G-Quadruplexes (GQs) are compact, stable structures in DNA and RNA comprised of two or more tiers of quartets whose G-rich motif of tracts of two or more G’s occurs commonly within genomes and transcriptomes. While thermodynamically stable in vitro, these structures remain difficult to study in vivo. One approach to understanding GQ in vivo behavior is to test whether conditions and molecules found in cells facilitate their folding. Polyamines are biogenic polycations that interact with RNA. Among common polyamines, spermine contains the highest charge and is found in eukaryotes, making it a good candidate for association with high-charge density nucleic acid structures like GQs. Using a variety of techniques, including ultraviolet-detected thermal denaturation, circular dichroism, size exclusion chromatography, and confocal microscopy, on an array of quadruplex sequence variants, we find that eukaryotic biological concentrations of spermine induce microaggregation of three-tiered G-rich sequences, but not of purely two-tiered structures, although higher spermine concentrations induce aggregation of even these. The formation of microaggregates can also be induced by addition of as little as a single G to a two-tiered structure; moreover, they form at biological temperatures, are sensitive to salt, and can form in the presence of at least some flanking sequence. Notably, GQ aggregation is not observed under prokaryotic-like conditions of no spermine and higher NaCl concentrations. The sequence, polyamine, and salt specificity of microaggregation reported herein have implications for the formation and stability of G-rich nucleic acid aggregates in vivo and for functional roles for understudied GQ sequences with only two quadruplex tiers.

中文翻译：

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在精胺存在的情况下，鸟嘌呤的长片段驱动 RNA G-四链体的聚集

G-四链体 (GQ) 是 DNA 和 RNA 中紧凑、稳定的结构，由两层或多层四重体组成，其富含 G 的两个或多个 G 的基序通常出现在基因组和转录组中。虽然在体外热力学稳定，但这些结构仍然难以在体内研究。一种在体内理解 GQ 的方法行为是测试细胞中发现的条件和分子是否促进它们的折叠。多胺是与 RNA 相互作用的生物聚阳离子。在常见的多胺中，精胺含有最高的电荷，并且存在于真核生物中，使其成为与 GQ 等高电荷密度核酸结构结合的良好候选者。使用多种技术，包括紫外检测热变性、圆二色性、尺寸排阻色谱和共聚焦显微镜，在一系列四链体序列变体上，我们发现精胺的真核生物浓度诱导三层富 G 序列的微聚集，但不是纯粹的两层结构，尽管更高的精胺浓度甚至会导致这些结构的聚集。微聚集体的形成也可以通过在两层结构中添加少至单个 G 来诱导；此外，它们在生物温度下形成，对盐敏感，并且可以在至少一些侧翼序列存在的情况下形成。值得注意的是，在没有精胺和较高 NaCl 浓度的类似原核生物的条件下，没有观察到 GQ 聚集。本文报道的微聚集的序列、多胺和盐特异性对富含 G 的核酸聚集体的形成和稳定性有影响体内和仅具有两个四链体层的未充分研究的 GQ 序列的功能作用。