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Nitric oxide (NO) is a key player in numerous physiological processes. Excessive NO induces DNA damage, but how plants respond to this damage remains unclear. We screened and identified an Arabidopsis NO hypersensitive mutant and found it to be allelic to TEBICHI/POLQ, encoding DNA polymerase θ. The teb mutant plants were preferentially sensitive to NO- and its derivative peroxynitrite-induced DNA damage and subsequent double-strand breaks (DSBs). Inactivation of TEB caused the accumulation of spontaneous DSBs largely attributed to endogenous NO and was synergistic to DSB repair pathway mutations with respect to growth. These effects were manifested in the presence of NO-inducing agents and relieved by NO scavengers. NO induced G2/M cell cycle arrest in the teb mutant, indicative of stalled replication forks. Genetic analyses indicate that Polθ is required for translesion DNA synthesis across NO-induced lesions, but not oxidation-induced lesions. Whole-genome sequencing revealed that Polθ bypasses NO-induced base adducts in an error-free manner and generates mutations characteristic of Polθ-mediated end joining. Our experimental data collectively suggests that Polθ plays dual roles in protecting plants from NO-induced DNA damage. Since Polθ is conserved in higher eukaryotes, mammalian Polθ may also be required for balancing NO physiological signaling and genotoxicity.

Objective To investigate the effect of baicalin on acute lung injury (ALI) induced by lipopolysaccharide (LPS), and to explore the roles of M1/M2 polarization of pulmonary macrophages and M1/M2 macrophage-mediated inflammation in the effect. Methods Sixty SD rats were divided into six groups: normal control group, LPS group, (10, 50, 100) mg/kg baicalin combined with LPS group, and dexamethasone (DEX) group. ALI models were established by intratracheal instillation of LPS. After 24 hours, bronchoalveolar lavage fluid (BALF) and bilateral lung tissues were collected. The pathological changes of rat lung tissue were observed by HE staining, and the wet/dry mass ratio (W/D) of lung tissue was measured; the contents of IL-1β, IL-6, TNF-α, and IL-10 in BALF were detected by ELISA; the M1 macrophage marker inducible nitric oxide synthase (iNOS) and the M2 macrophage marker CD206 in CD68 positive macrophages were detected by immunofluorescence cytochemical staining; the mRNA expressions of iNOS, IL-1β, Arg1, and CD206 were detected by real-time PCR, and the protein expressions of iNOS and Arg1 were detected by Western blot analysis. Results Baicalin significantly reduced lung lesions and lung water content in ALI rats, and down-regulated the secretion levels of pro-inflammatory cytokines TNF-α, IL-6, and IL-1β, while up-regulated the secretion of anti-inflammatory cytokine IL-10 in BALF. Baicalin significantly inhibited the lung macrophage polarization to M1 phenotype, and promoted the polarization to M2 phenotype. Baicalin significantly decreased the mRNA and protein expression levels of IL-1β and iNOS, while increased the mRNA and protein expression levels of CD206 and Arg1 in lung tissues. Conclusion Baicalin can inhibit the lung macrophage polarization to M1 phenotype, promote the polarization to M2 phenotype, and reduce the M1/M2 ratio, thereby alleviating the LPS-induced pulmonary inflammatory response in ALI rats.

Nitric oxide (NO), an active signaling molecule in plants, is involved in numerous physiological processes and adaptive responses to environmental stresses. Under high-salt conditions, plants accumulate NO quickly, and reorganize Na(+) and K(+) contents. However, the molecular connection between NO and ion homeostasis is largely unknown. Here, we report that NO lowers K(+) channel AKT1-mediated plant K(+) uptake by modulating vitamin B6 biosynthesis. In a screen for Arabidopsis NO-hypersensitive mutants, we isolated sno1 (sensitive to nitric oxide 1), which is allelic to the previously noted mutant sos4 (salt overly sensitive 4) that has impaired Na(+) and K(+) contents and overproduces pyridoxal 5'-phosphate (PLP), an active form of vitamin B6. We showed that NO increased PLP and decreased K(+) levels in plant. NO induced SNO1 gene expression and enzyme activity, indicating that NO-triggered PLP accumulation mainly occurs through SNO1-mediated vitamin B6 salvage biosynthetic pathway. Furthermore, we demonstrated that PLP significantly repressed the activity of K(+) channel AKT1 in the Xenopus oocyte system and Arabidopsis root protoplasts. Together, our results suggest that NO decreases K(+) absorption by promoting the synthesis of vitamin B6 PLP, which further represses the activity of K(+) channel AKT1 in Arabidopsis. These findings reveal a previously unidentified pivotal role of NO in modulating the homeostasis of vitamin B6 and potassium nutrition in plants, and shed light on the mechanism of NO in plant acclimation to environmental changes.

Heavy metals in the environment are harmful limiting factors for the normal growth and development of plants. Here, we isolated and identified an Arabidopsis thaliana T-DNA insertion mutant, named sro1-1, which showed a hyper-sensitive response to HgCl2. The SRO1 protein contains a WWE domain that mediates protein-protein interactions. Under HgCl2 treatment, when compared with the wild-type plants, the growth of sro1-1 was repressed dramatically and the number of true leaves was reduced and etiolated. The electrolyte leakage rates showed that cell membrane integrity in sro1-1 was damaged more severely than in the wild type. DAB (3,5-diaminobenzidine) staining and confocal microscopy showed that Hg2+ stress induced more hydrogen peroxide accumulation in sro1-1 than in the wild type. The qRT-PCR results indicated that the expression of some abiotic stress-induced genes, such as L-ascorbate peroxidase (APX1), was reduced under oxidative or Hg2+ stress. Transgenic plants containing a GFP::SRO1 fusion protein showed that SRO1 was localized in the nucleus of the cells. SRO1 was shown to be expressed in various tissues, and was most highly expressed in the vigorous tissues. Our results suggest that SRO1 may play an important role in the stress response of A. thaliana to heavy metals.

Low temperature restrains plant growth by inhibiting the cell cycle, and phytohormones play important roles in this case; however, the molecular mechanisms whereby phytohormones affect growth at low temperature are largely unknown. When grown at 23, 16, 10, and 4°C, we found that Arabidopsis thaliana could develop with normal morphology, but needed a prolonged period of cultivation. By screening mutants, we could implicate cytokinin and salicylic acid. At 4°C, both amp1 plants, which have an increased level of cytokinin, and wild-type plants treated with exogenous cytokinin, displayed relative growth rates greater than control by increasing total cell number. Additionally, transgenic NahG plants, which have lower salicylic acid content, grew faster than wild-type accompanied by larger cells. Expression of C-repeat binding transcription factors (CBFs), that mediate cold acclimation by stimulation of the expression of cold-inducible genes, was similar in all tested genotypes. Thus CBF expression did not correlate with the observed enhanced growth in mutants. The improved growth coincided with elevated expression of CYCD3;1, especially in NahG plants. At 4°C, enhanced endoreduplication took responsibility for larger cells in NahG plants, while enhanced cell division was observed in amp1 plants.