Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk, VA 23507, USA.
Old Dominion University, Department of Chemistry and Biochemistry, Norfolk, VA 23529, USA.
Biochim Biophys Acta Mol Cell Res. 2020 Jun;1867(6):118674. doi: 10.1016/j.bbamcr.2020.118674. Epub 2020 Feb 6.
Increased Pur-alpha (Pura) protein levels in animal models alleviate certain cellular symptoms of the disease spectrum amyotrophic lateral sclerosis/frontotemporal dementia (ALS/FTD). Pura is a member of the Pur family of evolutionarily conserved guanine-rich polynucleotide binding proteins containing a repeated signature PUR domain of 60-80 amino acids. Here we have employed a synthetic peptide, TZIP, similar to a Pur domain, but with sequence alterations based on a consensus of evolutionarily conserved Pur family binding domains and having an added transporter sequence. A major familial form of ALS/FTD, C9orf72 (C9), is due to a hexanucleotide repeat expansion (HRE) of (GGGGCC), a Pur binding element. We show by circular dichroism that RNA oligonucleotides containing this purine-rich sequence consist largely of parallel G-quadruplexes. TZIP peptide binds this repeat sequence in both DNA and RNA. It binds the RNA element, including the G-quadruplexes, with a high degree of specificity versus a random oligonucleotide. In addition, TZIP binds both linear and G-quadruplex repeat RNA to form higher order G-quadruplex secondary structures. This change in conformational form by Pur-based peptide represents a new mechanism for regulating G quadruplex secondary structure within the C9 repeat. TZIP modulation of C9 RNA structural configuration may alter interaction of the complex with other proteins. This Pur-based mechanism provides new targets for therapy, and it may help to explain Pura alleviation of certain cellular pathological aspects of ALS/FTD.
在动物模型中增加 Pur-alpha（Pura）蛋白水平可以缓解肌萎缩侧索硬化症/额颞叶痴呆（ALS/FTD）疾病谱的某些细胞症状。Pura 是 Pur 家族的成员，该家族是进化上保守的富含鸟嘌呤的多核苷酸结合蛋白，含有 60-80 个氨基酸的重复 PUR 结构域。在这里，我们使用了一种合成肽 TZIP，它类似于 Pur 结构域，但根据进化上保守的 Pur 家族结合结构域的共识进行了序列改变，并添加了转运序列。ALS/FTD 的一种主要家族形式 C9orf72（C9）是由于（GGGGCC）六核苷酸重复扩展（HRE）引起的，这是一个 Pur 结合元件。我们通过圆二色性表明，含有这种嘌呤丰富序列的 RNA 寡核苷酸主要由平行 G-四链体组成。TZIP 肽在 DNA 和 RNA 中均结合该重复序列。它与 RNA 元件结合，包括 G-四链体，具有高度特异性，而非随机寡核苷酸。此外，TZIP 结合线性和 G-四链体重复 RNA 以形成更高阶的 G-四链体二级结构。Pur 基肽的这种构象变化代表了一种调节 C9 重复内 G 四联体二级结构的新机制。TZIP 对 C9 RNA 结构构象的调节可能会改变该复合物与其他蛋白质的相互作用。这种基于 Pur 的机制为治疗提供了新的靶点，并可能有助于解释 Pura 缓解 ALS/FTD 的某些细胞病理方面。
