# Arabidopsis P4-ATPases ALA1 and ALA7 Enhance Resistance to Verticillium dahliae via Detoxifying Vd-Toxins

## TL;DR

Arabidopsis genes AtALA1 and AtALA7 help plants resist a harmful fungus by moving its toxins into vacuoles for detoxification.

## Contribution

The study identifies AtALA1 and AtALA7 as P4-ATPases that detoxify specific Vd-toxins via vesicle trafficking to vacuoles.

## Key findings

- AtALA1 detoxifies indazole, 4MBA, and 3ICD, while AtALA7 detoxifies CIA, 4MBA, and 3ICD.
- Overexpression of AtALA1 or AtALA7 increases Arabidopsis resistance to Vd-toxins.
- Combined expression of AtALA1 and AtALA7 further enhances resistance to Verticillium dahliae.

## Abstract

The phytopathogenic fungus Verticillium dahliae secretes diverse mycotoxins that disrupt host cellular processes, including alteration of microtubule cytoskeletons, nucleoli dysfunction, inhibition of sphingolipid synthesis, and disorder of hydrogen peroxide. However, the mechanisms underlying plant cellular detoxification of mycotoxins secreted by V. dahliae remain poorly understood. In this study, we showed that two Arabidopsis P4-ATPases, AtALA1 and AtALA7, are involved in detoxifying distinct Vd-toxins; AtALA1 is responsible for the cell detoxification of indazole, 4MBA, and 3ICD, whereas AtALA7 is essential for CIA, 4MBA, and 3ICD. Transcriptional profiling revealed significant upregulation of reactive oxygen species-associated genes in wild-type Arabidopsis in response to CIA, 3ICD, and indazole exposure. AtALA1- and AtALA7-associated vesicles compartmentalize Vd-toxins and transport them into vacuoles, thereby preventing their binding to intracellular targets and protecting host innate immunity. Notably, the expression of AtALA1 and AtALA7 simultaneously further enhances plant resistance against V. dahliae. This vesicle trafficking-mediated Vd-toxins detoxification mechanism provides a novel strategy to improve plants against various mycotoxin-producing pathogens, offering broad-spectrum applicability in agricultural biotechnology.

Background: Verticillium wilt, which is a soil-borne vascular disease, causes serious economic losses worldwide. Various toxins secreted by V. dahliae are key factors that lead to wilt symptoms. Methods: The Vd-toxins CIA, indazole, and 3ICD were labeled with fluorescence groups, respectively, to observe the transport pathway. Transcriptome sequencing and qRT-PCR were employed to assess the expression patterns under Vd-toxin treatment. Results: AtALA1 and AtALA7 were up-regulated by V. dahliae and LC-toxins, and overexpression of either AtALA1 or AtALA7 increased Arabidopsis resistance against LC-toxins. Overexpression of AtALA1 improved the resistance of Arabidopsis to 4MBA, 3ICD, and indazole, while AtALA7 enhanced resistance to 4MBA, 3ICD, and CIA. AtALA7-overexpressing plants showed a stronger capability to transport CIAFITC and 3ICD5-FAM into vacuoles, while AtALA1-overexpressing plants accumulated indazole5-FAM and 3ICD5-FAM. Aggregation of AtALA1 and AtALA7 enhances the resistance of plants to V. dahliae. Conclusions: Arabidopsis P4-ATPase genes AtALA1 and AtALA7 mediated cell detoxification by transporting different Vd-toxins to vacuoles for degradation, thereby increasing resistance to Verticillium wilt.