Ammonium improved cell wall and cell membrane P reutilization and external P uptake in a putrescine and ethylene dependent pathway.

Ammonium promotes rice P uptake and reutilization better than nitrate, under P starvation conditions; however, the underlying mechanism remains unclear. In this study, ammonium treatment significantly increased putrescine and ethylene content in rice roots under P deficient conditions, by increasing the protein content of ornithine decarboxylase and 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase compared with nitrate treatment. Ammonium treatment increased rice root cell wall P release by increasing pectin content and pectin methyl esterase (PME) activity, increased rice shoot cell membrane P release by decreasing phosphorus-containing lipid components, and maintained internal P homeostasis by increasing OsPT2/6/8 expression compared with nitrate treatment. Ammonium also improved external P uptake by regulating root morphology and increased rice grain yield by increasing the panicle number compared with nitrate treatment. The application of putrescine and ethylene synthesis precursor ACC further improved the above process. Our results demonstrate for the first time that ammonium increases rice P acquisition, reutilization, and homeostasis, and rice grain yield, in a putrescine- and ethylene-dependent manner, better than nitrate, under P starvation conditions.

Unearthing the alleviatory mechanisms of hydrogen sulfide in aluminum toxicity in rice.

Hydrogen sulfide (H2S) improves aluminum (Al) resistance in rice, however, the underlying mechanism remains unclear. In the present study, treatment with 30-µM Al significantly inhibited rice root growth and increased the total Al content, apoplastic and cytoplasm Al concentration in the rice roots. However, pretreatment with NaHS (H2S donor) reversed these negative effects. Pretreatment with NaHS significantly increased energy production under Al toxicity conditions, such as by increasing the content of ATP and nonstructural carbohydrates. In addition, NaHS stimulated the AsA-GSH cycle to decrease the peroxidation damage induced by Al toxicity. Pretreatment with NaHS significantly inhibited ethylene emissions in the rice and then inhibited pectin synthesis and increased the pectin methylation degree to reduce cell wall Al deposition. The phytohormones indole-3-acetic and brassinolide were also involved in the alleviation of Al toxicity by H2S. The transcriptome results further confirmed that H2S alleviates Al toxicity by increasing the pathways relating to material and energy metabolism, redox reactions, cell wall components, and signal transduction. These findings improve our understanding of how H2S affects rice responses to Al toxicity, which will facilitate further studies on crop safety.

Interaction of a Novel Zn2Cys6 Transcription Factor DcGliZ with Promoters in the Gliotoxin Biosynthetic Gene Cluster of the Deep-Sea-Derived Fungus Dichotomomyces cejpii.

Gliotoxin is an important epipolythiodioxopiperazine, which was biosynthesized by the gli gene cluster in Aspergillus genus. However, the regulatory mechanism of gliotoxin biosynthesis remains unclear. In this study, a novel Zn2Cys6 transcription factor DcGliZ that is responsible for the regulation of gliotoxin biosynthesis from the deep-sea-derived fungus Dichotomomyces cejpii was identified. DcGliZ was expressed in Escherichia coli and effectively purified from inclusion bodies by refolding. Using electrophoretic mobility shift assay, we demonstrated that purified DcGliZ can bind to gliG, gliM, and gliN promoter regions in the gli cluster. Furthermore, the binding kinetics and affinity of DcGliZ protein with different promoters were measured by surface plasmon resonance assays, and the results demonstrated the significant interaction of DcGliZ with the gliG, gliM, and gliN promoters. These new findings would lay the foundation for the elucidation of future gliotoxin biosynthetic regulation mechanisms in D. cejpii.

Insulin deficiency induces rat renal mesangial cell dysfunction via activation of IGF-1/IGF-1R pathway.

