Deciphering the negative regulatory role of TetR1 in organic solvent tolerance of Rhodococcus ruber through...

TetR family transcriptional regulators (TFRs) represent a major class of regulatory proteins that govern diverse cellular processes, including antibiotic biosynthesis, primary and secondary metabolism, and stress responses. In this study, the role of a specific TFR, TetR1, in organic solvent tolerance of Rhodococcus ruber SD3 (R. ruber SD3) was investigated through functional characterization. The interaction between TetR1 and its target DNA was confirmed via molecular docking and electrophoretic mobility shift assay. Under various organic solvent stresses, the relative expression level of the tetr1 gene was downregulated in R. ruber SD3, indicating that it acted as a negative regulator of organic solvent tolerance. Comparative growth analysis showed that the tetr1 knockdown strain exhibited enhanced growth compared to the wild-type under organic solvent stress, whereas the overexpression strain displayed impaired growth within 72 h. Transcriptomic profiling identified 804 differentially expressed genes between the tetr1 knockdown and wild-type strains, including 328 upregulated and 476 downregulated genes. Transcriptomic analysis further revealed that tetR1-KD triggered global transcriptional reprogramming. This included: upregulating ABC transporters while suppressing MCE-family importers to limit solvent influx; remodeling membrane lipid composition by altering fatty acid metabolism gene expression to enhance stability; and coordinately boosting xenobiotic biodegradation pathways to improve detoxification. These results provided new insights into the biological functions of TetR1 in R. ruber.