# Impact of V-box insertion on promoter activity and virus-inducibility in transgenic Arabidopsis thaliana

## TL;DR

This study shows how inserting V-box elements into a plant promoter can enhance gene expression and virus-inducibility in Arabidopsis.

## Contribution

The novel contribution is the creation and evaluation of three modified promoters with V-box insertions for improved inducibility and activity.

## Key findings

- V-box insertion significantly altered promoter activity and virus-inducibility in transgenic Arabidopsis.
- WGFSV showed the highest response to TMV and other inducers like ABA, SA, and Eth.
- A binary function relationship was observed between V-box insertion position and transcriptional activity.

## Abstract

Synthetic pathogen-inducible promoters (SPIPs) are essential for precise gene regulation in plant genetic engineering. However, natural promoters often exhibit limitations in expression strength and specificity. In this study, we modified the WGFS promoter by incorporating V-box dimers, elements known for their virus-inducible activities, at different positions within its sequence. We thus created three new SPIPs: VWGFS, WGVFS, and WGFSV. These modified promoters were then used in transgenic Arabidopsis thaliana to evaluate their transcriptional properties through β-glucuronidase gene (GUS) staining and Quantitative real-time PCR (qPCR) analysis. Results showed that the insertion of V-box elements at different positions significantly affected the basal transcriptional activity and virus-inducibility. Notably, WGVFS and WGFSV exhibited higher basal activity than VWGFS, with WGFSV showing the highest response to tobacco mosaic virus (TMV) induction. qPCR analysis further revealed that WGFSV had enhanced inducibility by various inducers such as TMV, abscisic acid (ABA), salicylic acid (SA), and ethylene (Eth). This suggests that V-box insertion can alter promoter activity and inducibility, with the midstream position yielding the most desirable characteristics. The transcriptional activities of WGVFS in response to TMV, ABA, SA, and Eth were 1.473, 1.109, 2.030, and 1.088, respectively. Additionally, a typical binary function relationship was observed between the V-box insertion position and transcriptional expression level. The maximum model-predicted value was 1.096 when the V-box was inserted at the 99 bp downstream position. These findings help optimize promoters for plant disease resistance gene control, which may have uses beyond viral induction. This study contributes to the development of synthetic promoters with specific activity for plant genetic engineering.