Istituto per la Protezione Sostenibile delle Piante (CNR), Bari, Italy.
Centro di Ricerca, Sperimentazione e Formazione in Agricoltura "Basile Caramia", Locorotondo, Italy.
Virus Res. 2019 Sep;270:197644. doi: 10.1016/j.virusres.2019.197644. Epub 2019 Jun 27.
In the absence of protein-coding ability, viroid RNAs rely on direct interactions with host factors for their infectivity. RNA structural elements are likely involved in these interactions. Therefore, preservation of a structural element, despite the sequence variability existing between the variants of a viroid population, is considered a solid evidence of its relevant role in vivo. In this study, apple hammerhead viroid (AHVd) was first identified in the two apple cultivars 'Mela Rosa Guadagno' (MRG) and 'Agostinella' (AG), which are cultivated since long in Southern Italy, thus providing the first solid evidence of its presence in this country. Then, the natural variability of AHVd viroid populations infecting MRG and AG was studied. The sequence variants from the two Italian isolates shared only 82.1-87.7% sequence identity with those reported previously from other geographic areas, thus providing the possibility of exploring the impact of this sequence divergence on the proposed secondary structure. Interestingly, all the AHVd sequence variants considered in this study preserved a branched secondary structure stabilized by a kissing-loop interaction, resembling the conformation proposed previously for variants from other isolates. Indeed, most mutations did not modify the proposed conformation because they were co-variations, conversions of canonical into wobble base-pairs, or vice versa, as well as changes mapping at loops. Importantly, a cruciform structural element formed by four hairpins, one of which is implicated in the proposed kissing-loop interaction, was also preserved because several nucleotide changes actually resulted into two, three and up to five consecutive co-variations associated with other changes that did not affect the secondary structure. These data provide very strong evidence for the relevance in vivo of this cruciform structure which, together with kissing-loop interaction, likely contribute to further stabilizing the branched AHVd secondary structure.
在缺乏编码蛋白质能力的情况下，类病毒 RNA 依赖于与宿主因子的直接相互作用来实现其感染力。RNA 结构元件可能参与这些相互作用。因此，尽管类病毒群体中的变体之间存在序列变异性，但结构元件的保留被认为是其在体内相关作用的可靠证据。在这项研究中，苹果锤状病毒（AHVd）首先在两个苹果品种“Mela Rosa Guadagno”（MRG）和“Agostinella”（AG）中被鉴定出来，这两个品种在意大利南部长期种植，因此提供了该病毒在该国存在的第一个确凿证据。然后，研究了感染 MRG 和 AG 的 AHVd 类病毒群体的自然变异性。这两个意大利分离株的序列变体与以前从其他地理区域报道的序列变体仅共享 82.1-87.7%的序列同一性，从而为探索这种序列差异对提出的二级结构的影响提供了可能性。有趣的是，本研究中考虑的所有 AHVd 序列变体都保留了一个分支的二级结构，由一个亲吻环相互作用稳定，类似于以前为来自其他分离株的变体提出的构象。事实上，由于大多数突变是共变异、将规范碱基对转换为摆动碱基对，或者反之亦然，以及在环中发生的变化，因此没有改变提出的构象。重要的是，一个由四个发夹组成的十字形结构元件也被保留下来，其中一个发夹参与了提出的亲吻环相互作用，因为几个核苷酸变化实际上导致了两个、三个甚至五个连续的共变与其他不影响二级结构的变化相关联。这些数据为这种十字形结构在体内的相关性提供了非常有力的证据，该结构与亲吻环相互作用一起可能有助于进一步稳定分支的 AHVd 二级结构。
