Unveiling a Virulence-Regulating Mechanism in Aeromonas hydrophila: a Quantitative Exoproteomic Analysis of an AraC-Like Protein

Abstract

Bacterial AraC is a transcription factor family that initiates transcription by recruiting RNA polymerase to the promoter and directly regulating various bacterial phenotypes. It also directly regulates various bacterial phenotypes. However, how this transcription factor regulates bacterial virulence and affects host immunity is still largely unknown. In this study, deleting the orf02889 (AraC-like transcription factor) gene in virulent Aeromonas hydrophila LP-2 affected several important phenotypes, such as increasing biofilm formation and siderophore production abilities. Moreover, Δorf02889 also significantly decreased the virulence of A. hydrophila and has promising attenuated vaccine potential. To better understand the effects of orf02889 on biological functions, a data independent acquisition (DIA)-based quantitative proteomics method was performed to compare the differentially expressed proteins between Δorf02889 and the wild-type strain in extracellular fractions. The following bioinformatics analysis suggested that ORF02889 may regulate various metabolic pathways, such as quorum sensing and ATP binding cassette (ABC) transporter metabolism. Moreover, 10 selected genes from the top 10 decreasing abundances in proteomics data were deleted, and their virulence to zebrafish was evaluated, respectively. The results showed that ΔcorC, Δorf00906, and Δorf04042 significantly reduced bacterial virulence. Finally, the following chromatin immunoprecipitation and polymerase chain reaction (ChIP-PCR) assay validated that the promoter of corC was directly regulated by ORF02889. Overall, these results provide insight into the biological function of ORF02889 and demonstrate its inherent regulatory mechanism for the virulence of A. hydrophila.

Quantitative Proteomics Analysis Reveals the Effect of a MarR Family Transcriptional Regulator AHA_2124 on Aeromonas hydrophila

“…Furthermore, it was reported that yeeY, a LysR-type transcriptional regulator (LTTR), negatively regulated furazolidone resistance according to the proteomics analysis [45]. Zhang et al reported that ORF02889, encoded by orf02889 (an AraC-like transcription factor), was involved in the regulatory mechanism for the virulence of Aeromonas hydrophila [46]. Thus, these transcriptional regulators described previously were all affected by the deletion of AHA_2124 in this study, indicating the important role of AHA_2124 in some physiological processes of Aeromonas hydrophila.…”

Unveiling the antibacterial mechanism of resveratrol against Aeromonas hydrophila through proteomics analysis

“…50μg of the protein sample was obtained for enzymatic hydrolysis using filter-aided sample preparation (FASP) method ( Wiśniewski, 2017 ). The protein was subjected to reduction and alkylation using dithiothreitol (DTT) and iodoacetamide (IAA), as previously described ( Zhang et al., 2023 ). The protein was digested with trypsin (the volume ratio of trypsin to sample was 1:50) for 16 h at 37°C.…”