Autophosphorylation desensitizes phytochrome signal transduction

Plant red/far-red photoreceptor phytochromes are known as autophosphorylating serine/threonine kinases. However, the functional roles of autophosphorylation and kinase activity of phytochromes are largely unknown. We recently reported that the autophosphorylation of phytochrome A (phyA) plays an important role in regulating plant phytochrome signaling by controlling phyA protein stability. Two serine residues in the N-terminal extension (NTE) region were identified as autophosphorylation sites, and phyA mutant proteins with serine-to-alanine mutations were degraded in plants at a significantly slower rate than the wild-type under light conditions, resulting in transgenic plants with hypersensitive light responses. In addition, the autophosphorylation site phyA mutants had normal protein kinase activities. Collectively, our results suggest that phytochrome autophosphorylation provides a mechanism for signal desensitization in phytochrome-mediated light signaling by accelerating the degradation of phytochrome A.Key words: phytochrome, autophosphorylation, phosphorylation, protein kinase, protein degradation, light signaling, signal desensitizationHigher plants continually adapt to their light environments to promote photosynthesis for optimal growth and development. Natural light conditions are monitored by various plant photoreceptors, including red (R)/far-red (FR) photoreceptor phytochromes.^1,2 Phytochromes are dimeric chromoproteins covalently linked to tetrapyrrole chromophore phytochromobilin, and exist as two photo-interconvertible species, red-light absorbing Pr and far-red-light absorbing Pfr forms. Phytochromes are biosynthesized as the Pr form in the dark, and are transformed to the Pfr form upon exposure to red light. This photoactivation of phytochromes induces a highly regulated signaling network for photomorphogenesis in plants.^3,4 Recently, phosphorylation and dephosphorylation have been suggested to play important roles in phytochrome-mediated light signaling;^5,6 for instance, a few phytochrome-associated protein phosphatases have been shown to act as positive regulators of phytochrome signaling.^7–9 However, the functional roles of phytochrome phosphorylation remain to be explored.     

Development of a reference material using methamphetamine abusers' hair samples for the determination of methamphetamine and amphetamine in hair

In the present study, we developed a reference material (RM) using authentic hair samples for the determination of methamphetamine (MA) and its main metabolite, amphetamine (AP) in human hair. MA abusers' hair samples were collected, homogenized and finally bottled. The concentration of each bottle was determined using two extraction methods, agitation with 1% HCl in methanol at 38 degrees C and ultrasonication with methanol/5M HCl (20:1), followed by gas chromatography/mass spectrometry (GC-MS) after derivatization with trifluoroacetic anhydride (TFAA). Both analytical procedures were fully validated and their extraction efficiency was compared. The homogeneity of analytes was evaluated and their property values were determined with their uncertainties. The two methods were acceptable to analyze MA and AP in human hair through the validation and comparative studies using spiked and authentic hair samples as well as NIST SRM 2379 certified reference material. Satisfying homogeneity was reached for MA and AP in the prepared RM. Finally, a human hair RM containing MA and AP is prepared at the level of 7.64+/-1.24 and 0.54+/-0.07 ng/mg, respectively. This material can be useful in forensic laboratories for internal quality control and external quality assurance.     

Gene Manipulation of a Heavy Metal Hyperaccumulator Species Thlaspi caerulescens L. via Agrobacterium-mediated Transformation

Thlaspi caerulescens L. is well known as a Zn/Cd hyperaccumulator. The genetic manipulation of T. caerulescens through transgenic technology can modify plant features for use in phytoremediation. Here, we describe the efficient transformation of T. caerulescens using Agrobacterium tumefaciens strain EHA105 harboring a binary vector pBI121 with the nptII gene as a selectable marker, the gus gene as a reporter and a foreign catalase gene. Based on the optimal concentration of growth regulators, the shoot cluster regeneration system via callus phase provided the basis of the genetic transformation in T. caerulescens. The key variables in transformation were examined, such as co-cultivation period and bacterial suspension density. Optimizing factors for T-DNA delivery resulted in kanamycin-resistant transgenic shoots with transformation efficiency more than 20%, proven by histochemical GUS assay and PCR analysis. Southern analysis of nptII and RT-PCR of catalase gene demonstrated that the foreign genes were integrated in the genome of transformed plantlets. Moreover, the activity of catalase enzyme in transgenic plants was obviously higher than in wild-type plants. This method offers new prospects for the genetic engineering of this important hyperaccumulator species.     

