Ethacrynic Acid Inhibits Root Gravitropism in Arabidopsis

Ethacrynic acid (EA) is a phenoxyacetic acid derivative diuretic with promising anticancer properties, but its effects on plant growth and development remain largely unexplored. Understanding plant responses to EA, a model inhibitor and substrate of glutathione-S-transferase that depletes glutathione levels, is crucial for various applications. Here, we provide genetic evidence for the essential role of a standard glutathione pool in Arabidopsis tolerance to EA using the glutathione-deficient mutant pad2-1. Strikingly, 60 mu M EA treatment significantly inhibited root directional growth, as measured by the reduced vertical growth index (VGI) and impaired root bending in response to gravistimulation, with minimal accompanying growth inhibition. The observed agravitropic phenotype was independent of endogenous glutathione levels, as EA-treated wild-type and pad2-1 roots exhibited similar VGI. The EA-induced inhibition of root gravitropism coincided with altered auxin distribution in the root tip, resembling the effects of known gravitropism inhibitors that interfere with polar auxin transport. Accordingly, molecular docking analysis indicated that EA acts as a competitive inhibitor of PIN auxin transporters by binding to the auxin-binding pocket of PIN proteins. Additionally, EA induced the expression of mitochondrial stress and retrograde signaling markers, providing another potential link to auxin homeostasis. This study highlights the potential of EA as a novel pharmacological tool with a unique mechanism of action, providing a promising lead structure for developing novel auxin-related inhibitors. This research also underscores its potential applications in deciphering intricate auxin signaling networks.