NbNAC42 and NbZFP3 Transcription Factors Regulate the Virus Inducible NbAGO5 Promoter in Nicotiana benthamiana

Abstract

Plant argonautes (AGOs) play important roles in the defense responses against viruses. The expression of Nicotiana benthamiana AGO5 gene (NbAGO5) is highly induced by Bamboo mosaic virus (BaMV) infection; however, the underlying mechanisms remain elusive. In this study, we have analyzed the potential promoter activities of NbAGO5 and its interactions with viral proteins by using a 2,000 bp fragment, designated as PN1, upstream to the translation initiation of NbAGO5. PN1 and seven serial 5′-deletion mutants (PN2–PN8) were fused with a β-glucuronidase (GUS) reporter and introduced into the N. benthamiana genome by Agrobacterium-mediated transformation for further characterization. It was found that PN4-GUS transgenic plants were able to drive strong GUS expression in the whole plant. In the virus infection tests, the GUS activity was strongly induced in PN4-GUS transgenic plants after being challenged with potexviruses. Infiltration of the transgenic plants individually with BaMV coat protein (CP) or triple gene block protein 1 (TGBp1) revealed that only TGBp1 was crucial for inducing the NbAGO5 promoter. To identify the factors responsible for controlling the activity of the NbAGO5 promoter, we employed yeast one-hybrid screening on a transcription factor cDNA library. The result showed that NbNAC42 and NbZFP3 could directly bind the 704 bp promoter regions of NbAGO5. By using overexpressing and virus-induced gene silencing techniques, we found that NbNAC42 and NbZFP3 regulated and downregulated, respectively, the expression of the NbAGO5 gene. Upon virus infection, NbNAC42 played an important role in regulating the expression of NbAGO5. Together, these results provide new insights into the modulation of the defense mechanism of N. benthamiana against viruses. This virus inducible promoter could be an ideal candidate to drive the target gene expression that could improve the anti-virus abilities of crops in the future.

E2F7 drives autotaxin/Enpp2 transcription via chromosome looping: Repression by p53 in murine but not in human carcinomas

“…Y1H screen was performed as described previously 38,39 . The coding region of suspected TFs (prey proteins) was cloned to pGADT7‐AD vector (Takara).…”

“…Y1H screen was performed as described previously. 38,39 The coding region of suspected TFs (prey proteins) was cloned to pGADT7-AD vector (Takara). Tandemly repeated ATX promoter regions (bait sequences) were amplified and cloned into the pABAi vector (Takara) with Aureobasidin A (AbA) resistance reporter gene (AbA r ).…”

The Role of Plant Transcription Factors in the Fight against Plant Viruses

“…The plant hormone ABA, the production of which has been demonstrated to be stimulated by BaMV infection, also contributes to the stimulation of NbAGO5 promoter activity. Triple gene block protein 1 (TGBp1), which is encoded by BaMV, may regulate NbAGO5 expression and plant defense response by activating the NbNAC42 expression [51].…”

“…Transcriptomic data highlighted the significant upregulation of 88 NAC transcripts in the sugarcane mosaic virus (SCMV)-resistant (B-48) genotype of sugarcane compared to those of the susceptible (Badila) genotype [67]. Recently, we identified NbNAC42, an activator of argonaute 5 (NbAGO5) from N. benthamiana [51], which was significantly upregulated under bamboo mosaic virus (BaMV) infection, inducing the activity level of NbAGO5, and leading to antiviral defense (Figure 2C). Overall, the findings suggest that NAC TFs play a crucial role in regulating viral infection, multiplication, and accumulation by interacting with viral proteins or upregulating pathogen-or defense-related genes during viral infection.…”

The phosphorylation of the movement protein TGBp1 regulates the accumulation of the Bamboo mosaic virus

“…TGBp1 is known to form an inclusion body within BaMV-infected cells, acting as a reservoir for active TGBp1 to bind to viral RNA and CP, forming a ribonucleoprotein complex [29, 31]. In addition, BaMV TGBp1 may induce NbAGO5 expression and trigger downstream defence against BaMV in N. benthamiana [32]. Although the role of TGBp1 phosphorylation in regulating virus movement is critical, the potential sites and their effects on BaMV infection remain unclear.…”