Original data

Aerobic respiration is a key driver of the Vibrio cholerae proliferation and infection process. Our previous transcriptome results suggest that degS knockout downregulates some genes involved in NADH and ATP synthesis in the aerobic respiratory pathway. In this study, nontargeted metabolomics results showed that the differential metabolites affected by degS knockout were associated with aerobic respiration. Further results suggested that the key products of aerobic respiration, NADH and ATP, were reduced upon degS deletion and were not dependent on the classical σE pathway. It has been shown that the two-component system response factor ArcA is involved in regulating NADH and ATP levels. By qRT-PCR, we found that DegS negatively regulates the transcription of the arcA gene, which also negatively regulates the expression of isocitrate dehydrogenase (ICDH), a key rate-limiting enzyme of the tricarboxylic acid cycle. Knockout of the arcA gene in theΔdegS strain partially restored NADH and ATP levels; whereas overexpression of ICDH in the ΔdegS strain also partially restored NADH and ATP levels. In the growth experiment, compared to the ΔdegS strain, the growth rates of ΔdegSΔarcA and ΔdegS-overexpressed icdh strains(ΔdegS+icdh) were partially restored during the logarithmic growth period. Intestinal colonization of suckling mice showed a significant reduction in the colonization ability of the ΔdegS strain, a comparable colonization ability of the ΔdegS::degS strain and the WT strain, and a partial recovery of the colonization ability of the ΔdegS+icdh strain. Overall, these findings suggest that the DegS protease regulates the expression of ICDH through ArcA, thereby affecting the NADH and ATP levels of V.cholerae and affecting its growth and intestinal colonization ability.