Molecular cloning and functional analysis of Arabidopsis thaliana genes involved in bromovirus multiplication

(1)明果黄斑溴化病毒（CYBV）可以系统有效地感染模式植物拟南芥（Arabidopsis thaliana），我们最近报道了另一种溴化病毒春美潜伏溴化病毒（SBLV）。CYBV和SBLV在部分拟南芥中表现出不同的感染性。构建了CYBV基因组rna的全长cDNA克隆，从中可转录感染性体外转录物。(2)对拟南芥感染SBLV后的症状发展进行了研究。包括S96在内的几个品种对SBLV感染敏感，而包括Col-0在内的其他品种无症状或出现轻度症状。遗传研究表明，S96的症状表型受单个半显性位点控制。我们把这个位点命名为SSB1。SBLV对植物和原生质体的侵染能力在Col-0和S96之间具有可比性，表明症状表型的变化不是由于SBLV侵染能力的不同而引起的。利用遗传标记，将SSB1定位于第IV染色体上的CAT2和nga1107之间。遗传学分析表明，SSB1控制的拟南芥症状发展依赖于水杨酸信号和乙烯信号。(3)溴花叶病毒（BMV）在拟蓝藻中的感染有限，而SBLV对拟蓝藻的感染有效。在对防御反应改变的拟南芥突变体的筛选中，发现BMV通过cpr5突变系统有效地感染拟南芥，已知cpr5突变可组成性地表达系统性获得性抗性（SAR）。cpr5的双突变体和三突变体在这些途径被破坏的情况下仍然允许BMV有效地感染假单胞菌，这表明cpr5突变通过独立于赋予SAR的机制增强了BMV的增殖。即使在原生质体中，BMV的增殖能力也低于SBLV，并被cpr5突变增强了。

(1)Cassia yellow blotch bromovirus(CYBV) was shown to infect systemically and efficiently a model plant species, Arabidopsis thaliana, as we recently reported for another bromovirus, Spring beauty latent bromovirus(SBLV). CYBV and SBLV showed different infectivities in some accessions of A.thaliana. Full-length cDNA clones of CYBV genomic RNAs were constructed, from which infectious in vitro transcripts can be transcribed. (2)Symptom development was examined in A.thaliana accessions infected with SBLV. Several accessions including S96 were sensitive to SBLV infection, while the other accessions including Col-0 were symptomless or developed mild symptoms. Genetic study suggested that the symptom phenotype of S96 is controlled by a single semidominant locus. We have designated this locus SSB1. Infectivity of SBLV in plants and protoplasts were comparable between Col-0 and S96, indicating that variation of symptom phenotype was not due to the different infectivity of SBLV. By using genetic markers, SSB1 was mapped between CAT2 and nga1107 on chromosome IV. Genetic analyses suggest that symptom development in A.thaliana controlled by SSB1 is dependent on salicylic acid signaling and ethylene signaling. (3)Brome mosaic bromovirus(BMV) shows limited infection in A.thaliana, while SBLV efficiently infects it. In a screening of Arabidopsis mutants with altered defense responses, BMV was found to infect A.thaliana systemically and efficiently by the cpr5 mutations, which have been known to constitutively express systemic acquired resistance(SAR). Double and triple mutants of cpr5 with these pathways disrupted still allowed BMV to infect A.thaliana efficiently, suggesting that cpr5 mutations enhance BMV multiplication by the mechanism independent of that to confer SAR. Even in protoplasts, multiplication ability of BMV was lower than that of SBLV and was enhanced by the cpr5 mutations.