Extensive use and disposal of 2,4,6-trinitrotoluene (TNT), a primary constituent of explosives, pollutes the environment and causes severe damage to human health. Complete mineralization of TNT via bacterial degradation has recently gained research interest as an effective method for the restoration of contaminated sites. Here, screening for TNT degradation by six selected bacteria revealed that Buttiauxella sp. S19-1, possesses the strongest degrading ability. Moreover, BuP34O (a gene encoding for protocatechuate 3,4-dioxygenase—P34O, a key enzyme in the β-ketoadipate pathway) was upregulated during TNT degradation. A knockout of BuP34O in S19-1 to generate S-M1 mutant strain caused a marked reduction in TNT degradation efficiency compared to S19-1. Additionally, the EM1 mutant strain (Escherichia coli DH5α transfected with BuP34O) showed higher degradation efficiency than DH5α. Gas chromatography mass spectrometry (GC-MS) analysis of TNT degradation by S19-1 revealed 4-amino-2,6-dinitrotolune (ADNT) as the intermediate metabolite of TNT. Furthermore, the recombinant protein P34O (rP34O) expressed the activity of 2.46 µmol/min·mg. Our findings present the first report on the involvement of P34O in bacterial degradation of TNT and its metabolites, suggesting that P34O could catalyze downstream reactions in the TNT degradation pathway. In addition, the TNT-degrading ability of S19-1, a Gram-negative marine-derived bacterium, presents enormous potential for restoration of TNT-contaminated seas.

2,4,6-Trinitrotoluene (TNT) is an aromatic pollutant that is difficult to be degraded in the natural environment. The screening of efficient degrading bacteria for bioremediation of TNT has received much attention from scholars. In this paper, transcriptome analysis of the efficient degrading bacterium Buttiauxella sp. S19-1 revealed that the monooxygenase gene (BuMO) was significantly up-regulated during TNT degradation. S-ΔMO (absence of BuMO gene in S19-1 mutant) degraded TNT 1.66-fold less efficiently than strain S19-1 (from 71.2% to 42.9%), and E-MO mutant (Escherichia coli BuMO-expressing strain) increased the efficiency of TNT degradation 1.33-fold (from 52.1% to 69.5%) for 9 h at 180 rpm at 27 °C in LB medium with 1.4 µg·mL⁻¹ TNT. We predicted the structure of BuMO and purified recombinant BuMO (rBuMO). Its specific activity was 1.81 µmol·min⁻¹·mg⁻¹ protein at pH 7.5 and 35 °C. The results of gas chromatography mass spectrometry (GC–MS) analysis indicated that 4-amino-2,6-dinitrotoluene (ADNT) is a metabolite of TNT biodegradation. We speculate that MO is involved in catalysis in the bacterial degradation pathway of TNT in TNT-polluted environment.

Dye wastewater pollution, particularly from persistent and toxic polycyclic organic pollutants, such as aniline blue, poses a significant environmental challenge. Aniline blue, a triphenylmethane dye widely used in the textile, leather, paper, and pharmaceutical industries, is notoriously difficult to treat owing to its complex structure and potential for bioaccumulation. In this study, we explored the capacity of Comamonas testosteroni (CT1) to efficiently degrade aniline blue, focusing on the underlying enzymatic mechanisms and degradation pathways. Through prokaryotic transcriptome analysis, we identified a significantly upregulated short-chain dehydrogenase (SDRz) gene (log₂FC = 2.11, p < 0.05) that plays a crucial role in the degradation process. The SDRz enzyme possessed highly conserved motifs and a typical short-chain dehydrogenase structure. Functional validation using an SDRz-knockout strain (CT-ΔSDRz) and an SDRz-expressioning strains (E-SDRz) confirmed that SDRz is essential for aniline blue degradation. The knockout strain CT-ΔSDRz exhibited a 1.27-fold reduction in the degradation efficiency, compared to CT1 strain after 12 h; while the expression strain E-SDRz showed a 1.24-fold increase compared to Escherichia coli DH5α after 12 h. Recombinant SDRz (rSDRz) was successfully produced, showing significant enzymatic activity (1.267 ± 0.04 mmol·L⁻¹·min⁻¹ protein), with kinetic parameters Vmax = 2.870 ± 0.0156 mmol·L⁻¹·min⁻¹ protein and Km = 1.805 ± 0.0128 mM·mL⁻¹. Under optimal conditions, the rSDRz achieved a degradation efficiency of 62.17% for aniline blue. Gas chromatography–mass spectrometry (GC-MS) analysis identified several intermediate metabolites in the degradation pathway, including benzeneacetaldehyde, a, a-diphenyl, 2-amino-4-methylbenzophenone, benzene, 1-dimethylamino-4-phenylmethyl, benzenesulfonic acid, methyl ester, further elucidating the biodegradation mechanism. These findings highlight SDRz as a critical enzyme in the biodegradation of aniline blue, offering valuable insights and a robust theoretical foundation for developing advanced bioremediation strategies to address dye wastewater pollution.

