Overexpression of the phosphatidylinositol synthase gene from Zea mays in tobacco plants alters the membrane lipids composition and improves drought stress tolerance.

Phosphatidylinositol (PtdIns) is an important lipid because it serves as a key membrane constituent and is the precursor of the inositol-containing lipids that are found in all plants and animals. It is synthesized from cytidine-diphosphodiacylglycerol (CDP-DG) and myo-inositol by PtdIns synthase (PIS). We have previously reported that two putative PIS genes from maize (Zea mays L.), ZmPIS and ZmPIS2, are transcriptionally up-regulated in response to drought (Sui et al., Gene, 426:47-56, 2008). In this work, we report on the characterization of ZmPIS in vitro and in vivo. The ZmPIS gene successfully complemented the yeast pis mutant BY4743, and the determination of PIS activity in the yeast strain further confirmed the enzymatic function of ZmPIS. An ESI-MS/MS-based lipid profiling approach was used to identify and quantify the lipid species in transgenic and wild-type tobacco plants before and after drought treatment. The results show that the overexpression of ZmPIS significantly increases lipid levels in tobacco leaves under drought stress compared to those of wild-type tobacco, which correlated well with the increased drought tolerance of the transgenic plants. Further analysis showed that, under drought stress conditions, ZmPIS overexpressors were found to exhibit increased membrane integrity, thereby enabling the retention of more solutes and water compared with the wild-type and the vector control transgenic lines. Our findings give us new insights into the role of the ZmPIS gene in the response of maize to drought/osmotic stress and the mechanisms by which plants adapt to drought stress.

Enhanced expression of phospholipase C 1 (ZmPLC1) improves drought tolerance in transgenic maize.

Phosphatidylinositol-specific phospholipase C (PI-PLC) plays an important role in a variety of physiological processes in plants, including drought tolerance. It has been reported that the ZmPLC1 gene cloned from maize (Zea mays L.) encoded a PI-PLC and up-regulated the expression in maize roots under dehydration conditions (Zhai SM, Sui ZH, Yang AF, Zhang JR in Biotechnol Lett 27:799-804, 2005). In this paper, transgenic maize expressing ZmPLC1 transgenes in sense or antisense orientation were generated by Agrobacterium-mediated transformation and confirmed by Southern blot analysis. High-level expression of the transgene was confirmed by real-time RT-PCR and PI-PLC activity assay. The tolerance to drought stress (DS) of the homogenous transgenic maize plants was investigated at two developmental stages. The results demonstrated that, under DS conditions, the sense transgenic plants had higher relative water content, better osmotic adjustment, increased photosynthesis rates, lower percentage of ion leakage and less lipid membrane peroxidation, higher grain yield than the WT; whereas those expressing the antisense transgene exhibited inferior characters compared with the WT. It was concluded that enhanced expression of sense ZmPLC1 improved the drought tolerance of maize.

Heterologous expression of vacuolar H(+)-PPase enhances the electrochemical gradient across the vacuolar membrane and improves tobacco cell salt tolerance.

The vacuolar H(+)-translocating inorganic pyrophosphatase (H(+)-PPase) uses pyrophosphate as substrate to generate the proton electrochemical gradient across the vacuolar membrane to acidify vacuoles in plant cells. The heterologous expression of H(+)-PPase genes (TsVP from Thellungiella halophila and AVP1 from Arabidopsis thaliana) improved the salt tolerance of tobacco plants. Under salt stress, the transgenic seedlings showed much better growth and greater fresh weight than wild-type plants, and their protoplasts had a normal appearance and greater vigor. The cytoplasmic and vacuolar pH in transgenic and wild-type cells were measured with a pH-sensitive fluorescence indicator. The results showed that heterologous expression of H(+)-PPase produced an enhanced proton electrochemical gradient across the vacuolar membrane, which accelerated the sequestration of sodium ions into the vacuole. More Na(+) accumulated in the vacuoles of transgenic cells under salt (NaCl) stress, revealed by staining with the fluorescent indicator Sodium Green. It was concluded that the tonoplast-resident H(+)-PPase plays important roles in the maintenance of the proton gradient across the vacuolar membrane and the compartmentation of Na(+) within vacuoles, and heterologous expression of this protein enhanced the electrochemical gradient across the vacuolar membrane, thereby improving the salt tolerance of tobacco cells.

[A clinical analysis of 23 cases of allergic bronchopulmonary aspergillosis].

OBJECTIVE: To describe the clinical characteristics of allergic bronchopulmonary aspergillosis (ABPA). METHODS: The clinical presentations, serologic results, lung function data, chest radiology and the results of treatment of 23 patients with ABPA in Peking Union Medical College Hospital were retrospectively analyzed. RESULTS: There were 11 males and 12 females, with a mean age of (34.0 +/- 13.2) yrs. Tuberculosis, pneumonia, lung cancer and Wegener's granulomatosis were initially diagnosed in 12, 3, 2 and 1 cases respectively. Cough was present in all patients and sputum production in 22 cases, wheeze in 18, sputum plugs in 16, fever in 15 (high degree of fever in 4), hemoptysis in 12, chest pain/backache in 8 and weight loss in 7 cases. Forced expired volume in one second (FEV(1)) was (54.7 +/- 24.1)% predicted, and FEV(1)/forced vital capacity (FVC) was (62.5 +/- 11.9)%. The FEV(1) reversibility was found in 56% (9/16) of the patients. Chest CT were performed in 22 cases. Patchy infiltrations were present in 21, central bronchiectasis in 17, nodular opacities in 9, mucoid impaction (glover-finger/band linear opacities) in 6, consolidation in 5, and mediastinal adenopathy in 11 cases, while 17 cases presented fleeting infiltrations. Twenty-two patients were treated with prednisone plus itraconazole and 17 patients were followed. The median follow up time was 26.2 months (13 days -19 years). Pulmonary infiltrations relapsed in 9 patients and not appeared for more than 6 years in only one patient. CONCLUSIONS: ABPA was mostly misdiagnosed as tuberculosis. Wheeze is present in almost all patients with ABPA, which can be useful in differentiation from tuberculosis. Obstructive ventilatory defect, peripheral blood eosinophilia, fleeting pulmonary infiltration and central bronchiectasis were features of ABPA. Measurement of total IgE, A.fumigatus-specific IgE levels and immediate cutaneous reaction to A.fumigatus are helpful for confirmation of the diagnosis.