Chang Y C, Borja M, Scholthof H B, Jackson A O, Morris T J
Department of Plant Biology, University of California, Berkeley 94720, USA.
Virology. 1995 Jun 20;210(1):41-53. doi: 10.1006/viro.1995.1315.
Passage of cucumber necrosis virus (CNV) containing defective interfering (DI) RNAs through cucumber plants decreased the accumulation of DI RNAs to undetectable levels. Subsequent passages in two Nicotiana species (Nicotiana benthamiana or N. clevelandii) resulted in the appearance of DI RNA species that were larger than the DI RNAs observed during exclusive serial passages of CNV through the Nicotiana species. Sequence analysis of cloned cDNAs corresponding to the two DI RNA populations indicated that the smaller CNV-DI RNAs contained the four conserved regions (I through IV) of the genome typical of tombusvirus DI RNAs, whereas the larger DI RNAs were of similar organization but had a direct repeat of the middle portion of the molecule. This result suggests that the host has an influence on the type of DI RNA that accumulates during consecutive high multiplicity of infection passages. A comparative analysis of deletions targeting the individual conserved regions in both CNV and tomato bushy stunt virus (TBSV) DI RNAs revealed that only region III was completely dispensable for accumulation of either DI RNA species. More refined deletion analyses in regions I and II indicated that smaller segments of 75 and 35 nucleotides (nt), respectively, could be deleted without abolishing infectivity. The dispensable sequences in region II of both TBSV and CNV DI RNAs mapped to the top portion of a putative stem-loop structure. These studies indicate that both essential and nonessential sequences are conserved in DI RNAs. The essential sequences in regions I, II, and IV likely contain important cis-acting elements, whereas nonessential regions such as region III may play secondary roles such as optimally spacing cis-acting elements or maintaining the DI RNA at an overall size that is stable.
携带缺陷干扰（DI）RNA的黄瓜坏死病毒（CNV）通过黄瓜植株传代后，DI RNA的积累量降至无法检测的水平。随后在两种烟草属植物（本氏烟草或克利夫兰烟草）中传代，出现了比仅通过烟草属植物连续传代CNV时观察到的DI RNA更大的DI RNA种类。对与两个DI RNA群体相对应的克隆cDNA进行序列分析表明，较小的CNV-DI RNA包含番茄病毒DI RNA基因组典型的四个保守区域（I至IV），而较大的DI RNA具有相似的结构组织，但分子中间部分有一个直接重复序列。这一结果表明，宿主对在连续高感染复数传代过程中积累的DI RNA类型有影响。对CNV和番茄丛生矮化病毒（TBSV）DI RNA中针对各个保守区域的缺失进行比较分析发现，只有区域III对于任何一种DI RNA的积累都是完全不必要的。对区域I和II进行更精细的缺失分析表明，分别删除75和35个核苷酸（nt）的较小片段不会消除感染性。TBSV和CNV DI RNA区域II中的可缺失序列定位于一个推定的茎环结构的顶部。这些研究表明，DI RNA中既有必需序列也有非必需序列是保守的。区域I、II和IV中的必需序列可能包含重要的顺式作用元件，而非必需区域如区域III可能起次要作用，如最佳地间隔顺式作用元件或使DI RNA保持在稳定的整体大小。