The COP9 signalosome (CSN) is a conserved protein complex, with CSN1 being one of the largest and most important subunits in the COP9 complex. To investigate the N-terminus function of OsCSN1, we edited the N-terminus of OsCSN1 and found that the mutant of OsCSN1 with 102 amino acids missing at the N-terminus showed insensitivity to red light in terms of the embryonic sheath, stem elongation, and main-root elongation. Moreover, the mutant was able to produce, develop, and bear fruit normally. The research results indicate that OsCSN1 is a negative regulator of stem elongation in rice seedlings regulated by red light. Under red-light treatment, OsCSN1 assembles into CSN, which degrades SLR1 through de NEDDylation, affecting PIL11 activity and ultimately inhibiting stem elongation. OsCSN1 also plays an important regulatory role in the inhibition of rice embryonic sheath elongation under red light. By regulating the degradation of SLR1 and PIL14 through the ubiquitin/26S protease pathway, the elongation of the embryonic sheath is ultimately inhibited. OsCSN1 forms a COP9 complex and is modified with RUB/NEDD8 of the E3 ligase of CUL1 to promote the degradation of SLR1 and PIL14, ultimately affecting the elongation of the embryonic sheath. The regulatory domain is located at the N-terminus of CSN1.

The COP9 signalosome (CSN) is a highly conserved multi-subunit protein complex, with CSN1 being its largest and most conserved subunit. The N-terminal function of CSN1 plays a pivotal and intricate role in plant photomorphogenesis and seedling development. Moreover, CSN is essential for far-red light-mediated photomorphogenesis in seedlings, but the function of OsCSN1 in seedling growth and development under far-red light conditions has not been determined. This study investigates the function of OsCSN1 under far-red light through phenotypic analysis of wild type and OsCSN1 mutant seedlings. Additionally, the effect of the N-terminal region of OsCSN1 on rice seedling growth and development was examined. The addition of exogenous hormone gibberellin (GA3) and gibberellin synthesis inhibitor paclobutrazol (PAC) resulted in notable changes in phenotypes and the expression of key proteins, including CUL4 and SLR1. The findings indicate that OsCSN1 functions as a positive regulator of plant height under far-red light and inhibits root elongation. Under far-red light, OsCSN1 integrates into the COP9 complex and regulates the nuclear localization of COP1. Through its interaction with CUL4 in the CULLIN-RING family, OsCSN1 facilitates the ubiquitin-mediated degradation of SLR1, thereby influencing the growth of rice seedlings. The regulatory function of OsCSN1 in seedling growth and development under far-red light predominantly relies on the 32 amino acids of its N-terminal region. The results of this study can provide new ideas for rice breeding and genetic improvement. Based on the study of key regulatory factors such as OsCSN1, new varieties that can make better use of far-red light signals can be cultivated to enhance crop adaptability and productivity.

The key flavonoid biosynthesis-related genes and their molecular features in rice have not been comprehensively and systematically characterized. In this study, we investigated the glumes of OsCSN1 mutants and OsCSN2 mutants and found the changes in the total flavonoid contents of the OsCSN2 mutants to be more pronounced than those of the OsCSN1 mutants and the changes in the anthocyanin contents of the OsCSN1 mutants to be more pronounced than those of the OsCSN2 mutants. In addition, key genes related to flavonoid synthesis, OsCHI, showed a more pronounced up-regulation trend, and the OsDFR gene, which encodes a precursor enzyme for anthocyanin synthesis, showed a clear down-regulation trend. And yeast two-hybrid experiments showed that OsCSN1 and OsCSN2 had the ability to interact with OsCUL4. In summary, OsCSN1 and OsCSN2 may regulate the metabolism of flavonoids in rice through CUL4-based E3 ligase, and the two subunits play different roles, laying a foundation for the study of the mechanism of flavonoid metabolism in monocotyledonous plants.