# ClpA affects the virulence of Paracidovorax citrulli on melon by regulating RepA

## TL;DR

This study shows that ClpA influences the virulence of Paracidovorax citrulli on melon plants by regulating RepA and other virulence-related genes.

## Contribution

The novel finding is that ClpA directly interacts with and negatively regulates RepA, affecting multiple virulence traits in P. citrulli.

## Key findings

- ClpA deletion reduces virulence, motility, and antioxidant capacity in P. citrulli.
- ClpA negatively regulates virB, pilR, pilA, and fliM, while positively regulating hrpG, hrcQ, and trbC.
- Deletion of RepA leads to loss of pathogenicity and altered gene expression patterns.

## Abstract

ClpA is a widely conserved protease in bacteria that plays a key role in virulence. To investigate its specific mechanism of action in the pathogenicity of Paracidovorax citrulli (formerly Acidovorax citrulli), we constructed a ClpA deletion mutant, ΔClpA. The ΔClpA mutant of P. citrulli displayed reduced virulence on melon seedlings, and reduced motility, swarming ability, and antioxidant capacity. On the other hand, the ClpA deletion of P. citrulli mutant reduced the resistance to elevated temperature and enhanced biofilm formation ability. Using qRT-PCR, we observed that ClpA negatively regulates the expression of the virulence-related genes virB, pilR, pilA, and fliM, while positively regulating hrpG, hrcQ, and trbC. Bacterial double hybrid and Glutathione-S-transferase pulldown (GST-pulldown) results showed that ClpA interacts directly with RepA, and negatively regulates the expression of RepA. After deletion of the RepA gene, the pathogenicity of P. citrulli was lost, biofilm formation ability was enhanced, and the expression of hrpG, pilR, and trbC was positively regulated. These results indicate that ClpA plays a key role in the regulation of several virulence traits of P. citrulli, paving the way for future studies to better elucidate the virulence mechanisms of this bacterial plant pathogen.