AIM: Diabetic nephropathy is one of the major complications of diabetes and the major cause of end-stage renal disease. In this study we investigated the insulin deficiency (ID) induced changes in renal mesangial cells (MCs) and in the kidney of STZ-induced diabetic rats. METHODS: Cultured rat renal MCs were incubated in ID media. Cell proliferation was analyzed using BrdU incorporation assay. The expression of insulin receptor (IR), insulin-like growth factor-1 receptor (IGF-1R), phosphorylated IGF-1R, fibronectin, and collagen IV was determined with Western blot analysis. STZ-induced diabetic rats were treated with an IGF-1R antagonist picropodophyllin (PPP, 20 mg·kg(-1)·d(-1), po) for 8 weeks. After the rats were euthanized, plasma and kidneys were collected. IGF-1 levels in renal cortex were measured with RT-PCR or ELISA. The morphological changes in the kidneys were also examined. RESULTS: Incubation in ID media significantly increased cell proliferation, the synthesis of fibronectin and collagen IV, and the expression of IGF-1 and IGF-1R and phosphorylated IGF-1R in renal MCs. Pretreatment of the cells with PPP (50 nmol/L) blocked ID-induced increases in cell proliferation and the synthesis of fibronectin and collagen IV; knockdown of IGF-1R showed a similar effect as PPP did. In contrast, treatment of the cells with IGF-1 (50 ng/mL) exacerbated ID-induced increases in cell proliferation. In the kidneys of diabetic rats, the expression of IGF-1, IGF-1R and phosphorylated IGF-1R were significantly elevated. Treatment of diabetic rats with PPP did not lower the blood glucose levels, but significantly suppressed the expression of TGF-ß, fibronectin and collagen IV in the kidneys, the plasma levels of urinary nitrogen and creatinine, and the urinary protein excretion. CONCLUSION: Insulin deficiency increases the expression of IGF-1 and IGF-1R in renal MCs and the kidney of diabetic rats, which contributes to the development of diabetic nephropathy.

P300-dependent STAT3 acetylation is necessary for angiotensin II-induced pro-fibrotic responses in renal tubular epithelial cells.

AIM: To explore the signal transducer and activator of transcription 3 (STAT3) signaling pathway, especially STAT3 acetylation, in angiotensin II (Ang II)-induced pro-fibrotic responses in renal tubular epithelial cells. METHODS: Rat renal tubular epithelial cell line (NRK-52E) was used. STAT3 acetylation and phosphorylation, as well as the expression of fibronectin, collagen IV and transforming growth factor-ß1 (TGF-ß1) were examined using Western blotting. The level and localization of STAT3 phosphorylation on Tyr705 were detected with fluorescence immunocytochemistry. The cells were transfected with a plasmid vector carrying p300 gene or siRNA targeting p300 to regulate p300 expression. RESULTS: Overexpression of p300 significantly increased STAT3 acetylation on Lys685, STAT3 phosphorylation on Tyr705, and the expression of TGF-ß1, collagen IV and fibronectin in the cells. Treatment of the cells with Ang II (1 µmol/L) significantly increased STAT3 phosphorylation on Tyr705 through JAK2 activation, and dose-dependently increased the expression of fibronectin, collagen IV and TGF-ß1. Pretreatment with curcumin, an inhibitor of JAK2 and p300, blocked Ang II-induced effects. Knockdown of p300 significantly decreased STAT3 acetylation on Lys685, and abolished Ang II-stimulated STAT3 phosphorylation on Tyr705, whereas pretreatment of the cells with C646, a selective inhibitor of p300, inhibited Ang II-induced STAT3 nuclear translocation and the expression of TGF-ß1, collagen IV and fibronectin. Pretreatment of the cells with AG490, a JAK2 inhibitor, markedly inhibited Ang II-induced STAT3 phosphorylation on Tyr705 and fibronectin expression. CONCLUSION: p300-dependent STAT3 acetylation is necessary for Ang II-induced STAT3 phosphorylation and the consequent pro-fibrotic responses in renal tubular epithelial cells in vitro.

Inhibition of STAT3 acetylation is associated with angiotesin renal fibrosis in the obstructed kidney.

AIM: To explore the relationship between the signal transducer and activator of transcription 3 (STAT3) signaling and renal fibrosis. METHODS: Rat renal tubular epithelial NRK-52E cells were treated with angiotesin II (Ang II), nicotinamide (an inhibitor of NAD+-dependent class III protein deacetylases, SIRT1-7), or resveratrol (an activator of SIRT1). Mice underwent unilateral ureteral obstruction (UUO) were used for in vivo studies. Renal interstitial fibrosis was observed with HE and Masson's trichrome staining. STAT3 acetylation and phosphorylation, fibronectin, collagen I, collagen IV, and α-smooth muscle actin (α-SMA) levels were examined using Western blotting. RESULTS: Nicotinamide (0.625-10 mmol/L) dose-dependently increased STAT3 acetylation on Lys685 and phosphorylation on Tyr705 in NRK-52E cells, accompanied by accumulation of fibronectin and collagen IV. Ang II increased STAT3 phosphorylation on Tyr705 and the expression of fibronectin, collagen IV and α-SMA in the cells. Pretreatment with resveratrol (12.5 µmol/L) blocked Ang II-induced effects in the cells. UUO induced marked STAT3 phosphorylation, fibronectin, collagen IV and α-SMA accumulation, and renal interstitial fibrosis in the obstructed kidneys, which were significantly attenuated by daily administration of resveratrol (100 mg/kg). CONCLUSION: STAT3 acetylation plays an important role in activation of STAT3 signaling pathway and consequent renal fibrosis.