Electroencephalographic (EEG) Measurements of Mindfulness-based Triarchic Body-pathway Relaxation Technique: A Pilot Study

Objective The “Triarchic body-pathway relaxation technique” (TBRT) is a form of ancient Chinese mindfulness-based meditation professed to give rise to positive emotions and a specific state of consciousness in which deep relaxation and internalized attention coexist. The purpose of this study was to examine the EEG pattern generated during the practice of this mindfulness exercise, and compare it to music listening which has been shown to induce positive emotions. Methods Nineteen college students (aged 19–22 years) participated in the study. Each participant listened to both the TBRT and music audiotapes while EEG was recorded. The order of presentation was counterbalanced to avoid order effect. Two EEG indicators were used: (1) alpha asymmetry index, an indicator for left-sided anterior activation, as measure of positive emotions, and (2) frontal midline theta activity, as a measure for internalized attention. Results Increased left-sided activation, a pattern associated with positive emotions, was found during both TBRT exercise and music conditions. However, only TBRT exercise was shown to exhibit greater frontal midline theta power, a pattern associated with internalized attention. Conclusions These results provided evidence to support that the TBRT gives rise to positive emotional experience, accompanied by focused internalized attention.     

Effect of grain-slag media for the treatment of wastewater in a biological aerated filter

Grain-slag was applied as the media of biological aerated filters (BAF). The performance of two lab-scale BAF was monitored for 6 months to compare the effect of grain-slag with haydite as media. Under ammonia nitrogen load rates varying from 0.49 to 1.21 kg NH(3)-N(m(3)d)(-1), the overall NH(3)-N reductions of the BAF supported by grain-slag and haydite averaged 84.30% and 80.87%, respectively. Higher ammonia nitrogen removal in the BAF with grain-slag was attributable to its buffering pH value capacity by stripping calcium carbonate (CaCO(3)). In terms of removing organic matter, turbidity and colourity, the efficiency of the BAF with grain-slag was lower than that with haydite, but more than 78%, 79% and 80% of fed organic matter, turbidity and colourity was still removed, respectively. So it is feasible for grain-slag to be applied as the media of BAF. The results obtained from the research of ammonia nitrogen removal rate versus pH values indicate that ammonia nitrogen removal rates were not distinctly dependent of pH values in the BAF supported by grain-slag. More than 85% of ammonia nitrogen was removed at pH values from 5.2 to 7.8 ranges. Grain-slag can strip CaCO(3) into the wastewater to buffer pH value and maintain optimal nitrification rates.     

Effects of Adaptive Support Ventilation and Synchronized Intermittent Mandatory Ventilation on Peripheral Circulation and Blood Gas Markers of COPD Patients with Respiratory Failure

The objective of the study was to investigate the effects of adaptive support ventilation (ASV) and synchronized intermittent mandatory ventilation (SIMV) on peripheral circulation of chronic obstructive pulmonary disease (COPD) patients with respiratory failure. 86 COPD patients with respiratory failure were recruited in this study. Self-control method was used to compare the effect of ASV and SIMV on the parameters of ventilation machine, heart rate, blood pressure, central venous pressure (CVP), and blood gas markers. When the patients in ASV and SIMV groups were compared, respiratory rate, tidal volume, and peak airway pressure (PIP) showed significant difference. When minute ventilation (MV) was compared, no significant difference was shown. When peripheral circulation parameters were compared, peripheral circulation heart rate, SBP, DBP, and CVP showed significant difference. Compared with SIMV group, PaO₂, pH, and SaO₂ values were remarkably increased (P < 0.01) while no significant difference was found for partial pressure of carbon dioxide (pCO₂) when two groups were compared. In conclusion, when mechanical ventilation was used in COPD patients with respiratory failure, ASV can significantly improve clinical outcomes.     

Cellular mechanisms in intermittent hypoxia-induced cardiac damage in vivo

Obstructive sleep apnea (OSA), characterized by intermittent hypoxia (IH) during sleep, is increasingly recognized as an independent risk factor of cardiovascular diseases. OSA is associated with changes in the levels of circulating oxidative stress/inflammatory markers and dyslipidemia, supporting their mediating roles in cardiovascular pathogenesis. Our aims were to investigate the effect of IH on heart tissue using an IH-exposed rat model and to explore the potential mechanisms involved in the occurrence of cardiac damage. Male Sprague–Dawley rats were exposed to IH and intermittent normoxia as control and sacrificed after 2 or 4 weeks. IH for 4 weeks caused elevation in serum malondialdehyde and cytokine-induced neutrophil chemoattractant-1 and reduction in serum adiponectin levels. In contrast, cardiac oxidative stress and pro-inflammatory markers were suppressed while cardiac adiponectin and cholesterol levels were elevated after IH exposure for 4 weeks. In parallel, there was an increase in apoptosis in the heart of IH-exposed rats, demonstrated by elevations of Bax and cleaved caspase-3 protein and TUNEL staining. Cardiac damage was further evident with decreased arterial vessel and capillary densities, increased cardiac fibrosis, and the loss of troponin I. Our data demonstrated that IH exposure paradoxically caused systemic oxidative and inflammatory responses and cardioprotective responses, i.e., anti-oxidative and anti-inflammatory responses. Despite such a local compensatory protective mechanism, cardiac damage was observed that might be due to IH-induced cholesterol accumulation in the heart and caspase-dependent apoptosis.