleus. Here, we analyzed the mechanism underlying the nuclear localization of Tst-1/Oct6, a member of this family that regulates events during neurogenesis and myelination. Nuclear localization of Tst-1/Oct6 was dependent on the POU domain, as its deletion prevented access to the nucleus, whereas its transfer to the amino terminus of beta-galactosidase was sufficient to prompt nuclear accumulation of this normally cytosolic protein. Interestingly, nuclear localization and high affinity DNA binding were two independent functions of the POU domain and could be separated in several mutants. While specific high affinity binding to DNA required the presence of both the POU-specific and the POU homeodomain, the POU-specific domain was dispensable for nuclear localization of Tst-1/Oct6. Rather, the nuclear localization function was selectively contained within the POU homeodomain. Specifically, a basic cluster (GRKRKKRT) preceding helix 1 of the homeodomain was shown by deletion mutagenesis to be involved in the nuclear localization of Tst-1/Oct6. This sequence, which is highly conserved among POU domain proteins, was by itself capable of translocating beta-galactosidase to the nucleus defining it as the bona fide nuclear localization signal of Tst-1/Oct6 and presumably other POU domain factors.
POU结构域蛋白基于其在细胞核中的转录活性，是发育和终末分化的重要调节因子。在此，我们分析了Tst-1/Oct6核定位的潜在机制，Tst-1/Oct6是该家族的一员，在神经发生和髓鞘形成过程中调节相关事件。Tst-1/Oct6的核定位依赖于POU结构域，因为其缺失会阻止其进入细胞核，而将其转移至β-半乳糖苷酶的氨基末端足以促使这种通常位于胞质溶胶中的蛋白质在细胞核中积累。有趣的是，核定位和高亲和力DNA结合是POU结构域的两个独立功能，并且在几个突变体中可以分开。虽然与DNA的特异性高亲和力结合需要POU特异性结构域和POU同源结构域同时存在，但POU特异性结构域对于Tst-1/Oct6的核定位是可有可无的。相反，核定位功能选择性地包含在POU同源结构域内。具体而言，通过缺失诱变表明，同源结构域第1螺旋之前的一个碱性簇（GRKRKKRT）参与Tst-1/Oct6的核定位。该序列在POU结构域蛋白中高度保守，其自身能够将β-半乳糖苷酶转运至细胞核，将其定义为Tst-1/Oct6以及大概其他POU结构域因子的真正核定位信号。
