Use of virus‐induced gene silencing to characterize genes involved in modulating hypersensitive cell death in maize,Molecular Plant Pathology

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Use of virus‐induced gene silencing to characterize genes involved in modulating hypersensitive cell death in maize
Molecular Plant Pathology ( IF 4.8 ) Pub Date : 2020-10-10 , DOI: 10.1111/mpp.12999
Colin Murphree ₁ , Saet-Byul Kim ₁ , Shailesh Karre ₁ , Rozalynne Samira _{1,

2} , Peter Balint-Kurti _{1,

3}

Affiliation

Plant disease resistance proteins (R‐proteins) detect specific pathogen‐derived molecules, triggering a defence response often including a rapid localized cell death at the point of pathogen penetration called the hypersensitive response (HR). The maize Rp1‐D21 gene encodes a protein that triggers a spontaneous HR causing spots on leaves in the absence of any pathogen. Previously, we used fine mapping and functional analysis in a Nicotiana benthamiana transient expression system to identify and characterize a number of genes associated with variation in Rp1‐D21‐induced HR. Here we describe a system for characterizing genes mediating HR, using virus‐induced gene silencing (VIGS) in a maize line carrying Rp1‐D21. We assess the roles of 12 candidate genes. Three of these genes, SGT1, RAR1, and HSP90, are required for HR induced by a number of R‐proteins across several plant–pathogen systems. We confirmed that maize HSP90 was required for full Rp1‐D21‐induced HR. However, suppression of SGT1 expression unexpectedly increased the severity of Rp1‐D21‐induced HR while suppression of RAR1 expression had no measurable effect. We confirmed the effects on HR of two genes we had previously validated in the N. benthamiana system, hydroxycinnamoyltransferase and caffeoyl CoA O‐methyltransferase. We further showed the suppression the expression of two previously uncharacterized, candidate genes, IQ calmodulin binding protein (IQM3) and vacuolar protein sorting protein 37, suppressed Rp1‐D21‐induced HR. This approach is an efficient way to characterize the roles of genes modulating the hypersensitive defence response and other dominant lesion phenotypes in maize.

中文翻译：

利用病毒诱导的基因沉默来表征参与调节玉米过敏性细胞死亡的基因

植物抗病蛋白（R-蛋白）检测特定的病原体衍生分子，触发防御反应，通常包括病原体渗透时的快速局部细胞死亡，称为过敏反应（HR）。玉米Rp1-D21基因编码一种蛋白质，该蛋白质会触发自发 HR，在没有任何病原体的情况下在叶子上产生斑点。此前，我们在烟草瞬时表达系统中使用精细作图和功能分析来识别和表征与Rp1-D21诱导的 HR 变异相关的许多基因。在这里，我们描述了一种在携带Rp1-D21的玉米品系中使用病毒诱导基因沉默 (VIGS) 来表征介导 HR 的基因的系统。我们评估了 12 个候选基因的作用。其中三个基因， SGT1 、 RAR1和HSP90 ，是多个植物病原体系统中许多 R 蛋白诱导的 HR 所必需的。我们证实，完整的Rp1-D21诱导 HR 需要玉米 HSP90。然而，SGT1 表达的抑制意外地增加了Rp1-D21诱导的 HR 的严重程度，而 RAR1 表达的抑制则没有可测量的效果。我们确认了之前在本塞姆氏烟草系统中验证过的两个基因（羟基肉桂酰转移酶和咖啡酰辅酶 A O-甲基转移酶）对 HR 的影响。我们进一步表明，两个先前未表征的候选基因IQ 钙调蛋白结合蛋白( IQM3 ) 和液泡蛋白分选蛋白 37的表达受到抑制，从而抑制了Rp1-D21诱导的 HR。这种方法是表征调节玉米超敏防御反应和其他显性病变表型的基因作用的有效方法。

更新日期：2020-11-27

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