评估捕食性细菌噬菌弧菌 DNA 四链体形成的潜力,Biochemistry

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评估捕食性细菌噬菌弧菌 DNA 四链体形成的潜力
Biochemistry ( IF 2.9 ) Pub Date : 2022-09-15 , DOI: 10.1021/acs.biochem.2c00443
Lucille H Tsao ₁ , Sally Shepardson-Fungairiño ₁ , Hikari Murayama ₁ , Amelia Cecere ₁ , Elizabeth Wren ₁ , Megan Núñez ₁

Affiliation

在其生命周期中，捕食性细菌噬菌蛭弧菌在攻击期和生长期之间切换，每个阶段都具有独特的基因表达模式。已在蛭弧菌基因组中鉴定出 21 个潜在的 G-四链体形成序列 (PQFS)。这些富含 G 的序列在开放阅读框中普遍存在，并且几乎均匀分布在模板和编码链之间，表明它们可以在基因表达和生命周期转换中发挥作用。已发表的转录组数据表明，最接近这些序列的基因在生命周期的同一阶段不会一起（去）激活。我们使用圆二色性 (CD) 光谱和凝胶电泳探索了三个已识别的 PQFS 的生物物理特性，并证明所有三个序列都折叠成具有不同拓扑的稳定单分子四链体。在存在互补链的情况下，各自形成双链体和四链体的平衡混合物，其中在较高温度下有利于四链体形成。一旦四链体折叠起来，当添加互补链时，它们就会缓慢地形成双链体，一个序列需要相当于许多蛭弧菌寿命才能完成。使用各种共溶质，我们表明模拟细胞条件的分子拥挤可以稳定四联体结构，并诱导结构转变为平行拓扑，无论原始拓扑如何。总而言之，这些实验表明蛭弧菌PQFS 能够在体内形成四链体，从而在基因表达中发挥作用。

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Assessing the Potential for DNA Quadruplex Formation in the Predatory Bacterium Bdellovibrio bacteriovorus

During its life cycle, the predatory bacterium Bdellovibrio bacteriovorus switches between an attack and a growth phase, each of which is characterized by a distinct pattern of gene expression. Twenty-one potential G-quadruplex-forming sequences (PQFS) have been identified in the Bdellovibrio genome. These G-rich sequences are prevalent within open reading frames and nearly evenly distributed between the template and the coding strand, suggesting that they could play a role in gene expression and life cycle switching. Published transcriptomic data show that the genes nearest these sequences are not (de)activated together during the same phases of the life cycle. We explored the biophysical properties of three identified PQFS using circular dichroism (CD) spectroscopy and gel electrophoresis and demonstrated that all three sequences fold into stable unimolecular quadruplexes with distinct topologies. In the presence of their complementary strands, each forms an equilibrium mixture of duplex and quadruplex in which quadruplex formation is favored at higher temperatures. Once the quadruplexes are folded, they are slow to form a duplex when the complementary strand is added, with one sequence requiring the equivalent of many Bdellovibrio lifetimes to do so. Using a variety of cosolutes, we showed that molecular crowding mimicking cellular conditions stabilizes the quadruplex structures and induces structural transitions to the parallel topology regardless of the original topology. Taken together, these experiments suggest that Bdellovibrio PQFS are capable of forming quadruplexes in vivo and thereby playing a role in gene expression.

更新日期：2022-09-15

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