Cooperative and Independent Functionality of tmRNA and SmpB: A Multifunctional Exploration Beyond Ribosome Rescue

The trans-translation system, mediated by transfer-messenger RNA (tmRNA, encoded by the ssrA gene) and its partner protein SmpB, helps release ribosomes stalled on defective mRNA and targets incomplete protein products for hydrolysis. Knocking out the ssrA and smpB genes in various pathogens leads to different phenotypic changes, indicating that they have both cooperative and independent functionalities. This study aims to clarify the functional relationships between tmRNA and SmpB in Aeromonas veronii, the pathogen that cause threats in aquaculture and human health. We characterized the expression dynamics of the ssrA and smpB genes at different growth stages of the pathogen, assessed the responses of deletion strains ΔssrA and ΔsmpB to various environmental stressors and carbon source supplementations, and identified the gene regulatory networks involving both genes by integrating transcriptomic and phenotypic analyses. Our results showed that the gene ssrA maintains stable expression throughout the bacterial growth period, while smpB exhibits upregulated expression in response to nutrient deficiencies. Compared to the wild type, both ΔssrA and ΔsmpB strains exhibit attenuated resistances to most stress conditions. However, ΔssrA independently responds to starvation, while ΔsmpB specifically shows reduced resistance to higher concentration of Fe³⁺/Na⁺ ions and higher utilization of the carbon source β-methylglucoside. Transcriptomic analysis supports these phenotypic results, demonstrating that tmRNA and SmpB cooperate under nutrient-deficient conditions but operate independently in nutrient-rich environments. Phenotypic experiments confirm that SsrA and SmpB collaboratively regulate genes involved in siderophore synthesis and iron uptake systems in response to extracellular iron deficiency. The findings of the present study provide crucial insights into the functions of the trans-translation system and highlight new roles for tmRNA and SmpB beyond trans-translation.