Studies Of The Molecular Interactions Between Maize Elongin C And VPg Of Sugarcane Mosaic Virus

Sugarcane mosaic virus (SCMV) is the major causal agent of maize dwarf mosaic disease in China. SCMV belongs to the genus Potyvirus. Potyviruses possess a single-stranded, positive-sense RNA genome. A viral genome-linked protein, VPg, is covalently attached to the5’terminus of the genomic RNA and has multiple roles during virus infection. VPg is involved in viral RNA replication and translation, and it also influences potyvirus movement. We identified a maize Elongin C (ZmElc) as an interactor with SCMV VPg though a yeast two-hybrid (Y2H) screen by using SCMV-BJ VPg as the bait. The interaction between SCMV-BJ VPg and ZmElc was studied and the role of ZmElc during SCMV infection was explored preliminarily.The interaction between SCMV-BJ VPg and ZmElc was further studied through a bimolecular fluorescence complementation (BiFC) assay in maize ptotoplasts. The interaction specificity between VPg and Elonglin C was investigated and the results showed that the VPg-Elongin C interaction was not potyvirus-and host-specific.The tissue-specific expression pattern of ZmELC and subcellular localization of ZmElc were studied. We found that ZmELC was expressed in all the tissues of maize seedlings and flowering maize plants. The highest ZmELC transcript level was in leaf blade in maize seedlings and in pistil in the flowering maize plants. In maize cells, ZmElc located in both cytoplasm and nucleus. These findings provide the first evidence of Elongin C tissue and subcellular localization in plants. We also determined that the subcellular localization of ZmElc in maize protoplasts was unchanged after SCMV infection.To study the role of ZmElc during SCMV infection, the influence of virus infection on ZmELC transcript level was determined by Real-Time quantitative RT-PCR. The result showed that the expression of ZmELC was up-regulated in inoculated and first-systemically infected leaves in virus-inoculated plants compared with equivalent leaves from the mock-inoculated plants at4and6dpi. This difference disappeared in the first-systemically infected leaves at10dpi.The accumulation of SCMV RNA was increased when ZmELC was transiently over-expressed in maize protoplasts, which suggested that ZmElc facilitated SCMV RNA replication. To determine further the function of ZmElc during SCMV infection, the inoculation method for Brome mosaic virus (BMV)-mediated gene silencing (VIGS) in maize was modified and ZmELC was transiently knocked down using this modified BMV VIGS method. The result showed that knockdown of ZmELC in maize plants did not influence maize plant growth and development, but did decrease RNA accumulation of two isolates of SCMV.To explore the pathway ZmElc may be involved in during SCMV infection, the correlation between ZmElc and ZmeIF4E was investigated. It was determined that SCMV VPg interacted with ZmeIF4E and ZmElc did not interact with ZmeIF4E by the Y2H and BiFC assay in maize protoplasts. Knockdown of ZmELC expression resulted in less ZmeIF4E expression only in the presence of SCMV. This result revealed that ZmElc and ZmeIF4E were related during SCMV infection.