Nguyen Thanh Duc, Lacombe Séverine, Bangratz Martine, Ta Hoang Anh, Vinh Do Nang, Gantet Pascal, Brugidou Christophe
IRD, UMR IPME, Avenue Agropolis, 34398, Montpellier Cedex, France.
IRD, LMI RICE, Agricultural Genetics Institute, Université des Sciences et Techniques d'Hanoi, Hanoi, Vietnam.
Virus Genes. 2015 Oct;51(2):267-75. doi: 10.1007/s11262-015-1229-2. Epub 2015 Jul 28.
In Vietnam, the two main viruses that cause disease in rice are the Rice grassy stunt virus (RGSV) and the Rice ragged stunt virus (RRSV). Outbreaks of these two viruses have dramatically decreased rice production in Vietnam. Because natural resistance genes are unknown, an RNAi strategy may be an alternative method to develop resistance to RGSV and RRSV. However, this strategy will be efficient only if putative silencing suppressors encoded by the two viruses are neutralized. To identify these suppressors, we used the classical green fluorescent protein (GFP) agroinfiltration method in Nicotiana benthamiana. Then, we investigated the effects of viral candidate proteins on GFP expression and GFP siRNA accumulation and their interference with the short- or long-range signal of silencing. RGSV genes s2gp1, s5gp2, and s6gp1 and RRSV genes s5gp1, s6gp1, s9gp1, and s10gp1 were selected for viral silencing suppressor investigation according to their small molecular weight, the presence of cysteines, or the presence of a GW motif in related protein products. We confirmed that protein p6 of RRSV displays mild silencing suppressor activity and affects long-range silencing by delaying the systemic silencing signal. In addition, we identified two new silencing suppressors that displayed mild activity: p2 of RGSV and p9 of RRSV.
在越南,导致水稻发病的两种主要病毒是水稻草状矮缩病毒(RGSV)和水稻锯齿叶矮缩病毒(RRSV)。这两种病毒的爆发已使越南的水稻产量大幅下降。由于未知天然抗性基因,RNA干扰策略可能是培育对RGSV和RRSV抗性的一种替代方法。然而,只有在中和这两种病毒编码的假定沉默抑制子时,该策略才会有效。为了鉴定这些抑制子,我们在本氏烟草中使用了经典的绿色荧光蛋白(GFP)农杆菌浸润法。然后,我们研究了病毒候选蛋白对GFP表达和GFP小干扰RNA积累的影响,以及它们对沉默的短程或长程信号的干扰。根据RGSV基因s2gp1、s5gp2和s6gp1以及RRSV基因s5gp1、s6gp1、s9gp1和s10gp1的小分子质量、半胱氨酸的存在情况或相关蛋白产物中GW基序的存在情况,选择它们进行病毒沉默抑制子研究。我们证实,RRSV的p6蛋白表现出轻度的沉默抑制活性,并通过延迟系统沉默信号来影响长程沉默。此外,我们鉴定出了两种具有轻度活性的新沉默抑制子:RGSV的p2蛋白和RRSV的p9蛋白。