Department of Microbiology and Immunology, W. R. Hearst Microbiology Research Center, Weill Medical College of Cornell University, 1300 York Avenue, New York, NY 10021, USA.
Proc Natl Acad Sci U S A. 2011 Dec 20;108(51):20370-5. doi: 10.1073/pnas.1017855108. Epub 2011 Jun 17.
The yeast telomerase regulatory protein Est3 is required for telomere maintenance in vivo, and shares intriguing structural and functional similarities with the mammalian telomeric protein TPP1. Here we report our physical and functional characterizations of Est3 homologues from Candida parapsilosis and Lodderomyces elongisporus, which bear unique N- and C-terminal tails in addition to a conserved central OB fold. We show that these Est3 homologues form stable complexes with the TEN domain of telomerase reverse transcriptase. Efficient complex formation requires both the N- and C-terminal tails, as well as conserved OB fold residues of Est3. Other Est3 homologues devoid of the tails failed to interact strongly with the cognate TEN domains. Remarkably, the C. parapsilosis Est3 alone exhibits no appreciable DNA-binding activity, but can be crosslinked to telomeric DNA in the presence of the TEN domain. A conserved basic residue on the putative DNA-binding surface of CpEst3 is required for efficient crosslinking. Mutating the equivalent residue in Candida albicans Est3 caused telomere attrition. We propose that interaction with the TEN domain unmasks a functionally important nucleic acid-binding activity in Est3. Our findings provide insights on the mechanisms and evolution of a widely conserved and functionally critical telomeric/telomerase component.
酵母端粒酶调节蛋白 Est3 是体内端粒维持所必需的，与哺乳动物端粒蛋白 TPP1 具有有趣的结构和功能相似性。在这里，我们报告了来自近平滑假丝酵母和 elongisporus 的 Est3 同源物的物理和功能特征，它们除了保守的中心 OB 折叠外，还具有独特的 N-和 C-末端尾巴。我们表明，这些 Est3 同源物与端粒酶逆转录酶的 TEN 结构域形成稳定的复合物。有效的复合物形成需要 Est3 的 N-和 C-末端尾巴以及保守的 OB 折叠残基。缺乏尾巴的其他 Est3 同源物未能与同源 TEN 结构域强烈相互作用。值得注意的是，单独的 C. parapsilosis Est3 没有表现出明显的 DNA 结合活性，但在 TEN 结构域存在的情况下可以与端粒 DNA 交联。CpEst3 假定的 DNA 结合表面上的保守碱性残基对于有效的交联是必需的。在 Candida albicans Est3 中突变等效残基会导致端粒损耗。我们提出，与 TEN 结构域的相互作用揭示了 Est3 中一种功能重要的核酸结合活性。我们的发现为广泛保守且功能关键的端粒/端粒酶成分的机制和进化提供了新的见解。
