Use of virus-induced gene silencing to characterize genes involved in modulating hypersensitive cell death in maize

Authors: MURPHREE, COLIN - North Carolina State University; KIM, SAET-BYUL - North Carolina State University; KARRE, SHAILESH - North Carolina State University; SAMIRA, ROZALYNNE - North Carolina State University; Balint-Kurti, Peter

Submitted to: Molecular Plant Pathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/1/2020
Publication Date: 8/1/2020
Citation: Murphree, C., Kim, S., Karre, S., Samira, R., Balint Kurti, P.J. 2020. Use of virus-induced gene silencing to characterize genes involved in modulating hypersensitive cell death in maize. Molecular Plant Pathology. 21:1662–1676.

Interpretive Summary: We used an engineered a plant virus to suppress the expression of several genes in maize and to assess their role in mediating the plant defense response. We identified three genes which increased the defense response when their expression was suppress and three others which had the opposite effect.

Technical Abstract: Plant disease resistance proteins (R-proteins) detect specific pathogen-derived molecules, triggering a defense 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 identified a number of candidate genes associated with variation in Rp1-D21-induced HR and validated the roles of several of them using a Nicotiana benthamiana transient expression system. However, functional analysis of these genes in maize itself was impractical due to the limitations of the maize experimental system. We have used a recently-developed system for virus-induced gene silencing (VIGS) in maize to assess the roles of twelve candidate genes in modulating Rp1-D21-induced HR. 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 gene 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 defense response.